3192 ENDO12L_M36A MEET-THE-PROFESSOR: CLINICAL - Clinical Management of Adrenocortical Carcinoma (1:00 PM - 1:45 PM) Clinical Management of Adrenocortical Carcinoma Martin Fassnacht University of Wuerzburg, Wuerzburg, Germany Nothing to Disclose: MF 2012-06-25T13:00:00 Room 320 2012-06-25T00:00:00 1899-12-30T13:00:00 6137 285 1945 M36 A2A Monday 258 2012


3193 ENDO12L_M38A MEET-THE-PROFESSOR: CLINICAL - Evaluation of Premature Adrenarche (1:00 PM - 1:45 PM) Evaluation of Premature Adrenarche Sharon E Oberfield Columbia University Medical Center, New York, NY Nothing to Disclose: SEO 2012-06-25T13:00:00 Room 310 2012-06-25T00:00:00 1899-12-30T13:00:00 6155 286 1946 M38 PED2A Monday 259 2012


3194 ENDO12L_M39A MEET-THE-PROFESSOR: CLINICAL - Pubertal Disruption in Chronic Inflammatory Disease (1:00 PM - 1:45 PM) Pubertal Disruption in Chronic Inflammatory Disease Mark Daniel DeBoer University of Virginia, Charlottesville, VA Disclosure Incomplete: MDD 2012-06-25T13:00:00 Room 360 2012-06-25T00:00:00 1899-12-30T13:00:00 6156 287 1947 M39 PED7A Monday 260 2012


3195 ENDO12L_M40A MEET-THE-PROFESSOR: CLINICAL - Specialized Lipid Testing: The Pre-Treatment Lipoprotein Size Consequences of Adiposopathy ([quot]Sick Fat[quot]) [amp] the Clinical Pitfalls of Post-Treatment Lipoprotein Particle Size Analyses (1:00 PM - 1:45 PM) Specialized Lipid Testing: The Pre-Treatment Lipoprotein Size Consequences of Adiposopathy ([quot]Sick Fat[quot]) [amp] the Clinical Pitfalls of Post-Treatment Lipoprotein Particle Size Analyses Harold Edward Bays L-MARC Research Center, Louisville, KY Disclosures: HEB: Advisory Group Member, Merck & Co.; Speaker, Merck & Co.; Researcher, Merck & Co., Orexigen, Vivus USA, Arena, Novo Nordisk. 2012-06-25T13:00:00 Room 371 2012-06-25T00:00:00 1899-12-30T13:00:00 6222 288 1948 M40 L1A Monday 261 2012


3196 ENDO12L_M41A MEET-THE-PROFESSOR: CLINICAL - What To Do If My Country Doesn[apos]t Have a FRAX? (1:00 PM - 1:45 PM) What To Do If My Country Doesn[apos]t Have a FRAX? Michael R McClung Oregon Osteoporosis Center, Portland, OR Nothing to Disclose: MRM 2012-06-25T13:00:00 Room 361 ABDE 2012-06-25T00:00:00 1899-12-30T13:00:00 6261 289 1949 M41 B11A Monday 262 2012


3197 ENDO12L_CMF9-1A CASE MANAGEMENT FORUM: CLINICAL - Management of Apparent Secondary Hypogonadism Due to Obesity, Opiates [amp] Anabolic Steroids (5:30 PM - 6:15 PM) Management of Apparent Secondary Hypogonadism Due to Obesity, Opiates [amp] Anabolic Steroids Bradley David Anawalt University of Washington, Seattle, WA Disclosure Incomplete: BDA 2012-06-25T17:30:00 Theater A 2012-06-25T00:00:00 1899-12-30T17:30:00 6169 343 2654 CMF9-1 CMF-MR2A Monday 308 2012


3198 ENDO12L_CMF9-2A CASE MANAGEMENT FORUM: CLINICAL - Management of Apparent Secondary Hypogonadism Due to Obesity, Opiates [amp] Anabolic Steroids (5:30 PM - 6:15 PM) Management of Apparent Secondary Hypogonadism due to Obesity, Opiates [amp] Anabolic Steroids Richard Arthur Bebb University of British Columbia, Vancouver, Canada Nothing to Disclose: RAB 2012-06-25T17:30:00 Theater A 2012-06-25T00:00:00 1899-12-30T17:30:00 6182 343 2655 CMF9-2 CMF-MR2A Monday 309 2012


3199 ENDO12L_M42A MEET-THE-PROFESSOR: CLINICAL - Assessing Readiness, Behavioral Modification [amp] Obesity (5:30 PM - 6:15 PM) Assessing Readiness, Behavioral Modification [amp] Obesity Daniel Holland Bessesen University of Colorado School of Medicine, Denver, CO Disclosure Incomplete: DHB 2012-06-25T17:30:00 Room 320 2012-06-25T00:00:00 1899-12-30T17:30:00 6164 344 2656 M42 OB1A Monday 310 2012


3200 ENDO12L_M43A MEET-THE-PROFESSOR: CLINICAL - Common Pitfalls in the Evaluation of Patients with Adrenal Incidentalomas (5:30 PM - 6:15 PM) Common Pitfalls in the Evaluation of Patients with Adrenal Incidentalomas Amir Hekmat Hamrahian Cleveland Clinic Foundation, Cleveland, OH The widespread use of abdominal imaging has resulted in an increased frequency of detected adrenal lesions. An adrenal incidentaloma is an adrenal mass, 1 cm or more in diameter, that is discovered serendipitously by radiological examination in the absence of symptoms or clinical findings suggestive of adrenal disease. When clinicians confront an adrenal incidentaloma, evaluation must address 2 issues: is the lesion functional or is it malignant and if so, is it primary or metastatic. In spite of the fact that the majority of adrenal incidentalomas are benign and non-functional, careful evaluation of all adrenal incidentalomas is warranted to ensure that tumors, such as primary adrenocortical carcinoma (PAC) and pheochromocytoma, that can be associated with significant morbidity and mortality, are not missed.[br]The adrenal computed tomographic (CT) is the cornerstone of imaging studies for adrenal tumors, providing information about size, density, presence of calcifications, areas of necrosis and extent of local invasion of the neoplasm. The three most important imaging criteria are: 1) size of the lesion, 2) the unenhanced CT attenuation value, and 3) the percentage washout. Several studies have shown that the non-contrast CT attenuation is superior to adrenal size in differentiating between benign and malignant adrenal tumors. Appropriate image interpretation can be a guide to a rational diagnostic workup and management.[br]There continues to be disagreement among experts on the choice and the extent of biochemical evaluation in patients with adrenal incidentalomas. All patients with an adrenal incidentaloma should be evaluated for autonomous cortisol secretion, termed subclinical Cushing[apos]s syndrome, (SCS), pheochromocytoma and, if hypertensive, primary aldosteronism. There is significant inconsistency among criteria used in the literature for the diagnosis of SCS. In spite of the lack of a single gold standard test to diagnose SCS, the 1-mg DST is currently considered the best diagnostic tool for evaluation of patients suspected to have SCS. Measurement of plasma metanephrines or 24 hour urine metanephrines are appropriate screening tests for pheochromocytoma. While measurement of plasma aldosterone to renin ratio (ARR) is considered as the best initial screening test for the evaluation of primary aldosteronism, there is a lack of consensus about what ARR cut-off should trigger further evaluation.[br][br]Nothing to Disclose: AHH 2012-06-25T17:30:00 Room 332 2012-06-25T00:00:00 1899-12-30T17:30:00 2459 345 2657 M43 A3A Monday 311 2012


3201 ENDO12L_M44A MEET-THE-PROFESSOR: CLINICAL - Management of Early Stage Cancer/Need for RAI (5:30 PM - 6:15 PM) Management of Early Stage Cancer/Need for RAI Bryan McIver Mayo Clinic, Rochester, MN Session supported by: Genzyme Corporation[br][br]Nothing to Disclose: BM 2012-06-25T17:30:00 Room 310 2012-06-25T00:00:00 1899-12-30T17:30:00 6160 346 2658 M44 TH3A Monday 312 2012


3202 ENDO12L_M45A MEET-THE-PROFESSOR: CLINICAL - Management of Gender Variant Youth (5:30 PM - 6:15 PM) Management of Gender Variant Youth Norman P Spack Children[apos]s Hospital Boston, Boston, MA Nothing to Disclose: NPS 2012-06-25T17:30:00 Room 370 2012-06-25T00:00:00 1899-12-30T17:30:00 6184 347 2659 M45 PED4A Monday 313 2012


3203 ENDO12L_M46A MEET-THE-PROFESSOR: CLINICAL - Minimally Invasive Surgery for Hyperparathyroidism (5:30 PM - 6:15 PM) Minimally Invasive Surgery for Hyperparathyroidism Robert Udelsman Yale University School of Medicine, New Haven, CT Nothing to Disclose: RU 2012-06-25T17:30:00 Room 361 ABDE 2012-06-25T00:00:00 1899-12-30T17:30:00 6208 348 2660 M46 B6A Monday 314 2012


3204 ENDO12L_M48A MEET-THE-PROFESSOR: CLINICAL - Update in FGF 23 (5:30 PM - 6:15 PM) Update in FGF 23 Michael John Econs Indiana University Medical Center, Indianapolis, IN Disclosures: MJE: Indiana University has a patent, but no clinical products at this time, Kirin Brewery; Consultant, Kirin Brewery. 2012-06-25T17:30:00 Room 352 DEF 2012-06-25T00:00:00 1899-12-30T17:30:00 6147 349 2661 M48 B10A Monday 315 2012


3205 ENDO12L_CMF10-1A CASE MANAGEMENT FORUM: CLINICAL - Management of Pediatric Thyroid Cancer (1:00 PM - 1:45 PM) Management of Pediatric Thyroid Cancer Catherine Anne Dinauer Yale University School of Medicine, Guilford, CT Session supported by: Genzyme Corporation[br][br]Nothing to Disclose: CAD 2012-06-26T13:00:00 Theater B 2012-06-26T00:00:00 1899-12-30T13:00:00 6185 389 2783 CMF10-1 CMF-TH2A Tuesday 367 2012


3206 ENDO12L_CMF10-2A CASE MANAGEMENT FORUM: CLINICAL - Management of Pediatric Thyroid Cancer (1:00 PM - 1:45 PM) Management of Pediatric Thyroid Cancer Gary Lee Francis Medical College of Virginia, Richmond, VA Session supported by: Genzyme Corporation[br][br]Nothing to Disclose: GLF 2012-06-26T13:00:00 Theater B 2012-06-26T00:00:00 1899-12-30T13:00:00 6186 389 2784 CMF10-2 CMF-TH2A Tuesday 368 2012


3207 ENDO12L_M49A MEET-THE-PROFESSOR: CLINICAL - Acromegaly (1:00 PM - 1:45 PM) Acromegaly Andrea Lynn Utz Vanderbilt University, Nashville, TN Session supported by: Ipsen Biopharmaceuticals, Inc.[br][br]Disclosures: ALU: Advisory Group Member, Ipsen. 2012-06-26T13:00:00 Room 310 2012-06-26T00:00:00 1899-12-30T13:00:00 6165 390 2785 M49 PIT4A Tuesday 369 2012


2996 ENDO12L_S37-2 SYMPOSIUM SESSION: BASIC - Endocrine Regulation of Lipid [amp] Carbohydrate Metabolism, with Consequences for Longevity: Insights from Animal Models (9:30 AM - 11:00 AM) Regulation of Adult Stem Cell Homeostasis by Metabolic Pathways Anne Brunet Stanford University, Palo Alto, CA Disclosure Incomplete: AB 2012-06-25T10:00:00 Room 342 ABDE 2012-06-25T00:00:00 1899-12-30T10:00:00 6063 261 1852 S37-2 T06-S01 Monday 277 2012


2997 ENDO12L_S37-3 SYMPOSIUM SESSION: BASIC - Endocrine Regulation of Lipid [amp] Carbohydrate Metabolism, with Consequences for Longevity: Insights from Animal Models (9:30 AM - 11:00 AM) Lifespan Extension in the Growth Hormone Receptor Disrupted Mouse John Joseph Kopchick Ohio University/Edison Biotechnical Institute, Athens, OH The growth hormone receptor gene disrupted (GHR-/-) mouse, a.k.a. [apos]The Laron Mouse[apos], was generated in our laboratory nearly two decades ago by targeting exon 4 of the GHR/BP gene. In doing so, both the GHR and GHBP encoded proteins were eliminated. These dwarf mice are insensitive (or resistant) to the action of GH with severely depressed levels of serum IGF-I and elevated levels of GH. GHR-/- mice are fertile; however, at times, breeding is difficult. They also are hypoinsulinemic and insulin sensitive and transition from hypo- to normoglycemic with age. Remarkably, these mice are obese yet long-lived. In terms of obesity, the GHR-/- mice have increased relative fat mass throughout life as compared to littermate controls with a preferential enlargement of the subcutaneous depot. Serum adipokine changes have been noted in these mice with both total adiponectin and high molecular weight (HMW) adiponectin increased. Interestingly, the ratio of the more biologically active HMW adiponectin to total adiponectin is increased in the GHR-/- mice relative to controls. This ratio may be important in the insulin sensitive state found in the mice. Significantly, the GHR-/- mouse mimics many phenotypic aspects of Laron Syndrome patients including depressed incidences of cancer and diabetes. Thus, the use of these mice as a research tool has been widespread and has increased our knowledge of the role of GH in growth, reproduction, longevity, obesity, cancer, insulin and glucose metabolism, the GH/IGF-I feedback loops, and in GH induced intracellular signaling. A review of results derived from these mice along with recent data will be presented.[br][br]The author would like to acknowledge additional authors on this abstract: Darlene Berryman and Ellen Lubbers, Edison Biotechnology Institute, Ohio University, Athens, Ohio.[br][br]Sources of Research Support: This work was supported in part by the State of Ohio[apos]s Eminent Scholar Program that includes a gift by Milton and Lawrence Goll; NIH grant P01AG031736-01A1; and by the AMVETS.[br][br]Nothing to Disclose: JJK 2012-06-25T10:30:00 Room 342 ABDE 2012-06-25T00:00:00 1899-12-30T10:30:00 2469 261 1853 S37-3 T06-S01 Monday 278 2012


2998 ENDO12L_S38-1 SYMPOSIUM SESSION: TRANSLATIONAL - New Insights in the Regulation of Puberty (9:30 AM - 11:00 AM) Lin 28A Transgenic Mice Models: The Human Phenotype Hao Zhu Children[apos]s Hospital Boston and Dana Farber Cancer Institute, Boston, MA Nothing to Disclose: HZ 2012-06-25T09:30:00 Room 362 2012-06-25T00:00:00 1899-12-30T09:30:00 6079 262 1854 S38-1 T07-S06 Monday 280 2012


2999 ENDO12L_S38-2 SYMPOSIUM SESSION: TRANSLATIONAL - New Insights in the Regulation of Puberty (9:30 AM - 11:00 AM) How Many Kisses Are Required To Change a Frog into a Princess? Robert A Steiner University of Washington, Seattle, WA Kisspeptin (a product of the [italic] Kiss1 [/italic] gene) and its receptor (GPR54 or Kiss1r) have emerged as key players in the regulation of reproduction. Mutations in humans and genetically targeted deletions in mice of either [italic] Kiss1 [/italic] or [italic] Kiss1r [/italic] silence the reproductive system, and early developmental expression of Kiss1 or activation of Kiss1r triggers precocious puberty. These observations suggest that kisspeptin signaling is a prerequisite for triggering the onset of puberty. However, recent evidence challenges this notion[mdash] reflecting the discovery that congenital ablation of virtually all ([gt] 97%) of kisspeptin neurons fails to disrupt normal puberty and fertility in mice. Although a few steely-eyed skeptics may question the precise role for kisspeptin in reproduction, others retort that just a few Kiss1-expressing neurons (and the most miniscule amount of Kiss1) may be sufficient to drive puberty and support reproduction. To distinguish between these interpretations and to determine whether males and females differ in their requirement for kisspeptin signaling to achieve reproductive success, we produced a mouse model that has a disruption in Kiss1 transcription ([italic] Kiss1 [sup] Cre/Cre [/sup] [/italic]) and markedly reduced expression of [italic] Kiss1 [/italic]. We found that male [italic] Kiss1 [sup] Cre/Cre [/sup] [/italic] mice exhibit a significant delay in the age of puberty onset and have lower testicular weights than wild type (WT) controls[mdash] however, these same [italic] Kiss1 [sup] Cre/Cre [/sup] [/italic] males can inseminate WT females, which produce normal-sized litters, and retain the ability to show a postcastration rise in gonadotropin secretion. In contrast, female [italic] Kiss1 [sup] Cre/Cre [/sup] [/italic] mice enter puberty at the same age as WT controls but exhibit a dramatically reduced incidence of ovulation and have markedly reduced fertility compared to WTs. Thus, we conclude that either a vanishingly small degree of Kiss1 expression is sufficient to support reproduction or that compensatory mechanisms help to rescue the reproductive phenotype of [italic] Kiss1 [sup] Cre/Cre [/sup] [/italic] mice; moreover, the absolute requirement for Kiss1 to initiate puberty and sustain reproduction in the adult differs between the sexes.[br][br]Sources of Research Support: NIH/NICHD 2 R01 HD049651 to RAS.[br][br]Nothing to Disclose: RAS 2012-06-25T10:00:00 Room 362 2012-06-25T00:00:00 1899-12-30T10:00:00 2532 262 1855 S38-2 T07-S06 Monday 281 2012


3000 ENDO12L_S38-3 SYMPOSIUM SESSION: TRANSLATIONAL - New Insights in the Regulation of Puberty (9:30 AM - 11:00 AM) Genes Implicated in GnRH Secretion Ursula B Kaiser Brigham and Women[apos]s Hospital/Harvard Medical School, Boston, MA The mechanisms controlling the timing and onset of puberty remain largely unknown but recent insights into the genetic causes of pubertal disorders have provided important advances in this field. Mutations in genes important for the development and function of hypothalamic-pituitary pathways controlling GnRH release and LH and FSH secretion have been identified in patients with normosmic hypogonadotropic hypogonadism, Kallmann syndrome, and central precocious puberty. Many of the genes implicated encode G protein-coupled receptors and their ligands, including: (1) [italic]KISS1/KISS1R,[/italic] encoding kisspeptin and its receptor (KISS1R), (2) [italic]TAC3/TACR3[/italic], encoding neurokinin B and the neurokinin 3 receptor (NK3R), (3) [italic]PROK2/PROKR2[/italic], encoding prokineticin 2 and prokineticin receptor 2 (PROKR2), and (4) [italic]GNRH1/GNRHR[/italic], encoding GnRH itself and its receptor, GnRHR. Mutations in these GPCRs, located in diverse functional domains of the receptors, have been described in both heterozygous and homozygous states in patients with varying degrees of GnRH dysregulation. Elucidating the structure-function relationships for these GPCRs and the key mechanisms by which the activation of these receptors by their ligands mediate cellular and biological responses have become increasingly important as reproductive abnormalities are attributed to mutations in these genes. The identification of the domains of these receptors important for cell surface expression, ligand binding, and activation of cellular signaling pathways will advance our understanding of the function of these receptors in the control of GnRH release.[br][br]Nothing to Disclose: UBK 2012-06-25T10:30:00 Room 362 2012-06-25T00:00:00 1899-12-30T10:30:00 2549 262 1856 S38-3 T07-S06 Monday 282 2012


3001 ENDO12L_S39-1 SYMPOSIUM SESSION: CLINICAL - New Insights into Androgen Replacement Therapy (9:30 AM - 11:00 AM) Does Androgen Replacement Drive Prostate Disease? Stephanie T Page University of Washington Medical Center, Seattle, WA Androgen replacement in men with low serum testosterone concentrations can improve symptoms of hypogonadism, such as low libido and reduced sexual function, and has positive effects on body composition and strength. However, concern exists that increases in circulating testosterone concentrations might increase the risk for prostate disease, such as benign prostatic hyperplasia or prostate cancer. Advanced age is the primary, identified risk factor for both diseases of the prostate and hypogonadism and it is likely that these conditions co-exist in significant numbers of men. Unfortunately, randomized-controlled trials appropriately powered to address the relationship, if any, between prostate disease risk and androgen replacement have not been performed. Therefore, in this session we will review the available data addressing separate and important issues regarding testosterone replacement, prostate biology and disease risk.[br]First, do exogenous androgens, acting as a growth factor, drive the normal prostate towards a proliferative or de-differentiated state that might progress to prostate cancer? Androgen replacement can increase prostate size in men with long-standing androgen deficiency. However, recent data suggest that raising serum androgen levels does not increase intraprostatic androgen concentrations in parallel, suggesting that there might be few biologic consequences within the prostate epithelium when serum androgen levels are increased (within the normal range).[br]Second, does androgen replacement in men who have been treated for prostate cancer increase their risk for recurrence? Androgen withdrawal is standard therapy for prostate cancer, and initial responsiveness is essentially universal, suggesting a pivotal role for androgens in maintaining prostate carcinoma once it has developed. Recent reports have challenged the notion that androgens stimulate cancer, citing small series of men who have undergone radical prostatectomy for prostate cancer, developed hypogonadism, and been treated with testosterone without biochemical recurrence. Men with longstanding biochemical remission prior to the initiation of testosterone replacement appear to be less likely to experience increases in PSA. However, these reports must be treated with extreme caution, as controlled clinical trials in this area are lacking. Thus, the true risks and benefits of androgen replacement in men who have been treated for prostate cancer remain largely unknown.[br][br]Disclosures: STP: Principal Investigator, Abbott Laboratories. 2012-06-25T09:30:00 Theater B 2012-06-25T00:00:00 1899-12-30T09:30:00 2489 263 1857 S39-1 T05-S05 Monday 284 2012


3002 ENDO12L_S39-2 SYMPOSIUM SESSION: CLINICAL - New Insights into Androgen Replacement Therapy (9:30 AM - 11:00 AM) When Is Testosterone an Appropriate Treatment for Erectile Dysfunction? Jacques Buvat CETPARP, Lille, France Erectile dysfunction (ED) is prevalent in young hypogonadal men. Some randomized, placebo-controlled trials (RCPTs) demonstrated that testosterone therapy (Tth) normalizes sexual function in such men. This has led to the belief that hypogonadism is an easily curable cause of ED. Serum testosterone (T) is [lt] 10.4 nmol/L in 15% of middle-aged and older men presenting with ED. Observational studies of the effects of Tth on erectile function of such men show limited success rate ([lt] 40%). Several meta-analyses of the RPCTs of Tth also analyzed the effects of this therapy on erectile function of men of any age. A first one, not restricted to men with sexual complaints, concluded to an improvement of erections in men with baseline T [lt] 12 nmol/L. Two further meta-analyses, restricted to men with ED or desire problems, did not confirm the benefit of Tth for erectile function, except in young hypogonadal men. Several reasons may account for such differences in the effects of Tth on erectile function of young and older hypogonadal men with ED. Above all vascular comorbidities are prevalent in older men with ED and late onset hypogonadism (LOH).These may prevent improvement of erections with Tth alone, and may require combination therapy with T and phosphodiesterase type V inhibitor (PDE5I) for optimal results. Moreover ED-associated LOH is often mild or moderate, and may not be the real cause of ED, since 2 population-based studies found that in older men low T is associated with ED, but only below a threshold of 8 nmol/L. In certain cases, the low T associated with ED might be a consequence of reduced sexual activity, as suggested by an increase in T following improvement of erectile function after non-hormonal treatment of ED. Lastly, evidence of requirement of a minimum T level for a complete efficacy of PDE5I therapy recently emerged. The corresponding thresholds seem close to 10.4 nmol/L for total T, 2.1 nmol/L for bioavailable T, and 180 pml/L for calculated Free T. In conclusion, the lower the patient[apos]s age and T level, the higher the chance of success of Tth alone in a hypogonadal patient with ED. Low T is probably a plausible cause of ED only below 8 nmol/L, it but may reduce sexual desire at less decreased levels. After the age of 50, the frequent association of endothelial and vascular factors often requires PDE5I therapy, that should be combined with Tth in case of T [lt] 10.4 nmol/L, due to the need for a minimum T level for a full PDE5I efficacy.[br][br](1) Buvat J, Maggi M, Gooren L, Guay A, Kaufman J, Morgentaler A, Schulman C, Tan H.M, Torres L.O, Yassin A, Zitzmann M. Endocrine Aspects of Male Sexual Dysfunctions. J Sex Med 2010;7:1627-1656. (2) Buvat J, Montorsi F, Maggi M, Porst H, Kaipia A, Colson MH, Cuzin B, Moncada I, Martin-Morales A, Yassin A, Meuleman E, Eardley I, Dean JD, Shabsigh R. Hypogonadal men nonresponders to the PDE5 inhibitor tadalafil benefit from normalization of testosterone levels with a 1% hydroalcoholic testosterone gel in the treatment of erectile dysfunction (TADTEST study). J Sex Med 2011;8:284-293.[br][br]Sources of Research Support: Grant of Bayer Schering Pharma.[br][br]Disclosures: JB: Advisory Group Member, Lilly USA, LLC; Investigator, Bayer Schering Pharma, Johnson & Johnson. 2012-06-25T10:00:00 Theater B 2012-06-25T00:00:00 1899-12-30T10:00:00 2478 263 1858 S39-2 T05-S05 Monday 285 2012


3003 ENDO12L_S39-3 SYMPOSIUM SESSION: CLINICAL - New Insights into Androgen Replacement Therapy (9:30 AM - 11:00 AM) Is Serum Testosterone Concentration in Men Genetically Determined? Claes Ohlsson Institute of Medicine, Sahlgrenska Academy, Gothenburg, Sweden Testosterone is the most important testicular androgen in men. Low serum testosterone concentrations are associated with cardiovascular morbidity, metabolic syndrome, type 2 diabetes mellitus, atherosclerosis, osteoporosis, sarcopenia, and increased mortality risk. Thus, there is growing evidence that serum testosterone is a valuable biomarker of men[apos]s overall health status. Studies in male twins indicate that there is a strong heritability of serum testosterone. We recently perform a large-scale genome-wide association study to examine the effects of common genetic variants on serum testosterone concentrations.[sup]1[/sup] By examining 14,429 men, we show that genetic variants in the sex hormone-binding globulin ([italic]SHBG[/italic]) locus and on the X chromosome are associated with a substantial variation in serum testosterone concentrations and increased risk of low testosterone. Two single-nucleotide polymorphisms at the sex hormone-binding globulin [italic](SHBG)[/italic] locus (17p13-p12) were identified as independently associated with serum testosterone concentration (rs12150660, p=1.2x10[sup]-41[/sup] and rs6258, p=2.3x10[sup]-22[/sup]). Subjects with [ge]3 risk alleles of these variants had 6.5-fold higher risk of having low serum testosterone than subjects with no risk allele. The rs5934505 polymorphism near [italic]FAM9B[/italic] on the X chromosome was also associated with testosterone concentrations (p=5.6x10[sup]-16[/sup]). The reported associations may now be used in order to better understand the functional background of recently identified disease associations related to low testosterone. Importantly, we identified the first known genetic variant, which affects SHBG[apos]s affinity for binding testosterone and the free testosterone fraction and could therefore influence the calculation of free testosterone. This finding suggests that individual-based SHBG-testosterone affinity constants might be required depending on the genotype of this single-nucleotide polymorphism.[br][br]1) [bold][underline]Ohlsson C[/underline][/bold][bold], Wallaschofski, H et al [/bold]2011 Genetic Determinants of Serum Testosterone Concentrations in men. [bold]PloS Genetics[/bold] [italic]Oct;7(10):e1002313[/italic].[br][br]Nothing to Disclose: CO 2012-06-25T10:30:00 Theater B 2012-06-25T00:00:00 1899-12-30T10:30:00 2399 263 1859 S39-3 T05-S05 Monday 286 2012


3004 ENDO12L_S40-1 SYMPOSIUM SESSION: BASIC - New Mechanisms for HPA Axis Regulation (9:30 AM - 11:00 AM) Circadian Clock Genes [amp] Regulation of Adrenal Glucocorticoid Secretion Henrik Oster University of Lubeck, Lubeck, Germany Besides acute activation by stress hypothalamus-pituitary-adrenal (HPA) axis regulation follows strong circadian ([sim]24h period) and ultradian ([lt]24h period) regulation. Under baseline conditions adrenal glucocorticoid (GC) secretion shows a robust circadian rhythm with blood levels peaking around wake-up time (i.e. in the morning in humans and at the beginning of the night in nocturnal rodents). While circadian HPA axis activity is ultimately controlled by the circadian pacemaker residing in the suprachiasmatic nuclei (SCN) of the hypothalamus, different so called peripheral clocks modulate HPA signal propagation at various levels. Circadian oscillators have been described in the paraventricular nucleus, the pituitary and the adrenal cortex. In the latter clock genes regulate the sensitivity of the steroidogenic machinery to ACTH stimulation and, hence, the GC responsiveness during the course of the day. Using a combination of molecular, genetic and surgical approaches we describe the mechanisms of SCN-to-adrenal clock interaction in the regulation of GC rhythms in the mouse and the role of GCs in the synchronization of the circadian machinery in other tissues. Recent data suggest that besides diurnal regulation, clock genes in the adrenal cortex also directly interfere with stress axis activation. In this way, the circadian system might regulate long-term psychological and physiological consequences of repetitive stress exposure.[br][br]Sources of Research Support: German Research Foundation (DFG) Emmy Noether Grant OS 353/4-1; Volkswagen Foundation Lichtenberg Grant.[br][br]Nothing to Disclose: HO 2012-06-25T09:30:00 Room 310 2012-06-25T00:00:00 1899-12-30T09:30:00 1649 264 1860 S40-1 T01-S06 Monday 288 2012


3005 ENDO12L_S40-2 SYMPOSIUM SESSION: BASIC - New Mechanisms for HPA Axis Regulation (9:30 AM - 11:00 AM) Fast Glucocorticoid Feedback Via Endocannabanoid Signaling Jeffrey G Tasker Tulane University, New Orleans, LA Stress leads to activation of corticotropin releasing hormone (CRH) cells in the hypothalamic paraventricular nucleus (PVN) and stimulation of the hypothalamic-pituitary-adrenal (HPA) axis, resulting in an increase in the systemic secretion of glucocorticoids. Glucocorticoids exert both rapid, transient actions and delayed, long-lasting actions in various target tissues throughout the organism, including in multiple areas of the brain. The delayed glucocorticoid effects are mediated by canonical nuclear receptor-mediated transcriptional actions of the steroid, while the rapid effects appear to be mediated by a membrane-associated receptor and cell signaling mechanisms. The acute glucocorticoid actions in the hypothalamus and pituitary gland cause rapid feedback inhibition of the HPA axis. We have investigated the cellular mechanisms responsible for the rapid glucocorticoid feedback inhibition of the HPA axis in a series of in vitro and in vivo studies. We have identified a novel mechanism of rapid glucocorticoid modulation of synaptic signaling in hypothalamic neuroendocrine cells that involves the activation of a membrane glucocorticoid receptor and the rapid synthesis of endocannabinoids in PVN neurons. The retrograde release at glutamate synapses of the endocannabinoid 2-arachidonoylglycerol leads to a presynaptic CB1 receptor-mediated suppression of glutamate release onto the PVN neurons, which causes an inhibition of the excitatory synaptic input to the PVN neurons. The rapid glucocorticoid actions appear to be mediated by the classical intracellular glucocorticoid receptor (GR) located at the membrane, since knocking out the GR causes the loss of the rapid glucocorticoid-induced eCB effect. In vivo, peripheral application of a CB1 antagonist blocked the rapid glucocorticoid suppression of HPA activation induced by restraint stress. Direct infusion of membrane-impermeant glucocorticoid into the PVN inhibited restraint-induced HPA activation, suggesting rapid feedback actions at the PVN. Infusion into the PVN of a CB1 receptor antagonist blocked the rapid glucocorticoid inhibition of the HPA axis. Our findings suggest, therefore, that the rapid glucocorticoid feedback inhibition of the HPA axis is mediated, in part, by the glucocorticoid activation of a membrane-associated GR and the GR-induced retrograde endocannabinoid suppression of the excitatory synaptic drive to the CRH cells.[br][br]Sources of Research Support: NIH grant MH069879; the Catherine and Hunter Pierson Chair in Neuroscience; the Tulane Research Enhancement Fund.[br][br]Nothing to Disclose: JGT 2012-06-25T10:00:00 Room 310 2012-06-25T00:00:00 1899-12-30T10:00:00 2545 264 1861 S40-2 T01-S06 Monday 289 2012


3006 ENDO12L_S40-3 SYMPOSIUM SESSION: BASIC - New Mechanisms for HPA Axis Regulation (9:30 AM - 11:00 AM) Molecular Regulation of CRH Gene Expression Greti Aguilera NIH/NICHD, Bethesda, MD Adequate regulation of hypothalamic corticotropin releasing hormone (CRH) secretion and expression is essential for circadian basal HPA axis activity and for stress adaptation. Transcriptional regulation of the CRH gene depends on cyclic AMP/protein kinase A signaling and binding of phospho-CREB to a CRE at [ndash]270 in the CRH promoter. This CRE is essential for activation of the CRH promoter, and DNA methylation at the internal CpG of this site reduces CREB binding to the promoter affecting CRH expression. However, phospho-CREB alone is not sufficient for driving CRH transcription and emerging evidence indicates that transcriptional activation requires the CREB co-activator, Transducer Of Regulated CREB activity (TORC) and its recruitment by the CRH promoter. In basal conditions, TORC is in the cytoplasm, phosphorylated by the Ser/Thr protein kinases, salt inducible kinase 1 and 2 (SIK1 and 2), and SIK inactivation by PKA allows TORC translocation to the nucleus. Over-expression of TORC1, 2 or 3 potentiates cyclic-AMP-stimulated CRH transcription, while TORC2 and 3 silencing using siRNA blunts it. Immunohistochemistry shows the presence of TORC2 in the cytoplasm of most CRH neurons in basal conditions. Restraint stress causes transient nuclear translocation of TORC2 in about 60% of CRH neurons by 30 min (returning to basal levels by 3 h). Activation of CRH transcription in vitro and in vivo is associated with recruitment of phospho-CREB and TORC2 by the CRH promoter. Increasing evidence indicate that activation/inactivation of TORC in the CRH neuron involves the TORC kinases SIK1 and SIK2. This includes marked induction of SIK1 concomitantly with the declining phase of CRH transcription, and the fact that over-expression of both SIK1 and SIK2 reduces nuclear translocation of TORC and CRH transcription, while the non-selective SIK inhibitor, staurosporin stimulates CRH transcription. Selective silencing of SIK1 or SIK2 using shRNA has revealed differential effects of both isoforms, suggestive that SIK2 is responsible for TORC sequestration in the cytoplasm in basal conditions, while induction of SIK1 probably inactivates TORC in the nucleus and limit CRH transcriptional responses. The overall evidence indicates that TORC is essential for activation of CRH transcription, and suggests that regulation of the SIK/TORC system by stress-activated signaling pathways acts as a sensitive switch mechanism for rapid activation and inactivation of CRH transcription.[br][br]Sources of Research Support: Intramural Research Program, Eunice Kennedy Shriver National Institute of Child Health and Human Development.[br][br]Nothing to Disclose: GA 2012-06-25T10:30:00 Room 310 2012-06-25T00:00:00 1899-12-30T10:30:00 2542 264 1862 S40-3 T01-S06 Monday 290 2012


3007 ENDO12L_S41-1 SYMPOSIUM SESSION: TRANSLATIONAL - Novel Signaling Pathways during Bone Formation (9:30 AM - 11:00 AM) IGF-I Crosstalk with Other Signaling Mechanisms during Bone Formation Daniel D Bikle VA Medical Center 111N-UCSF, San Francisco, CA IGF-I crosstalk with other signaling mechanisms during bone formation.[br]DD Bikle*1 and Y Wang1. Dept Medicine VA Medical Center and University of California San Francisco, San Francsico,CA[br]IGF-I is produced by chondrocytes, osteoblasts, osteocytes, and osteoclasts, and its receptor (IGF-IR) is found in all these cells. The relative contributions of circulating (primarily liver produced) vs paracrine (produced in bone) IGF-I to bone formation is not clear, but both are involved. The actions of IGF-I are mediated by IGF-IR, a tetrameric membrane bound receptor homologous to the insulin receptor. When activated this receptor sets into motion two signaling pathways, the PI3K/Akt pathway and the ras/raf/MAPK/ERK pathway. Akt activation reduces apoptosis by inhibiting BAD, promotes protein synthesis via mTOR, and facilitates [beta]-catenin activation by suppressing GSK3[beta]. Selected integrins are critical for the ability of IGF-I to activate its receptor, modulating the response of bone to IGF-I during skeletal loading and unloading. The global deletion of IGF-I or the chondrocyte specific deletion of IGF-IR leads to a profound reduction in proliferation, increased apoptosis, and blunted differentiation of chondrocytes in the growth plate associated with a reduction in Ihh expression but with increased PTHrP expression presumably accounting for the failure in differentiation. Global deletion of IGF-I or the osteoblast specific deletion of IGF-IR leads to osteopenia and decreased bone formation resistant to the anabolic actions of PTH. Even when IGF-IR is deleted in mature osteoblasts, the ability of PTH to stimulate CFUs (osteoprogenitors) is blocked. Similarly, deletion of IGF-I in osteoblasts and chondrocytes or IGF-IR receptor in osteoclasts blocks osteoclastogenesis and its stimulation by PTH. These results can best be understood by mechanisms involving cell cell signaling. Indeed two such intercellular signaling mechanisms, RANKL/RANK and ephrinB2/EphB4, are dependent for their expression on an intact IGF-I signaling pathway. Disruption of this pathway interferes with the normal regulation of chondrocyte proliferation and differentiation by Ihh/PTHrP and bone remodeling by PTH.[br][br]Sources of Research Support: NIH grants RO1AR055924 and RO1DK054793 awarded to DDB.[br][br]Nothing to Disclose: DDB 2012-06-25T09:30:00 Room 332 2012-06-25T00:00:00 1899-12-30T09:30:00 2463 265 1863 S41-1 T03-S05 Monday 292 2012


3008 ENDO12L_S41-2 SYMPOSIUM SESSION: TRANSLATIONAL - Novel Signaling Pathways during Bone Formation (9:30 AM - 11:00 AM) EphrinB2/EphB4 Interactions Regulating Stromal Cell/Osteoblast Differentiation [amp] Bone Formation Natalie A Sims St Vincent[apos]s Institute of Medical Research, Fitzroy, Australia Ephrins and their receptors (Ephs) are receptor tyrosine kinases that regulate tissue remodeling, including cell migration, angiogenesis and axon guidance. Through direct cell contact, interaction of membrane-bound ephrin ligands with their receptors simultaneously activates signaling through both the receptor (forward) and ligand (reverse). We have reported that ephrinB2 expression by osteoblasts is stimulated by parathyroid hormone (PTH) and its related protein (PTHrP), suggesting endocrine and paracrine regulation of this pathway. In cell culture, specific inhibitors that block signaling of ephrinB2 through EphB4 impaired mineralization by both human and murine osteoblast precursors, and strongly inhibited markers of late stage osteoblast / osteocyte expression. In contrast, they stimulated osteoblastic expression of RANKL, a critical factor for osteoclast differentiation and bone resorption.[br]To study the role of ephrinB2/EphB4 signaling in PTH anabolic action, we treated mice with PTH in the presence and absence of soluble EphB4 (sEphB4) to inhibit ephrinB2/EphB4 signaling. PTH treatment increased trabecular number and thickness, and dramatically increased all osteoblast and bone formation parameters without increasing osteoclast parameters. sEphB4 treatment, with or without PTH, increased osteoblast formation even above PTH-treated levels, but production of osteoid and formation of mineralized bone was not increased, consistent with inhibition of late stage osteoblast markers in vitro. Strikingly, sEphB4 treatment converted PTH anabolic effect to a resorptive response: osteoclast formation was increased and trabecular number reduced. This influence of sEphB4 was confirmed in vitro, where both osteoblastic RANKL production and osteoclast formation in response to PTH or 1,25-dihydroxyvitaminD3 was increased with addition of sEphB4. This enhancing effect of receptor blockade on osteoclast formation might result from the increased RANKL production, but could also relate to sEphB4 preventing ephrinB2 reverse signaling within the osteoclast lineage that has also been reported to limit osteoclast differentiation.[br]These results indicate that ephrinB2/EphB4 signaling enhances late stages of osteoblast differentiation, and communicates with the hemopoietic lineage to restrict osteoclast formation. The increased resorption resulting from ephrinB2/EphB4 receptor blockade needs further assessment in view of the interest in EphB4 blockade as anti-cancer therapy.[br][br]Nothing to Disclose: NAS 2012-06-25T10:00:00 Room 332 2012-06-25T00:00:00 1899-12-30T10:00:00 2455 265 1864 S41-2 T03-S05 Monday 293 2012


3009 ENDO12L_S41-3 SYMPOSIUM SESSION: TRANSLATIONAL - Novel Signaling Pathways during Bone Formation (9:30 AM - 11:00 AM) The Role of Sclerostin in Mediating the Anabolic and Catabolic Effects of PTH in Cortical and Trabecular Bone Alexander G Robling Indiana University School of Medicine, Indianapolis, IN Parathyroid hormone (PTH) is a potent calcium-regulating factor that has skeletal anabolic effects when administered intermittently, or catabolic effects when maintained at consistently high levels. Bone cells express PTH receptors, but the cellular responses to PTH in bone are incompletely understood. Wnt signaling has recently been implicated in the osteo-anabolic response to the hormone. Specifically, the Sost gene, a major antagonist of Wnt signaling, is down-regulated by PTH exposure. We investigated this mechanism by treating Sost-deficient mice and their wild-type littermates with anabolic and catabolic regimens of PTH, and measuring the skeletal responses. Male Sost+/+ and Sost-/- mice were injected daily with human PTH 1-34 (0, 30, or 90 [micro]g/kg) for 6 weeks. Female Sost+/+ and Sost-/- mice were infused with vehicle or high dose PTH (40 [micro]g/kg/day) for 3 weeks. DEXA-derived measures of intermittent PTH (iPTH)-induced bone gain were impaired in Sost-/- mice. Further probing revealed normal or enhanced iPTH-induced cortical bone formation rates but concomitant increases in cortical porosity among Sost-/- mice. Trabecular bone was highly responsive to iPTH in Sost-/- mice. Continuous PTH (cPTH) infusion resulted in equal bone loss in Sost+/+ and Sost-/- mice as measured by DEXA. However, distal femur trabecular bone was spared the bone-wasting effects of cPTH in Sost-/- mice. These results suggest that changes in Sost expression are not required for iPTH-induced anabolism. iPTH-induced resorption of cortical bone might be over-stimulated in Sost deficient environments. Furthermore, Sost deletion protects the trabecular but not cortical compartments from bone loss induced by high-dose PTH infusion.[br][br]Nothing to Disclose: AGR 2012-06-25T10:30:00 Room 332 2012-06-25T00:00:00 1899-12-30T10:30:00 2429 265 1865 S41-3 T03-S05 Monday 294 2012


3010 ENDO12L_S42-1 SYMPOSIUM SESSION: CLINICAL - State of the Art Management of Nodular Thyroid Disease (9:30 AM - 11:00 AM) Application of Molecular Analysis to Clinical Practice Electron Kebebew National Cancer Institute, Bethesda, MD Nothing to Disclose: EK 2012-06-25T09:30:00 Theater A 2012-06-25T00:00:00 1899-12-30T09:30:00 6018 266 1866 S42-1 T02-S03 Monday 296 2012


3011 ENDO12L_S42-2 SYMPOSIUM SESSION: CLINICAL - State of the Art Management of Nodular Thyroid Disease (9:30 AM - 11:00 AM) Follow Up Recommendation for Benign Nodular Disease Edward K Tomimori Sao Paulo University School of Medicine, Sao Paulo, Brazil Disclosure Incomplete: EKT 2012-06-25T10:00:00 Theater A 2012-06-25T00:00:00 1899-12-30T10:00:00 6019 266 1867 S42-2 T02-S03 Monday 297 2012


3256 ENDO12L_M12 MEET-THE-PROFESSOR: CLINICAL - Medical Management of Obesity (2:45 PM - 3:30 PM) Medical Management of Obesity Florencia Halperin Brigham and Women[apos]s Hosp, Boston, MA 2012-06-23T14:45:00 Room 372 2012-06-23T00:00:00 1899-12-30T14:45:00 6303 86 864 M12 OB2 Saturday 12 2012


3257 ENDO12L_M33A MEET-THE-PROFESSOR: CLINICAL - CGMS Sensors: Who Benefits; How [amp] When To Use? (1:00 PM - 1:45 PM) CGMS Sensors: Who Benefits; How & When to Use Joseph Anthony Aloi ' Session supported by: Dexcom, Inc. [amp] Medtronic Diabetes 2012-06-25T13:00:00 Room 352 DEF 2012-06-25T00:00:00 1899-12-30T13:00:00 6288 282 1941 M33 D3A Monday 33 2012


3258 ENDO12L_M12A MEET-THE-PROFESSOR: CLINICAL - Medical Management of Obesity (5:30 PM - 6:15 PM) Medical Management of Obesity Florencia Halperin Brigham and Women[apos]s Hosp, Boston, MA 2012-06-23T17:30:00 Room 372 2012-06-23T00:00:00 1899-12-30T17:30:00 6303 102 902 M12 OB2A Saturday 12 2012


19 ENDO12L_OR04-1 ORAL SESSION: Female Reproduction: The Endometrium [amp] Pregnancy (11:15 AM-12:45 PM) Increased Pregnancy Losses and Poor Neonatal Outcomes in Women with First-Trimester TSH Levels between 2.5 and 4 mIU/L Compared to Euthyroid Women with TSH Less Than or Equal to 2.5 Jubbin Jagan Jacob, Kewal Aditya, Shweta Achint, Tapasya Dhar, Kumkum Avasti Christian Medical College and Hospital, Ludhiana, India; Christian Medical College and Hospital, Ludhiana, India Context: Though maternal and foetal outcomes in overt hypothyroidism have been well studied, data regarding outcomes in subclinical hypothyroidism is conflicting. A recent publication has shown an increase in pregnancy losses among antibody negative women with TSH levels between 2.5 and 5.0mIU/L.(1)[br]Objective: To evaluate pregnancy, maternal and neonatal outcomes in women with first trimester TSH levels [gt] 2.5 but [le] 4mIU/L compared to euthyroid women with TSH [le] 2.5mIU/L irrespective of thyroid antibody levels.[br]Design: The study is part of a prospective study evaluating routine screening of thyroid function tests among pregnant women in the first trimester (Thyroid Screening in Urban Ludhiana Pregnancy Study: TULIPS). Outcomes evaluated include early and late pregnancy losses, 12 pregnancy related maternal complications including preterm delivery, birth weights and 5 neonatal complications. All patients with TSH [gt] 4mIU/L were treated with levothyroxine. For the present study the women were divided into two groups based on the first trimester TSH value, group A with TSH [le] 2.5mIU/L and group B with TSH [gt] 2.5 but [le] 4mIU/L.[br]Setting: The study was conducted in the antenatal clinic of a university affiliated teaching hospital.[br]Patients: Of the 1000 patients recruited, data was available for 951, of whom 790 delivered/aborted in the study hospital and in another 161 patients; the outcomes were obtained over the telephone.[br]Results: There were 533 patients in group A and 263 in group B. There was a significant increase in early ([le] 20 wks.) and late ([gt] 20 wks.) pregnancy losses in group B vs. group A (Early; 8.4% vs. 4.1% p[lt]0.001 [OR (95%CI) 2.1(1.1-3.9)], Late; 4.2% vs. 0.6% p[lt]0.001 [OR (95%CI) 7.7(2.1-27.8)] and combined 12.5% vs. 4.7%, p[lt]0.001[OR (95%CI) 2.9(1.6-5.0)]). The odds (95%CI) of preterm delivery ([lt]37wks.) in group B was 2.02(1.1-3.7) compared to group A. There were no statistically significant differences between the two groups among the other 11 maternal outcomes. Neonates born to women in group B had significantly higher odds of being LBW (24.2% vs. 16.6% p=0.01 [OR (95%CI) 1.6(1.1-2.3)]), SGA (18.6% vs. 10.7%, p[lt]0.001 [OR (95%CI) 1.9(1.2-2.9)]) and had more instances of neonatal hyperbilirubinemia (12.1% vs. 7.5% p=0.04).[br]Conclusions: The clear cut increase in pregnancy losses with TSH [gt] 2.5 but [le] 4mIU/L and significant adverse neonatal outcomes provides another evidence for lowering the TSH range in pregnant women.[br][br]Negro R et al.,J Clin Endocrinol Metab 2010;E44-48.[br][br]Nothing to Disclose: JJJ, KA, SA, TD, KA 2012-06-23T11:15:00 352DEF 2012-06-23T00:00:00 1899-12-30T11:15:00 970 37 74 OR04-1 OR200-01 Saturday 19 2012


20 ENDO12L_OR04-2 ORAL SESSION: Female Reproduction: The Endometrium [amp] Pregnancy (11:15 AM-12:45 PM) Evidence for Bone Marrow Origin of Endometrial Cells in a Murine Model Sara S Morelli, Pranela Rameshwar, Laura T Goldsmith New Jersey Medical School of UMDNJ, Newark, NJ; New Jersey Medical School of UMDNJ, Newark, NJ Mechanisms used in cyclic regeneration of endometrial cellular compartments are unknown. Recent evidence supports the existence of resident stem cell population(s) which give rise to mature endometrial parenchymal cell types. Their origin is unknown. Few studies have attempted to determine the cellular origin of endometrial cells. Current studies determined whether bone marrow (BM) is a source of endometrial epithelial and/or stromal cells.[br]BM cells, harvested from Green Fluorescent Protein (GFP) transgenic donor mice, were injected into lethally irradiated, immature female mice (two independent expts, n=7 or 8 recipients in each). Flow cytometry was used to detect GFP positive cells in peripheral blood of recipient mice. Vaginal cytology determined whether irradiated mice were undergoing estrus cycles. Four animals with successful BM reconstitution, confirmed to be cycling, were sacrificed at either 5 months (n=2, 1 from each expt) or 9 months (n=2, 1 from each expt) post transplant, and hysterectomy was performed. Uteri were assessed by confocal laser microscopy to identify and localize GFP positive cells and fluorescently stained nuclei in recipient uterine tissue sections. Cell counting was performed using computer-assisted image analysis. Data are expressed as GFP positive cell number/total number of stromal compartment cells.[br]5/8 (Expt 1) and 6/7 (Expt 2) recipient mice were successfully reconstituted with donor BM at 7-8 weeks post transplant. In the 11 reconstituted animals, the range of GFP positive peripheral blood cells was 81.3 to 92.2%. At 5 months post transplant, 5.5% of cells (148/2690, Expt 1, n=1 animal) and 3.6% of cells (95/2659, Expt 2, n=1) in the endometrial stromal compartment were GFP positive. At 9 months post transplant, 21.5% of cells (584/2721, Expt 1, n=1) and 18.2% of cells (305/1678, Expt 2, n=1) in the endometrial stromal compartment were GFP positive. In distinct contrast, no endometrial glandular or luminal epithelial cells were GFP positive.[br]These data demonstrate that bone marrow is a significant source of endometrial cells in the stromal, but not epithelial, compartment. The contribution of bone marrow derived-cells to the endometrial stromal compartment increases with time/with increasing number of estrus cycles. These novel data provide an important advance in our understanding of the cellular source of regenerative cells of the endometrium.[br][br]Sources of Research Support: American Board of Obstetrics and Gynecology/American Association of Obstetricians and Gynecologists Foundation Research Training Scholarship; American Society for Reproductive Medicine Career Development Award in Reproductive Medicine and Biology.[br][br]Nothing to Disclose: SSM, PR, LTG 2012-06-23T11:30:00 352DEF 2012-06-23T00:00:00 1899-12-30T11:30:00 1745 37 75 OR04-2 OR200-01 Saturday 20 2012


21 ENDO12L_OR04-3 ORAL SESSION: Female Reproduction: The Endometrium [amp] Pregnancy (11:15 AM-12:45 PM) Absence of Kr[uuml]ppel-Like Factor 9 (KLF9) Promotes Ectopic Endometrial Lesion Development in a Mouse Model of Endometriosis Melissa E Heard, Christian D Simmons, Kim E Light, Frank A Simmen, Rosalia CM Simmen University of Arkansas for Medical Sciences, Little Rock, AR; University of Arkansas for Medical Sciences, Little Rock, AR; Arkansas Children[apos]s Research Institute, Little Rock, AR Endometriosis is a benign gynecological disorder commonly associated with uterine progesterone (P) -resistance and reduced fertility. Kr[uuml]ppel-like factors (KLF) are zinc finger-containing transcription factors which regulate diverse physiological processes in the female reproductive tract. Our group has identified KLF9 as a transcriptional co-regulator of estrogen and P receptor (PGR) signaling in endometrial stroma and glandular epithelial cells [italic]in vitro[/italic]. We recently demonstrated that [italic]KLF9[/italic] mRNA levels are decreased in eutopic endometria of women with vs. without endometriosis. Mice null for [italic]Klf9[/italic] are sub-fertile due to decreased numbers of implantation sites and exhibit attenuated P-sensitivity of uterine endometrium. To determine a causal relationship between P-resistant endometrium due to loss of KLF9 transcriptional control and predisposition to endometriosis, we utilized a mouse model of endometriosis induced by intraperitoneal (IP) injection of minced donor endometrial tissue into immune competent recipient mice. Initial studies with endometria from intact mice expressing enhanced green fluorescent protein administered IP into mice of the same strain confirmed the formation of peritoneal ectopic lesions. To compare predisposition of [italic]Klf9[/italic]-expressing (WT) and [italic]Klf9[/italic] null (KO) endometria for ectopic lesion development, WT recipient mice (8-10 weeks) in the estrous phase were administered IP with minced endometrium ([sim]40 mg tissue in 400[micro]l PBS) from WT or KO mice (10-12 weeks, estrus phase) or with PBS alone (sham control). Eight weeks later, recipient mice (n=10 and 9, respectively for WT and KO donors; one donor to one recipient) were analyzed for presence, size, and histology of endometrial lesions and for body weight changes pre- and post-endometriosis. While 50% of mice injected with donor WT tissue developed detectable lesions, all recipients of donor KO endometrium showed established peritoneal lesions. Donor WT and KO endometrial tissues generated similar numbers of lesions in WT recipient mice; however, lesions from KO donors were larger than those from WT donors (30.3[plusmn]11.3 vs. 19.8[plusmn]12.0 mm[sup]3[/sup]). Mice with and without ectopic lesions did not differ in body, uterine or ovarian weights. Eutopic endometria of WT mice with ectopic lesions did not differ in expression levels of [italic]Pgr[/italic], [italic]Igfbp1[/italic], [italic]Wnt4[/italic] and[italic] Wnt2[/italic] as a function of donor source at sac. Our findings suggest that endometrial KLF9 deficiency maybe causal to the etiology of human endometriosis.[br][br]Sources of Research Support: NIH-HD21961 and ABI-ACHRI Student and Clinical Staff Intramural Grants.[br][br]Nothing to Disclose: MEH, CDS, KEL, FAS, RCMS 2012-06-23T11:45:00 352DEF 2012-06-23T00:00:00 1899-12-30T11:45:00 324 37 76 OR04-3 OR200-01 Saturday 21 2012


22 ENDO12L_OR04-4 ORAL SESSION: Female Reproduction: The Endometrium [amp] Pregnancy (11:15 AM-12:45 PM) A Novel Role for Oviductal Epithelial Estrogen Receptor [alpha] during Fertilization and Early Embryo Development Wipawee Winuthayanon, Sylvia C Hewitt, Elizabeth Padilla-Banks, Grant D Orvis, Richard R Behringer, Carmen J Williams, Kenneth S Korach National Institute of Environmental Health Sciences (NIEHS)/National Institute of Health (NIH), Research Triangle Park, NC; Sloan-Kettering Institute, New York, NY; University of Texas MD Anderson Cancer Center, Houston, TX Loss of epithelial estrogen receptor [alpha] (ER[alpha]) from the uterus causes female infertility, in part, due to an implantation defect. We hypothesized that loss of oviductal epithelial ER[alpha] also contributed to their infertility. By flushing embryos at various times after mating, we showed that epithelial ER[alpha] conditional knockout (cKO) mice lost all of their embryos between days 1 and 2 of pregnancy. Although similar numbers of oocytes were ovulated, only [sim]50% of oocytes collected from the cKO oviducts were fertilized when compared to control littermates, and no 2-cell embryos were ever recovered on pregnancy day 2. The fertilization rate of the cKO oocytes was restored to that of control littermates only when the oocytes were collected directly from the ovaries prior to ovulation and then fertilized in vitro Furthermore, when wild type zygotes were transferred into oviducts of pseudopregnant cKO mice, the majority of the embryos were under-developed when flushed from the oviduct at 3.5 dpc, and no embryos were found in the uterus. In contrast, all transferred embryos collected from pseudopregnant control littermates were morulas or blastocysts, and all of these embryos had reached the uterus. These findings suggest that ablation of ER[alpha] in the oviductal epithelium results in alterations in the oviductal microenvironment that completely disrupts fertilization, preimplantation embryo development and embryo transport. Microarray analysis of cKO and control oviducts collected at 0.5 and 1.5 dpc demonstrated significant alterations in expression of genes involved in interleukin 17 signaling and prostaglandin D2 synthesis. These findings were confirmed at the protein level by analysis of IL17 and prostaglandin D synthase. It can be concluded that epithelial ER[alpha] is required to establish the appropriate oviduct environment for successful mouse preimplantation embryo development. Our findings demonstrate a highly novel and unexpected regulatory phenomenon of epithelial ER[alpha] in the female reproductive tract that has the potential to provide a better understanding of idiopathic infertility in women.[br][br]Nothing to Disclose: WW, SCH, EP-B, GDO, RRB, CJW, KSK 2012-06-23T12:00:00 352DEF 2012-06-23T00:00:00 1899-12-30T12:00:00 988 37 77 OR04-4 OR200-01 Saturday 22 2012


23 ENDO12L_OR04-5 ORAL SESSION: Female Reproduction: The Endometrium [amp] Pregnancy (11:15 AM-12:45 PM) Bone Morphogenic Protein Receptor Type 2 (BMPR2) Is Required for Establishment of an Adequate Uterine Environment To Support Normal Placentation Takashi Nagashima, Qinglei Li, John P Lydon, Francesco J DeMayo, Martin M Matzuk Baylor College of Medicine, Houston, TX; Baylor College of Medicine, Houston, TX; Baylor College of Medicine, Houston, TX; Baylor College of Medicine, Houston, TX; Texas A[amp]M University, College Station, TX Bone morphogenetic proteins (BMPs), known to play a critical role in reproductive function, are members of transforming growth factor-[beta] (TGF-[beta]) superfamily and regulate a variety of cellular functions. BMP receptor type 2 (BMPR2) transduces BMPs signals by forming heteromeric complexes with BMP type 1 receptors. Of note, mutant embryos homozygous for a [italic]Bmpr2[/italic] null allele ([italic]Bmpr2[sup]-/-[/sup][/italic]) die during gastrulation. To overcome the early embryonic lethality and understand the [italic]in vivo[/italic] roles of BMPR2 in reproduction, we used progesterone receptor Cre ([italic]Pgr-Cre[/italic]) to conditionally disrupt the [italic]Bmpr2[/italic] gene in the female reproductive tract. [italic]Bmpr2[/italic] conditional knockout ([italic]Bmpr2[/italic] cKO: [italic]Bmpr2[sup]flox/-[/sup]Pgr[sup]cre/+[/sup][/italic]) female mice are complete infertile. However, the ovarian function remains normal, as evidenced by normal ovarian morphology, follicle growth, cumulus expansion, ovulation, oocyte fertilization, and hormone levels, indicating the infertility of [italic]Bmpr2[/italic] cKO mice arises from uterine dysfunction. [italic]Bmpr2[/italic] cKO uteri showed reduced uterine weight after artificial decidualization, which likely results from defective proliferation and differentiation of uterine stromal cells and reduction of blood vessel diameter. However, implantation and early embryo development occurred normally in [italic]Bmpr2[/italic] cKO mice, as shown by normal embryo morphology and weight, number of implantation sites, and implantation- and trophoblast-specific-gene expressions. Strikingly, [italic]Bmpr2[/italic] cKO mice showed severe hemorrhage in trophoblast and decidual tissues of postimplantation uteri at the initiation of vascular remodeling. This defect was accompanied by multiple abnormalities, leading to fetal-growth retardation and loss of the fetus before birth. These results indicate that BMPR2 is indispensable for the establishment of an adequate uterine environment to support normal placentation in mice.[br][br]Sources of Research Support: NIH Grant HD32067 from the Eunice Kennedy Shriver National Institute of Child Health and Human Development.[br][br]Nothing to Disclose: TN, QL, JPL, FJD, MMM 2012-06-23T12:15:00 352DEF 2012-06-23T00:00:00 1899-12-30T12:15:00 1069 37 78 OR04-5 OR200-01 Saturday 23 2012


24 ENDO12L_OR04-6 ORAL SESSION: Female Reproduction: The Endometrium [amp] Pregnancy (11:15 AM-12:45 PM) Transcription Factor ZEB1 Acting Via Specific microRNAs (miRNA/miR) Plays a Pivotal Role in Hormonal Control of Uterine Quiescence and Contractility during Pregnancy and Labor Koriand[apos]r Williams, Nora E Renthal, Carole R Mendelson University of Texas Southwestern Medical Center, Dallas, TX Uterine quiescence is maintained during pregnancy by progesterone (P[sub]4[/sub]) acting via its nuclear receptors (PR) to inhibit expression of genes encoding contraction-associated proteins (CAPs) ([italic]e.g[/italic]. oxytocin receptor (OTR), connexin-43 (CX43) and cyclooxygenase-2 (COX-2)) in the myometrium. This occurs, in part, via P[sub]4[/sub]/PR anti-inflammatory actions. On the other hand, parturition is associated with a decline in PR function, resulting in an increased inflammatory response within the myometrium. Moreover, circulating estradiol (E[sub]2[/sub]) levels and/or estrogen receptor [alpha] activation, which increase markedly near term, contribute to the uterine inflammatory response and antagonize the anti-inflammatory actions of P[sub]4[/sub]/PR. Previously, we found that during the transition to labor the decline in PR function resulted in decreased expression of ZEB1/2, which allowed for the induction of miR-200 expression and increased transcription of OTR and CX43. Interestingly, we recently identified a conserved cluster of miRNAs, miR-199a and miR-214, that were significantly downregulated in pregnant mouse and human myometrium near term and during labor, whereas, their target, COX-2, a critical enzyme in the production of contractile prostaglandins, was coordinately increased. This pattern of expression also was observed in an inflammatory (lipopolysaccharide-induced) mouse model of preterm labor. Physiological relevance of these miRNAs was suggested by the finding that overexpression of miR-199a-3p/214 mimics in cultured human myometrial cells inhibited COX-2 protein expression and blocked TNF-[alpha]-induced myometrial cell contractility. Notably, E[sub]2[/sub] treatment of ovariectomized mice suppressed uterine miR-199a-3p/214 expression and coordinately induced COX-2 protein, whereas P[sub]4[/sub] treatment enhanced miR-199a-3p/214 expression and repressed COX-2. Intriguingly, these opposing hormonal effects were mediated by ZEB1, which we found to activate miR199a/214 expression. Remarkably, ZEB1 was inhibited by E[sub]2[/sub] and stimulated by P[sub]4[/sub] in uterine tissues of ovarectomized mice. Consequently, binding of endogenous ZEB1 to the miR-199a/214 promoter was found to be diminished in the pregnant mouse uterus at term. Furthermore, in co-transfected COS-7 cells, ZEB1 overexpression induced miR-199a/214 promoter activity. Our collective findings point to the key pivotal roles of myometrial ZEB1 and its miRNA targets as hormonally-controlled regulators of inflammatory and contractile gene expression in the pregnant uterus.[br][br]Sources of Research Support: P01-HD-11149, Prematurity Research Initiative Grant (#21FY11), March of Dimes Foundation, Pharmacological Sciences Training Grant.[br][br]Nothing to Disclose: KW, NER, CRM 2012-06-23T12:30:00 352DEF 2012-06-23T00:00:00 1899-12-30T12:30:00 1678 37 79 OR04-6 OR200-01 Saturday 24 2012


25 ENDO12L_OR05-1 ORAL SESSION: Hormone Dependent Tumors (11:15 AM-12:45 PM) Mitochondrial Estrogen Receptor Beta Dictates Tamoxifen Resistance or Sensitivity in Breast Cancer Ali Pedram, Mahnaz Razandi, Suzanne Fuqua, Ellis R Levin University of California, Irvine, CA; Long Beach VA Medical Center, Long Beach, CA; Baylor College of Medicine, Houston, TX Tamoxifen (TAM) resistance in breast cancer occurs in approximately 50% of women. TAM[apos]s effectiveness importantly involves a cyto-toxic action and underlies responses to TAM. We investigated several TAM sensitive or resistant cell lines most notably MCF7-BK (sensitive) or MCF7 BK-TR (resistant) cells. At 24 hours, TAM induced strong mitochondrial ROS generation in BK cells that caused a 13-fold increase in apoptosis compared to no TAM. Increased ROS and apoptosis resulted from 70% decreased MnSOD activity, a mitochondrial enzyme that processes superoxide ROS. Decreased mitochondrial MnSOD activity was due to TAM inducing nitric oxide-dependent nitrosylation at tyrosine 34 of MnSOD, by our mutational analysis required for inhibition of MnSOD activity. TAM-generated ROS induced PKCdelta and JNK kinase activation, BAX translocation to mitochondria, and the TAM-induced secretion of cytochrome c (apoptosome component) directly from isolated mitochondria. These functions in BK cells resulted from TAM acting as an ERbeta antagonist, shown by siRNA knockdown of ERbeta or ERalpha, and opposition to TAM by DPN (ERbeta agonist) but not PPT (ERalpha agonist). ERbeta was mainly localized to the mitochondria of the cells by fluorescent microscopy and sub-cellular immuno-blotting. In contrast, TAM acted as an ERbeta agonist in BK-TR (resistant) cells, decreasing nitrosylation to stimulate MnSOD activity, diminishing ROS levels and preventing apoptosis. Knocking down MnSOD with siRNA resulted in reversal of resistance to TAM in BK-TR cells, increasing cell death 15-fold. BK-TR tumors were grown in both flanks of nude mice (n=6) under TAM pellet administration for 5 weeks to 1200mm[sup]3[/sup]on average. Tumors were then injected with either lentiviral control or MnSOD shRNAs. MnSOD knockdown was seen only in the latter tumors that substantially regressed to 480mm[sup]3[/sup] at 8 weeks while control shRNA-injected tumors grew to 1690mm[sup]3[/sup], all in the presence of TAM. Caspase 7 was cleaved and apoptosis was confirmed (Annexin V histological staining) only in the MnSOD shRNA-injected tumors. MnSOD lentiviral shRNA in matrigel at initiation almost completely prevented in-vivo tumor formation over 5 weeks, compared to control shRNA and BK-TR cells in matrigel. Thus, mitochondrial ERbeta serves as an ROS rheostat to determine breast tumor fate. Inhibiting the tumor MnSOD and engaging ERbeta as antagonist overcomes TAM resistance and promotes apoptosis in human breast cancer.[br][br]Sources of Research Support: NCI Grant CA100366 (ERL).[br][br]Nothing to Disclose: AP, MR, SF, ERL 2012-06-23T11:15:00 342ABDE 2012-06-23T00:00:00 1899-12-30T11:15:00 24 38 80 OR05-1 OR42-01 Saturday 25 2012


26 ENDO12L_OR05-2 ORAL SESSION: Hormone Dependent Tumors (11:15 AM-12:45 PM) SLIRP Represses Retinoic Acid, SOX9 and Notch Signaling in Colorectal Cancer and Is a Good Prognostic Factor Patrick Aaron Candy, Michael Phillips, Andrew Redfern, Lisa Stuart, John A Davidson, Barry Iacopetta, Ben Wood, Shane M Colley, Peter J Leedman University of Western Australia, Western Australian Institute for Medical Research (WAIMR), Perth, Australia; University of Western Australia, Perth, Australia; Royal Perth Hospital, Perth, Australia; Sir Charles Gairdner Hospital and QEII Medical Centre, Perth, Australia; University of Western Australia, Perth, Australia Colorectal cancer (CRC) is the second highest cause of cancer death and the third most common malignancy. Aberrant Notch signaling in CRC promotes epithelial to mesenchymal transition, chemoresistance and metastasis. Its effects are mediated through transcription factors (TFs), in particular HES1 and HEY1, which influence COX2, NF-[kappa]B and Wnt signaling. In CRC, Notch signaling can be activated by retinoic acid, through the nuclear receptor (NR) RAR[alpha] and the early developmental TF SOX9, which activates HES1. Using CRC tissue microarrays (TMAs), we found that SLIRP, a nuclear receptor (NR) corepressor, is a good prognostic factor in CRC, with a tumor suppressor phenotype. High tumor SLIRP expression in a TMA cohort of 967 patients correlated with improved 5 year survival (p[lt]0.01) and inversely with tumor stage and lymph node invasion. In two CRC cDNA microarray sets, high SLIRP mRNA expression correlated with improved disease free survival over three years following surgery (p[lt]0.05). Here we investigated the mechanism for this clinical advantage and the hypothesis that SLIRP is a repressor Notch signaling. In human CRC cell transfection studies using luciferase (Luc) reporters, siRNA mediated depletion of SLIRP increased the activity of ligand-stimulated RAR[alpha], HES1 and SOX9 signaling, resulting in increased expression of downstream targets, including NOTCH2, NOTCH3, NFKB1, NFKB2, LMO2, HES1 and SOX9. In ChIP studies, SLIRP was recruited (with RAR[alpha]) to the HES1 and SOX9 promoters. When SLIRP was depleted from the cells with siRNA, ChIP studies revealed an increase in RAR[alpha] recruitment to the HES1 and SOX9 promoters, along with increased binding of SOX9 to the HES1 promoter, and a decrease in binding of HES1 to its own promoter. Further, we found that siRNA mediated SLIRP depleted CRC cells were more invasive in matrigel assays and more resistant to the standard CRC chemotherapeutic agents, 5-Fluorouracil and irinotecan. Taken together, these data suggest that SLIRP is a potent suppressor of Notch and RAR signaling in CRC and provide insight into the mechanisms by which SLIRP functions as a tumor suppressor to reduce invasion and enhance sensitivity to chemotherapy in this disease.[br][br]Nothing to Disclose: PAC, MP, AR, LS, JAD, BI, BW, SMC, PJL 2012-06-23T11:30:00 342ABDE 2012-06-23T00:00:00 1899-12-30T11:30:00 1055 38 81 OR05-2 OR42-01 Saturday 26 2012


27 ENDO12L_OR05-3 ORAL SESSION: Hormone Dependent Tumors (11:15 AM-12:45 PM) The Prostate Cancer Specific TMPRSS2:ERG (T/E) Fusion Cooperates with the Vitamin D Receptor To Induce CYP24A1, the Enzyme Responsible for Inactivation of 1,25-Dihydroxyvitamin D[sub]3[/sub] Jung-Sun Kim, Justin Michael Roberts, William Edward Bingman, Nancy Lynn Weigel Baylor College of Medicine, Houston, TX Results of epidemiological and clinical studies assessing the potential benefit of vitamin D in prostate cancer (PCa) have been conflicting even though pre-clinical studies in PCa models have shown that activation of the vitamin D receptor (VDR) reduces prostate cell proliferation and tumor growth. Although the majority of human PCas express a TMPRSS2:ETS factor fusion gene (T/E) which links the androgen regulated promoter of TMPRSS2 to the coding regions of ETS transcription factors (most commonly ERG), most pre-clinical studies have been performed in models lacking T/E. The level of the active metabolite of vitamin D, 1,25-dihydroxyvitamin D[sub]3[/sub] (1,25D), is controlled in part by VDR dependent induction of CYP24A1 which inactivates 1,25D. Since ETS factors can cooperate with VDR to induce CYP24A1, we hypothesized that T/E would cause aberrant induction of CYP24A1 limiting the beneficial effects of VDR action. In VCaP cells, ERG specific siRNAs reduced T/E to 30% of control and reduced 1,25D mediated induction of CYP24A1 to less than 10% of control. In contrast, 1,25D dependent induction of another vitamin D target gene (TRPV6) was not reduced. Induction of CYP24A1 by 1,25D was greatly increased in an LNCaP cell line that overexpresses a T/E isoform (Type VI +72). In T/E overexpressing LNCaP cells, 1 nM 1,25D caused an induction of CYP24A1 equivalent to 100 nM 1,25D in control LNCaP cells. T/E expression also increased maximal CYP24A1 expression at optimal levels of 1,25D suggesting that ETS factors are limiting in cells lacking T/E. Thus, one of the early effects of T/E in prostate cells is a reduction in intracellular 1,25D, which may lead to increased proliferation. Our ChIP analyses showed that ERG bound to the CYP24A1 promoter region predicted to contain both VDR and ETS factor binding sites. Preliminary data show that 1,25D enhances the extent of ERG binding suggesting an interaction between the factors. YK-4-279, an ERG inhibitor, did not reduce 1,25D mediated CYP24A1 induction in VCaP cells. The mechanism by which YK-4-279 inhibits ERG activity in PCa cells is unknown, but it does not prevent DNA binding and cannot prevent cooperation with VDR. Thus, although this compound would be beneficial in counteracting other activities of T/E, it would not prevent the aberrant induction of CYP24A1 by T/E. These new findings suggest that presence of T/E may reduce beneficial actions of VDR by increasing expression of CYP24A1 which inactivates 1,25D.[br][br]Nothing to Disclose: J-SK, JMR, WEB, NLW 2012-06-23T11:45:00 342ABDE 2012-06-23T00:00:00 1899-12-30T11:45:00 1127 38 82 OR05-3 OR42-01 Saturday 27 2012


28 ENDO12L_OR05-4 ORAL SESSION: Hormone Dependent Tumors (11:15 AM-12:45 PM) Macrophage-Derived Amphiregulin Causes Estrogen Receptor-[alpha] Down-Regulation in Tamoxifen-Resistant Breast Cancer Cells Fabio Stossi, Zeynep Madak-Erdogan, Rosa Ventrella, Benita S Katzenellenbogen Baylor College of Medicine, Houston, TX; University of Illinois at Urbana-Champaign, Urbana, IL Breast cancer is the second leading cause of death in US women. Approximately 70% of primary breast tumors express the estrogen receptor alpha (ER), which is routinely used as a prognostic marker and is pharmacologically targeted by endocrine therapies (e.g. tamoxifen or aromatase inhibitors). However, up to 1/3 of women that receive tamoxifen treatment will have a recurrence that has become resistant to the therapy. In this study, we wanted to follow up on our previous observation (1) that macrophage-derived factors can cause down-regulation of ER expression, thus directly impinging on the efficacy of endocrine therapies. We therefore analyzed the actions of macrophages on breast cancer cells that are resistant to tamoxifen treatment because relatively little is known about the role of elements of the tumor microenvironment in the development of endocrine resistance.[br]In this study, we found that macrophage derived factors caused ER down-regulation also in tamoxifen resistant breast cancer cells. Interestingly this happened under different macrophage polarization conditions when compared with tamoxifen sensitive cells. In resistant cells, conditioned media from unpolarized or M2 (pro-tumoral)-like macrophages was responsible for ER[alpha] loss while M1 (anti-tumoral)-like caused this in tamoxifen sensitive cells. By analyzing macrophage conditioned media activated pathways in tamoxifen resistant cells; we found that ER[alpha] down-regulation was mediated via an EGFR-dependent mechanism. Through protein arrays, recombinant factor treatments and antibody-depletion studies, we identified amphiregulin as the main EGFR ligand that elicits reduction of ER[alpha] levels in breast cancer cells. Of note, our findings revealed that this amphiregulin/EGFR paracrine, not autocrine, loop switches the growth potential of tamoxifen resistant breast cancer cells from ER-dependent to EGFR-dependent. No effect of amphiregulin or EGFR inhibitors was seen in tamoxifen sensitive cells. These observations further validate the importance of understanding which factors are present in the tumor microenvironment that will directly impinge on tumor cells, altering the signaling and growth milieu while directly contributing to the success or failure of therapeutic approaches.[br][br](1)Stossi, F., Madak-Erdogan, Z., and Katzenellenbogen, B.S. Macrophage-derived Soluble Factors Cause Loss of Estrogen Receptor Alpha in Breast Cancer Cells via Hyper-activation of ERK2 and c-Jun. Oncogene in press, PMID: 21860415.[br][br]Sources of Research Support: Grant from The Breast Cancer Research Foundation (BSK).[br][br]Nothing to Disclose: FS, ZM-E, RV, BSK 2012-06-23T12:00:00 342ABDE 2012-06-23T00:00:00 1899-12-30T12:00:00 1250 38 83 OR05-4 OR42-01 Saturday 28 2012


29 ENDO12L_OR05-5 ORAL SESSION: Hormone Dependent Tumors (11:15 AM-12:45 PM) Role of microRNA-141 in Hormone-Dependent Breast Cancer Jessica Finlay-Schultz, Diana M Cittelly, Peter Hendricks, Purvi R Patel, Dawn M Cochrane, Britta M Jacobsen, Jennifer K Richer, Carol A Sartorius University of Colorado Denver Anschutz Medical Campus, Aurora, CO; University of Colorado Denver Anschutz Medical Campus, Aurora, CO MicroRNAs (miRNAs) are involved in many aspects of cancer biology, including tumor progression, chemoresistance, and metastasis. In breast cancer specifically, miRNAs are more abundant in well differentiated cancer cells, and many are downregulated or lost in more de-differentiated cancer cells. Luminal breast cancers are estrogen receptor (ER) and progesterone receptor (PR) positive. These cancers are hormone dependent and require estrogens for growth; the role of progesterone is less clear. Progesterone is known to rapidly downregulate the expression of several miRNAs in breast cancer cell lines. This includes miR-141, a member of the miR-200 family, of which miR-200c is a well-known tumor suppressor. Interestingly, of the miR-200 family, only miR-141 was found to be significantly hormone regulated in breast cancer cells, and is also reported to be downregulated in breast cancer stem cells (CSCs). Therefore, we hypothesize that loss of miR-141 plays a role in triggering expansion of the CSC population in breast tumors.[br]Preliminary studies of stable inhibition of miR-141 showed significant upregulation of the CSC marker CK5/6 mRNA and an expansion of CK5/6+ cells with progestin treatment. To investigate the role of miR-141 in hormone dependent breast cancer, we have measured levels of miR-141 and family in multiple luminal and basal breast cancer cell lines and in CK5/6+ and CK5/6- cells. We have also created cells with stable knockdown and overexpression of miR-141 to measure mRNA and protein levels of basal and luminal markers. We have validated potential miR-141 targets, including CK6, using luciferase reporter assays and miR-141 mimics to show direct targeting of miR-141 to their 3[apos]-UTRs. Finally, we have measured miR-141 levels in fractionated CSC and non-CSC populations and show that miR-141 specifically downregulated in the CSC fraction.[br]These data support a role for miR-141 in potentiating the transition from luminal cancer cells to CSCs. This is important as a specific role of miR-141 has not yet been described in the maintenance of differentiated phenotype, as it has for other family members. Defining the molecular mechanisms that promote breast CSCs will help design better strategies to prevent or target these cells in tumors.[br][br]Nothing to Disclose: JF-S, DMC, PH, PRP, DMC, BMJ, JKR, CAS 2012-06-23T12:15:00 342ABDE 2012-06-23T00:00:00 1899-12-30T12:15:00 1992 38 84 OR05-5 OR42-01 Saturday 29 2012


30 ENDO12L_OR05-6 ORAL SESSION: Hormone Dependent Tumors (11:15 AM-12:45 PM) Regulation of Type 1 CRH-R Splicing and MCF-7 Cell Responsiveness to CRH by Estrogens: Implications for Breast Cancer Tumor Biology Suchita Lal, Anna Allan, Dimitris Grammatopoulos University of Warwick, Coventry, UK Corticotropin-releasing hormone (CRH), the 41 amino acid hypothalamic peptide that plays a fundamental role in adaptation to stressful stimuli, is emerging as an important regulator of breast cancer cell biology. Previous studies supported a positive effect of CRH on tumour growth by showing increased proliferation, migration and actin polymerization in the 4T1 mouse mammary tumour cell line (1,2). This proposed tumor-promoting action of CRH has been challenged by other studies suggesting an anti-neoplastic potential of CRH through anti-proliferative and differentiation-inducing effects (3,4). Moreover, the use of corticorelin acetate (CrA), a synthetic CRH analog in the preclinical model of MX-1 breast carcinoma, demonstrated substantial antitumor activity, by exerting antiangiogenic effects (5). To understand the role of CRH in breast cancer biology, we used the oestrogen-responsive MCF-7 cell line and we identified a complex signalling network activated by type 1 CRH receptors (CRH-R1) involving activation of pro-survival and migration signalling molecules such as Akt, Hsp27 and p38 MAPK, and Focal Adhesion kinase (FAK) as well GSK3[beta]. Phosphorylation of the latter at Tyr216 was associated with a 50% decrease in [beta]-catenin protein levels, thus suggesting a negative effect on malignant cellular pathways. Interestingly, the signalling potency of CRH was reduced when cells were pre-treated with 10 nM 17[beta]-estradiol (E2). We investigated the profile of CRH-R1 mRNA variants and we identified significant increase by 4-fold of mRNA transcripts of the signaling impaired exon 12-missing variant CRH-R1d, in response to E2 treatment. This was associated with an increase in cytoplasmic CRH-R1 immunoreactivity, a characteristic of CRH-R1d protein expression. In silico studies identified the Serine/Arginine-rich splicing factor 6 (SRp55) as a potential regulator of exon 12 retention in the CRH-R1 mRNA transcript. Indeed, E2 treatment significantly attenuated SRp55 mRNA and protein expression in MCF-7 cells. Attenuation of SRp55 expression by either siRNA or E2-treatment led to a significant increase in CRH-R1d mRNA. These results suggest that CRH actions in MCF-7 cells are sensitive to estrogen-driven pathways that down-regulate expression of SRp55. This signaling modification drives CRHR1 gene splicing mechanisms towards generation of the signaling impaired CRH-R1d and would result in dampened responses to CRH.[br][br]1.Androulidaki A et al. Mol Cancer 2009; 8: 30. 2.Arranz A et al. Mol Cancer 2010; 9: 261. 3. Tjuvajev J et al In Vivo. 1998; 12:1-10. 4. Graziani G et al Mol Cell Endocrinol. 2007 264:44-94. 5. Gamez I et al. 2011; 67: 1415-22.[br][br]Sources of Research Support: The Pathological Society of Great Britain.[br][br]Nothing to Disclose: SL, AA, DG 2012-06-23T12:30:00 342ABDE 2012-06-23T00:00:00 1899-12-30T12:30:00 1271 38 85 OR05-6 OR42-01 Saturday 30 2012


31 ENDO12L_OR06-1 ORAL SESSION: Benign Thyroid Disease (11:15 AM-12:45 PM) Nutrient Restriction during Critical Illness Accelerates Recovery While Further Aggravating the Low T3 Syndrome: A Randomized Controlled Clinical Study Lies Langouche, Sarah Vander Perre, Mirna Bastos Marques, Anita Boelen, Michael Casaer, Greet Van den Berghe KU Leuven, Leuven, Belgium; Academic Medical Center, Amsterdam, Netherlands Critically ill patients reveal low circulating T3 and/or T4 levels in the absence of a rise in TSH. This constellation is labeled [apos]low T3 syndrome[apos] or [apos]nonthyroidal illness syndrome (NTI)[apos]. Interestingly, in healthy subjects, a low T3 syndrome is evoked by fasting in which it presumably mediates adaptation to reduced macronutrient availability. The contribution of restricted nutrition during human critical illness in bringing about the NTI currently remains unclear.[br]We recently performed a large randomized, controlled, multicenter trial, in which we compared early initiation of parenteral nutrition to supplement insufficient enteral feeding ([apos]early PN[apos] group) with tolerating pronounced nutritional deficit during the first week in ICU ([apos]late PN[apos] group). Tolerating the nutritional deficit resulted in faster recovery from organ failure and fewer complications, as compared with early supplementation with parenteral feeding (1). The aim of the current study was to investigate the impact of early nutritional deficit versus early feeding on the NTI of critical illness, in relation to outcome.[br]Of a subgroup of patients in this trial, identifiable upon ICU admission, for whom early enteral nutrition was surgically contraindicated and for whom the study actually compared early PN with full fasting for the first week in ICU, we randomly selected 280 patients for this analysis. In the [apos]early PN[apos] group (n=140), parenteral nutrition was initiated within 48 hours after ICU admission, whereas in the [apos]late PN[apos] group (N=140), parenteral nutrition was only initiated on day 8 when still in ICU. Serum isolated from blood samples collected on admission, on ICU day 3 and 5 and on the last day in ICU were quantified for circulating levels of TT3, TT4 and leptin using commercial assays.[br]The decrease over time in TT3 was significantly blunted by early PN, whereas the decrease in TT4 was only transiently affected. These changes coincided with a pronounced increase in circulating leptin over time, although the circulating levels of leptin did not correlate with circulating thyroid hormone levels. Analyses for circulating rT3 and TSH are ongoing and will be presented. Also, further analyses will assess the extent to which the changes in thyroid hormones, brought about by the nutritional management, explain the clinical benefits of nutrient restriction.[br][br](1) Casaer M et al., 2011 NEJM; 365:506.[br][br]Nothing to Disclose: LL, SVP, MBM, AB, MC, GVdB 2012-06-23T11:15:00 Theater B 2012-06-23T00:00:00 1899-12-30T11:15:00 663 39 86 OR06-1 OR39-01 Saturday 31 2012


32 ENDO12L_OR06-2 ORAL SESSION: Benign Thyroid Disease (11:15 AM-12:45 PM) Hypothyroidism in Critically Ill Patients: Outcomes and Effects of Thyroid Hormone Replacement Katayoun Edalat Parsi, Marianne Ghobrial, Ivance Pugoy, Vijay Balasubramanian, Soe Naing UCSF Fresno Center for Medical Education and Research, Fresno, CA; Fresno Medical Education Program, University of California, San Francisco, Fresno, CA; Fresno Medical Education Program, University of California, San Francisco, Fresno, CA BACKGROUND:[br]Hypothyroidism has been associated with increased mortality in critically ill patients. However, there is a paucity of literature evaluating impact of thyroid hormone replacement (THR) on the mortality rate (MR) in critically ill patients with hypothyroidism.[br]AIM: To evaluate the outcomes and effects of THR in critically ill patients with hypothyroidism in medical intensive care unit (MICU).[br]STUDY DESIGN:[br]This is a retrospective study of the patients admitted to the MICU at a community hospital from 2004 to 2010. Hypothyroidism was defined as admitting thyroid stimulating hormone (TSH) of [gt]30mU/L. The patients were divided into 3 groups based upon total levothyroxine (LT4) dose received during first week of MICU stay: [lt]500 mcg (low dose group or LD), 500-1000mcg (medium dose group or MD) and [gt]1000mcg (high dose group or HD).[br]RESULTS:[br]We studied a total of 36 patients (mean age 68.1years (SD[plusmn]17.7), men 47.2%, BMI 28.2 (SD[plusmn]9.6), mean TSH 92.5mU/L (SD[plusmn]95.4), Hispanics 50%, known primary hypothyroidism 92%, mean total LT4 dose received during the first week of ICU stay 846mcg (SD[plusmn]425), total hospital MR 31%). There were 8 patients in LD, 13 in MD and 15 in HD. None of the patients received liothyronine (T3). There was no statistically significant difference in mean age (68.8 vs 68.1 vs 67.7 years), gender (men 38 vs 62 vs 46%), mean BMI (29.7 vs 25.8 vs 29.6), mean TSH (105 vs 71 vs 104;NR 0.35-5.5 mU/L), mean FT4 (0.6 vs 0.6 vs 0.7;NR 0.89-1.76 ng/dl), mean APACHE II score (22.3 vs 19.1 vs 19.4), mean Glasgow Coma Scale (8.6 vs 12.7 vs 13.2), portions of the patients who received intravenous LT4 (25% vs 38% vs 40%) among 3 groups (LD vs MD vs HD). However the differences in mean total LT4 dose (272 vs 725 vs 1260 mcg) were statistically significant (P[lt]0.0001). Overall, total hospital MR was 50%, 38% and 13% in LD, MD and HD, respectively. MICU length of stay was 9.1, 9.2 and 15.6 days and total hospital stay was 15.8, 22.8 and 27.9 days in LD, MD and HD, respectively.[br]CONCLUSIONS:[br]1. Total hospital MR was 31% in critically ill patients with hypothyroidism who had admission TSH of [gt]30.[br]2. MR was inversely proportional to the total dose of THR received during the first week of MICU stay.[br]3. Aggressive THR during the first week of MICU stay in critically ill hypothyroid patients may improve survival.[br]Further studies with larger number of cases are needed to confirm the findings.[br][br]Nothing to Disclose: KEP, MG, IP, VB, SN 2012-06-23T11:30:00 Theater B 2012-06-23T00:00:00 1899-12-30T11:30:00 682 39 87 OR06-2 OR39-01 Saturday 32 2012


33 ENDO12L_OR06-3 ORAL SESSION: Benign Thyroid Disease (11:15 AM-12:45 PM) Cardiovascular Morbidity after Surgical Treatment of Hyperthyroidism [mdash] A Nationwide Cohort Study with a Long-Term Follow-Up Essi Ryodi, Jorma Salmi, Matti Valimaki, Anssi Auvinen, Pia Jaatinen, Heini Huhtala, Rauni Saaristo, Saara Metso Tampere University Hospital, Tampere, Finland; Tampere University Hospital, Tampere, Finland; Helsinki University Central Hospital, Helsinki, Finland; University of Tampere, Tampere, Finland; University of Tampere, Tampere, Finland; Tampere University Hospital, Tampere, Finland [bold]OBJECTIVE[/bold] Previous studies suggest that hyperthyroid patients remain at an increased risk of cardiovascular morbidity after restoring euthyroidism. The mechanisms of the increased risk and its dependency on different treatment modalities of hyperthyroidism remain unclear. The aim of this long-term follow-up study was to compare the cardiovascular causes of hospitalization in hyperthyroid patients treated with thyroidectomy with those of an age- and gender-matched reference population.[br][bold]PATIENTS AND MEASUREMENTS[/bold] A population-based cohort study was conducted among the 4334 hyperthyroid patients treated with thyroidectomy between years 1986 and 2007 in Finland and among 12,991 reference subjects. Information on hospitalizations was obtained from the nationwide Hospital Discharge Registry. New events were analyzed as the main outcome, including only the first hospitalization due to the given indication.[br][bold]RESULTS[/bold] The median follow-up time of the patients was 10.5 years. The hospitalization rate due to cardiovascular diseases was higher among the patients with hyperthyroidism than in the control population (240.5 vs 206.2 per 10 000 patient-years, rate ratio (RR) 1.17, 95% CI 1.09-1.26). The risk was sustained up to two decades after thyroidectomy. Hospitalizations due to arrhythmias (RR 1.50), hypertension (RR 1.27), and other cardiovascular diseases (RR 1.54) including non-bacterial endo-, peri- and myocardial diseases, valvular diseases, and cardiomyopathy were significantly more frequent among the patients than the controls. No significant difference was found between the patients and controls for coronary artery disease (RR 1.04), cerebrovascular diseases (RR 1.12) and diseases of other arteries and veins (RR 1.11) or heart failure (RR 1.18).[br][bold]CONCLUSIONS[/bold] In line with previous findings on patients treated with radioactive iodine, the current study shows that also hyperthyroid patients treated with thyroidectomy have an increased risk of hospitalization due to cardiovascular diseases. The risk is sustained up to two decades after surgery, which is regarded as an effective treatment modality for hyperthyroidism.[br][br]Sources of Research Support: This work was supported by grants from the Research Funding of the Pirkanmaa Hospital District.[br][br]Nothing to Disclose: ER, JS, MV, AA, PJ, HH, RS, SM 2012-06-23T11:45:00 Theater B 2012-06-23T00:00:00 1899-12-30T11:45:00 673 39 88 OR06-3 OR39-01 Saturday 33 2012


34 ENDO12L_OR06-4 ORAL SESSION: Benign Thyroid Disease (11:15 AM-12:45 PM) Age-Related Modifications of the Relationship between Thyroid Status and Lipoprotein Profile: Results from a Large Cross-Sectional Study Sara Tognini, Antonio Polini, Giuseppe Pasqualetti, Silvia Cottone, Silvia Ursino, Fabio Monzani University of Pisa, Pisa, Italy Background: The relationship between serum lipids and subclinical hypothyroidism is controversial. We evaluated the possible influence of age and gender in a large cohort of patients attending a thyroid outpatient clinic.[br]Patients [amp] Methods: 2308 consecutive patients (1874 F, mean age 47[plusmn]4.1 years) were submitted to complete clinical workup. Thyroid status and lipoprotein profile were assessed after an overnight fast. Study participants were divided into subgroups according to age (1st: 10-29, 2nd: 30-49, 3rd: 50-64, 4th: [gt]65 years) and serum TSH values (1st: [lt]0.36; 2nd: [gt]0.36 and [lt]3.6; 3rd: [gt]3.6 and [lt]10.0; 4th: [gt]10.0 mIU/l).[br]Results: Males and females were matched for age and blood pressure while body mass index was slightly higher in men (p=0.05). The prevalence of autoimmunity was significantly higher in women than men (55.4% Vs 39.9%, p=0.0002) but no gender difference in TSH frequency distribution was observed. Total (TC) and LDL cholesterol (LDLc) values (p[lt]0.0003 and p[lt]0.003, respectively) as well as LDL/HDLc ratio (p[lt]0.03) were significantly elevated in unselected women of the 4th TSH group ([gt]10 mIU/l) but, while analyzing women of different age groups, also those of the 3rd TSH group older than 50 years showed an atherogenic lipid profile (TC and LDLc p=0.01 and LDL/HDLc ratio p=0.02 Vs euthyroid women). Among unselected men only those of the 4th TSH group had significantly elevated triglyceride (TG) (p[lt]0.0001) but not cholesterol values. However, men of the 3rd and 4th TSH groups older than 65 years showed significantly higher TC, LDLc values and LDL/HDLc ratio too (p=0.03, p=0.02 and p=0.01, respectively Vs euthyroid men). In the final model of stepwise regression analysis, which explained almost 40% of serum TC and LDLc variations, age had the highest standardized coefficient (0.36 and 0.37, respectively), followed by TSH (0.20 and 0.11, respectively) and FT4 (-0.11 and [ndash]0.09, respectively).[br]Conclusions: The current study, while confirming a gender related differentiation in the relationship between thyroid function and lipid profile, firstly documents that the impact of mild thyroid failure on serum lipids is substantially influenced by age in both genders. Overall, our data delineate a definite interrelationship between thyroid function, age and gender and may, at least partially, explain the conflicting results of the literature regarding the impact of slightly elevated TSH value on circulating lipid parameters.[br][br]Nothing to Disclose: ST, AP, GP, SC, SU, FM 2012-06-23T12:00:00 Theater B 2012-06-23T00:00:00 1899-12-30T12:00:00 1075 39 89 OR06-4 OR39-01 Saturday 34 2012


35 ENDO12L_OR06-5 ORAL SESSION: Benign Thyroid Disease (11:15 AM-12:45 PM) Subclinical Thyroid Dysfunction Is Not Associated with Risk of Cognitive Decline or Dementia in Older Adults Eve Bloomgarden, Alice M Arnold, Michelle Carlson, Lewis Kuller, Oscar Lopez, Anne R Cappola University of Pennsylvania, Philadelphia, PA; University of Washington, Seattle, WA; Johns Hopkins University, Bloomberg School of Public Health, Baltimore, MD; University of Pittsburgh, Pittsburgh, PA; University of Pittsburgh, Pittsburgh, PA [bold]Background:[/bold] Subclinical thyroid dysfunction is common in older individuals and has been linked to neurocognitive impairment. We assessed the risks of cognitive decline and dementia in older adults with subclinical thyroid dysfunction.[br][bold]Methods:[/bold] Annual Modified Mini Mental Examination (3MSE), Digit Symbol Substitution Test (DSST) scores and incident dementia were analyzed from US community-dwelling men and women aged [ge]65 y in the Cardiovascular Health Study (CHS) who were not taking thyroid hormone preparations. Generalized estimating equations were used to estimate the difference in rate of change in 3MSE and DSST scores comparing subclinical hypothyroid (Shypo; n=624) and subclinical hyperthyroid (Shyper; n=74) groups to the euthyroid group (n=3894). Informative missingness was assessed in sensitivity analyses using weights defined by the inverse probability of dropping out. Cox proportional hazard models were used to examine the relationship between baseline thyroid status and incident dementia. All models were adjusted for age, sex, race, and education. Analyses were repeated in a subset of participants who had a second set of thyroid measurements to define persistent, transient, or progressive Shypo and Shyper.[br][bold]Results:[/bold] There was no difference in rate of change of 3MSE or DSST score by baseline or persistent thyroid category, although there was a suggestion of greater decline in DSST scores in the baseline Shyper group compared to the euthyroid group (-0.79 vs. -0.47; p=0.06), which was confirmed in the sensitivity analysis (-0.94 vs. -0.44; p=0.004). In the small group of Shypo participants who progressed to overt hypothyroidism, a greater decline in DSST was observed, approaching statistical significance (-1.96 vs. -0.85; p=0.07). There were 402 cases (339 euthyroid, 58 Shypo, 5 Shyper) of incident dementia over a mean follow-up of 5.4 years. We found no association of thyroid status with incident dementia. The hazard ratio was 1.06 (0.80- 1.40) for the Shypo group and 1.32 (0.54-3.19) for the Shyper group. Findings were similar in models of persistence.[br][bold]Conclusions: [/bold]We found no difference in cognitive decline or development of dementia in older individuals with baseline or persistent Shypo. There may be a greater decline in those who progress to overt hypothyroidism and in those with Shyper. We find no evidence for a clinically significant impact of subclinical thyroid dysfunction on cognitive function in this population.[br][br]Sources of Research Support: R01 AG032317 to ARC.[br][br]Nothing to Disclose: EB, AMA, MC, LK, OL, ARC 2012-06-23T12:15:00 Theater B 2012-06-23T00:00:00 1899-12-30T12:15:00 657 39 90 OR06-5 OR39-01 Saturday 35 2012


36 ENDO12L_OR06-6 ORAL SESSION: Benign Thyroid Disease (11:15 AM-12:45 PM) Women with High Early Pregnancy Urinary Iodine Levels Are at Risk of Hyperthyroid Newborns: The Generation R Study Marco Medici, Akhgar Ghassabian, Willy Visser, Sabine MPF de Muinck Keizer-Schrama, Herbert Hooijkaas, Jacoba J Bongers-Schokking, Henning Tiemeier, Alec Ross, Theo J Visser, Yolanda B de Rijke, Robin P Peeters Erasmus Medical Center, Rotterdam, Netherlands; Erasmus Medical Center [ndash] Sophia Children[apos]s Hospital, Rotterdam, Netherlands; Erasmus Medical Center [ndash] Sophia Children[apos]s Hospital, Rotterdam, Netherlands; Erasmus Medical Center [ndash] Sophia Children[apos]s Hospital, Rotterdam, Netherlands; Erasmus Medical Center, Rotterdam, Netherlands; Erasmus Medical Center, Rotterdam, Netherlands; Radboud University Nijmegen Medical Centre, Nijmegen, Netherlands Introduction:[br]Both iodine deficiency and iodine excess can lead to thyroid dysfunction. During pregnancy, iodine requirements increase and maternal iodine deficiency can result in developmental brain damage. Most studies on iodine and pregnancy focus on regions with (mild) iodine deficiency. As iodine intake is highly variable within populations, it is remarkable to note that little data on the effects of variation in iodine status on maternal and fetal thyroid function are available from iodine-sufficient populations.[br]Materials and methods:[br]Urinary iodine, serum TSH, FT4 and TPO-antibody (TPOAb) levels were determined in early pregnancy (13.3 (1.7) wk) in 1154 pregnant women from the Generation R Study, a population-based cohort study in Rotterdam, The Netherlands, which is considered an iodine-sufficient area. Cord serum TSH and FT4 levels were available in 1068 newborns. Informed consent was obtained from all participants.[br]Results:[br]The median urinary iodine level was 222.5 [mu]g/L (range: 9.3 [ndash] 1743.5 [mu]g/L), indicating an iodine-sufficient population (1). 30.8% and 11.5% had urinary iodine levels [lt] 150 and [gt] 500 [mu]g/L, respectively.[br]When comparing mothers with urinary iodine levels [lt] 150 vs [ge] 150 [mu]g/L, and [gt] 500 vs [le] 500 [mu]g/L, there were no differences in the risk of maternal hypothyroxinemia, (subclinical) hypo- or hyperthyroidism. However, mothers with urinary iodine levels [gt] 500 [mu]g/L had a 2.7 times increased risk (5.6[plusmn]1.4 vs 2.1[plusmn]0.5 %, [italic]P[/italic] = 0.035) of a newborn with suppressed cord TSH levels (TSH [lt] 3.10 mU/L ([lt] 2.5[sup]th[/sup] percentile)), as well as a 5.2 times increased risk (3.1[plusmn]0.9 vs 0.6[plusmn]0.3 %, [italic]P[/italic] = 0.016) of a hyperthyroid newborn (TSH [lt] 3.10 mU/L ([lt] 2.5[sup]th[/sup] percentile) and FT4 [gt] 30.1 pmol/L ([gt] 97.5[sup]th[/sup] percentile)). These mothers had newborns with higher cord FT4 levels (21.7[plusmn]0.3 vs 21.0[plusmn]0.1 pmol/L, [italic]P[/italic] = 0.036).[br]Maternal urinary iodine levels [lt] 150 [mu]g/L were not associated with newborn thyroid dysfunction.[br]Conclusion:[br]In an iodine-sufficient population, women with higher urinary iodine levels have a substantial increased risk of a hyperthyroid newborn. In contrast, women with lowest urinary iodine levels did not have a higher TSH. These data provide insight into the effects of variation in maternal iodine status on maternal and fetal thyroid dysfunction.[br][br](1) World Health Organization United Nations Children[apos]s Fund, International Council for the Control of Iodine Deficiency Disorders. 2007 Assessment of Iodine Deficiency Disorders and Monitoring Their Elimination. 3rd ed. Geneva: WHO.[br][br]Nothing to Disclose: MM, AG, WV, SMPFdMK-S, HH, JJB-S, HT, AR, TJV, YBdR, RPP 2012-06-23T12:30:00 Theater B 2012-06-23T00:00:00 1899-12-30T12:30:00 826 39 91 OR06-6 OR39-01 Saturday 36 2012


1 ENDO12L_OR01-1 ORAL SESSION: Endocrine Gene Regulatory Circuits (11:15:00 AM-12:45:00 PM) A Pitx2-miRNA-200c-Noggin Regulatory Axis Regulates BMP Signaling and Anterior Pituitary Cell Type Specification Huojun Cao, Xiao Li, Jianbo Wang, Zhao Sun, Zichao Zhang, Brad A Amendt Texas A[amp]M Health Science Center, Institute of Biosciences and Technology, Houston, TX The microRNA-200 family plays a critical role in cancer cell progression and regulation of the epithelial-to-mesenchymal transition during embryonic development and cancer. We previously reported a role for mature miRNAs in pituitary development, hormone levels and gene expression. However, the specific miRNAs that regulate pituitary cell types remains unknown. The miRNA-200 family is specifically up-regulated during pituitary development and in gondadotropes, somatotropes and lactotropes, but not in corticotropes. The miRNA-200 family expression increases with differentiation. BMP signaling is required for invagination of Rathke[apos]s pouch and over-expression of Noggin (a BMP2/4 antagonist) inhibits pituitary cell lineage differentiation. BMP4 signaling and therefore repressed Noggin expression is required for the continued progression of pituitary development. We demonstrate that miRNA-200c targets Noggin and allows for BMP signaling to regulate gene expression and differentiation of pituitary lineage cells. Interestingly, miRNA-200c and 200b expression are increased in PITX2C over expression mouse epithelial tissues. Chromatin immunoprecipiation assays reveal endogenous PITX2 binding to the miRNA-200c-141 promoter. Transfection of PITX2 in cells activates endogenous miRNA-200c and the miRNA-200c enhancer reporter. The Pitx2 transcription factor regulates early pituitary developmental programs and we demonstrate Pitx2 regulates the miRNA-200 family during development. The activation of miRNA-200c indirectly regulates BMP signaling by targeting and repressing Noggin expression. This process allows for pituitary progenitor cells to become differentiated cells of the anterior pituitary. These studies reveal a multistep transcriptional program involving a Pitx2-miRNA200-Noggin axis during pituitary cell differentiation.[br][br]Sources of Research Support: National Institute of Dental and Craniofacial Research grants DE13941 and DE018885 awarded to BAA.[br][br]Nothing to Disclose: HC, XL, JW, ZS, ZZ, BAA 2012-06-23T11:15:00 320 2012-06-23T00:00:00 1899-12-30T11:15:00 2137 34 56 OR01-1 OR16-01 Saturday 1 2012


2 ENDO12L_OR01-2 ORAL SESSION: Endocrine Gene Regulatory Circuits (11:15:00 AM-12:45:00 PM) Evidence That microRNA-29 Downregulates Multiple Growth-Promoting Genes and Thus Contributes to Juvenile Growth Deceleration Fariha Kamran, Anenisia C Andrade, Samuel Clokie, Geoffrey A Rezvani, Ola Nilsson, Jeffrey Baron, Julian C Lui National Institute of Child Health and Human Development/National Institutes of Health, Bethesda, MD; Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden; St Christopher[apos]s Hospital for Children, Philadelphia, PA Infancy is characterized by rapid body growth, which slows with increasing age and eventually ceases due to a progressive decline in cell proliferation that occurs simultaneously in multiple organs. We previously showed evidence that this decline in proliferation is driven in part by postnatal downregulation of a large set of growth-promoting genes in multiple organs (Endocrinology 150:1791-800, 2009; FASEB J 24:3083-92, 2010). We hypothesized that this growth-limiting genetic program is orchestrated by microRNAs, which are short non-coding RNAs that bind to target sequences in multiple mRNAs to inhibit translation or stimulate mRNA degradation. A bioinformatic algorithm (Partek Genomics Suite 6.6) was used to identify microRNA target sequences in genes that are commonly downregulated in mouse kidney and lung as growth slows from 1 to 4 weeks of age. The analysis predicted that the downregulated genes are strongly enriched with target sequences for microRNA-29 family members (miR-29a, b, and c, P [lt] 0.00001 each). Concomitantly, we analyzed changes in microRNA expression in juvenile mice as growth slows with age, from 1 to 6 weeks by microarray (Agilent Mouse Genome VI). miR-29a, b, and c were strongly upregulated with age in both kidney and lung (all [gt]8-fold increase, false discovery rate [lt] 0.05). Real-time PCR confirmed significant (P [lt]0.05) upregulation with age, from 1 to 4 weeks, of miR-29a (lung, 15-fold; kidney, 27-fold), miR-29b (lung, 9-fold; kidney, 32-fold) and miR 29c (lung, 16-fold; kidney, 58-fold). We next focused on 3 age-downregulated genes that are predicted targets of miR-29: Igf-1, Igf2bp1, and Mest. The 3[apos]-UTR of each gene was placed downstream of a luciferase reporter and transfected into HEK293 cells. Co-transfection with miR-29a, b or c mimics suppressed luciferase activity, confirming that the mRNAs are actual targets of miR-29. In summary: (i) miR-29a, b and c are strongly upregulated with age in multiple organs during juvenile life, (ii) miR-29 target sequences are strongly overrepresented in genes that are downregulated with age in multiple organs and (iii) some of these predicted targets have been experimentally confirmed. Taken together, the findings suggest that upregulation of miR-29a, b, and c during juvenile life downregulates multiple growth-promoting genes, thus contributing to physiological slowing and eventual cessation of body growth.[br][br]Nothing to Disclose: FK, ACA, SC, GAR, ON, JB, JCL 2012-06-23T11:30:00 320 2012-06-23T00:00:00 1899-12-30T11:30:00 1392 34 57 OR01-2 OR16-01 Saturday 2 2012


3 ENDO12L_OR01-3 ORAL SESSION: Endocrine Gene Regulatory Circuits (11:15:00 AM-12:45:00 PM) [italic]ARTD1[/italic] Deletion Causes Increased Hepatic Lipid Accumulation in Mice on High-Fat Diet and Impairs Adipocyte Function and Differentiation Suheda Erener, Ali Mirsaidi, Mareike Hesse, Andre N Tiaden, Helga Ellingsgaard, Radina Kostadinova, Marc Y Donath, Peter J Richards, Michael O Hottiger University of Zurich, Zurich, Switzerland; University of Zurich, Zurich, Switzerland; University Hospital Basel, Basel, Switzerland ADP-ribosyltransferase Diphtheria toxin-like 1 (ARTD1) is a chromatin-associated enzyme involved in regulating metabolic homeostasis. We provided earlier evidence that ARTD1-dependent ADP-ribose polymer (PAR) formation increases during adipocyte development and, at late time points of adipogenesis, is involved in the sustained expression of [italic]PPAR[gamma]2[/italic] and of PPAR[gamma]2 target genes. During adipogenesis, ARTD1 is recruited to [italic]PPAR[gamma]2[/italic] target genes such as [italic]CD36[/italic] or [italic]aP2[/italic] in a PAR-dependent manner. Interestingly, activation of ARTD1 enzymatic activity is prevented with a Toposiomerase II inhibitor (1).[br]The liver is at the core of glucose and lipid metabolism and significantly affected by obesity and the metabolic syndrome. Here, we will show that on high fat diet (HFD), mice lacking [italic]ARTD1[/italic] developed exacerbated hepatic steatosis. [italic]ARTD1[/italic][sup]-/-[/sup] mice had a 19% higher liver weight than wildtype (WT) animals and also exhibited an increased serum concentration of free fatty acids (42% increase) and cholesterol (38% increase) and impaired glucose tolerance. In addition, adipocyte function and size were significantly reduced in [italic]ARTD1 [/italic][sup]-/-[/sup] mice on HFD (7794 [mu]m[sup]2[/sup] for WT and 5579 [mu]m[sup]2[/sup] for [italic]ARTD1[/italic][sup]-/-[/sup] mice). The significantly reduced adipogenic differentiation of adipose-derived stromal cells (ASCs) isolated from [italic]ARTD1 [/italic][sup]-/-[/sup] mice (28% vs. 11% Oil red O positive cells in WT and [italic]ARTD1 [/italic][sup]-/-[/sup] ASCs, respectively) suggested adipogenesis as the underlying cause for this adipose tissue malfunction. This function of ARTD1 was specific for adipogenesis since osteogenic differentiation was not affected by the [italic]ARTD1[/italic] deletion. Our data suggest a novel role for ARTD1 in the development of hepatic steatosis as a consequence of impaired adipogenesis and adipocyte malfunction, which can subsequently lead to non-alcoholic fatty liver disease.[br][br](1) Erener S et al., Mol Endocrinol. 2012; 26(1):79-86.[br][br]Sources of Research Support: This work was supported in part by the University Research Priority Program [ldquo]Integrative Human Physiology[rdquo] at the University of Zurich, the Swiss National Science Foundation Grant 31-122421 and 31-138667 and the Kanton of Zurich (to M.O.H.).[br][br]Nothing to Disclose: SE, AM, MH, ANT, HE, RK, MYD, PJR, MOH 2012-06-23T11:45:00 320 2012-06-23T00:00:00 1899-12-30T11:45:00 1436 34 58 OR01-3 OR16-01 Saturday 3 2012


4 ENDO12L_OR01-4 ORAL SESSION: Endocrine Gene Regulatory Circuits (11:15:00 AM-12:45:00 PM) The miR-200 Family and ZEB Transcription Factors Serve Important Roles in Developmental Induction of Surfactant Protein-A (SP-A) Gene Expression in Human Fetal Lung Houda Benlhabib, Carole R Mendelson University of Texas Southwestern Medical Center, Dallas, TX Differentiation of the fetal lung and development of the capacity to synthesize pulmonary surfactant are essential for transition of the fetus from an aqueous to an air-breathing environment. In recent years, considerable progress has been made in our understanding of the transcriptional mechanisms for differentiation of the developing lung and expression of genes encoding the surfactant proteins. To further define mechanisms for type II cell differentiation and developmental induction of surfactant synthesis, we are investigating the potential role of microRNAs (miRNAs, miRs). miRNAs are evolutionarily conserved [sim]22 nucleotide, single-stranded noncoding RNAs that, inhibit gene expression by binding to the 3[apos]-untranslated regions of their target mRNAs through base-pairing, resulting in inhibition of mRNA translation and/or increased mRNA degradation. Previously, we observed that midgestation human fetal lung explants differentiate spontaneously in organ culture in serum-free medium. The rate of type II cell differentiation and expression of the gene encoding the major surfactant protein, SP-A, are markedly induced by cAMP. To identify and characterize differentially regulated miRNAs in mid-gestation human fetal lung explants upon type II pneumocyte differentiation in culture, we performed miRNA microarray analysis of RNA isolated from epithelial cells from human fetal lung explants before and after 48 and 96 h of culture [plusmn] Bt[sub]2[/sub]cAMP. Interestingly, the miR-200 family was found to be significantly upregulated with type II cell differentiation. Moreover, the induction of miR-200 family members was inversely correlated with expression of their known targets, transcription factors ZEB1 and ZEB2. Interestingly, overexpression of ZEB1 or ZEB2 in cultured type II cells using recombinant adenoviruses blocked cAMP stimulation of SP-A expression. On the other hand, cAMP, which promotes type II cell differentiation, caused a marked inhibition of ZEB expression. Notably, culture of human fetal lung epithelial cells with active TGF-[beta]1, known to induce epithelial to mesenchymal transition in epithelial cells and to inhibit type II cell differentiation, induced expression of ZEBs and inhibited SP-A expression. These findings suggest that the miR-200 family and ZEB1/2, which exist in a double-negative feedback loop, serve important regulatory roles in the developmental regulation of type II cell differentiation and SP-A expression in human fetal lung.[br][br]Sources of Research Support: NIH R01 HL050022.[br][br]Nothing to Disclose: HB, CRM 2012-06-23T12:00:00 320 2012-06-23T00:00:00 1899-12-30T12:00:00 1931 34 59 OR01-4 OR16-01 Saturday 4 2012


5 ENDO12L_OR01-5 ORAL SESSION: Endocrine Gene Regulatory Circuits (11:15:00 AM-12:45:00 PM) Influence of Prenatal Steroid Exposure on Insulin Sensitivity and DNA Methylation in Mice Tiffany Sue Musick, Scott A Sands, Jill D Jacobson Children[apos]s Mercy Hospitals and Clinics, Kansas City, MO; Univeristy of Missouri-Kansas City School of Medicine, Kansas City, MO Objective: Stimulatory G proteins play a key role in energy metabolism. Loss of function mutations in the [italic]Gnas[/italic] gene, encoding the guanine nucleotide alpha subunit (G[alpha]s), lead to obesity and insulin resistance. Maternally inherited mutations of the [italic]Gnas[/italic] gene are known to cause obesity in both mice and humans. We have recently found that mice bearing inactivating mutations in the [italic]Gnas [/italic]gene display low estradiol levels during pregnancy. We hypothesized that alterations in the prenatal hormonal milieu would affect later insulin sensitivity, G protein expression, and/or epigenetic phenomenon, specifically DNA methylation.[br]Methods: All experiments were approved by the UMKC Animal Care and Use Committee. Pregnant female mice were treated with 17 [beta]-estradiol (.1 [micro]g/day) on days 16-18 or 5-21, dihydrotestosterone (DHT) (250 [micro]g/day) on days 16-18, or with vehicle consisting of charcoal-stripped, delipidated corn oil on days 5-21. At 6 months of age, we performed intraperitoneal glucose tolerance tests (ipgtt) and obtained insulin and leptin measurements in female mice. DNA, RNA, and protein were purified from hypothalamus and white adipose tissue (WAT). We performed quantitative PCR for G[alpha]s and G[alpha]q mRNA. Global DNA methylation was quantitated by ELISA.[br]Results: Prenatal exposure to estradiol led to a significant decrease in serum insulin levels at the peak time point (60 minute) in the ipgtt test[(mean [plusmn] SEM) 0.1796 [plusmn] 0.0269 vs. 0.4978 [plusmn] 0.0826]. In contrast, prenatal exposure to DHT resulted in significant increases in insulin levels at the same time point (0.8046 [plusmn] 0.1725 vs. 0.4978 [plusmn] 0.0826). Prenatal exposure to estradiol led to increased expression of G[alpha]s mRNA in the hypothalamus (166.4 [plusmn] 14.9 vs. 117.8 [plusmn] 14.2) and WAT (155.3 [plusmn] 15.7 vs. 54 [plusmn] 14.1). G protein mRNA expression in the hypothalamus correlated inversely with log 60 minute glucose measurements (rho = -0.62; p[lt]0.001). Prenatal exposure to estradiol lead to an increase in G[alpha]s mRNA expression in parallel with a decrease in global DNA methylation in the hypothalamus (0.87 [plusmn] 0.07 vs. 1.23 [plusmn] 0.11 % control).[br]Conclusions: Our study may provide insight into possible mechanisms for the effects of sex steroids on later development of insulin resistance. Prenatal exposure to estradiol increased insulin sensitivity and G protein expression in hypothalamus and WAT. Aberrant levels of estrogens or androgens may contribute to altered expression of G[alpha]s and insulin sensitivity in adipose cells.[br][br]Sources of Research Support: Endocrine Fellows Foundation Marilyn Fishman Grant for Diabetes Research Fall 2010.[br][br]Nothing to Disclose: TSM, SAS, JDJ 2012-06-23T12:15:00 320 2012-06-23T00:00:00 1899-12-30T12:15:00 725 34 60 OR01-5 OR16-01 Saturday 5 2012


6 ENDO12L_OR01-6 ORAL SESSION: Endocrine Gene Regulatory Circuits (11:15:00 AM-12:45:00 PM) T-Box 15 Is a Molecular Determinant of Fiber Type Distribution in Skeletal Muscle Kevin Y Lee, Cecile Vernochet, Stephane Gesta, C Ronald Kahn Joslin Diabetes Center, Boston, MA Tbx15 is a member of the T-box gene family that has been shown to have critical roles in mesoderm development and regulating mitochondrial mass and activity. We have previously shown that Tbx15 expression is increased in subcutaneous fat and decreased in the visceral fat of obese humans, and that Tbx15 overexpression in adipocytes in vitro impairs mitochondrial oxidation.[br]Skeletal muscle plays an important role in maintaining normal whole-body glucose metabolism and energy homeostasis, in part, due to its large mass and intrinsic oxidative capacity. In mammals, skeletal muscles are composed of a mosaic of different fiber types, and fiber type distribution correlates with glucose uptake and insulin resistance. Red oxidative fibers (Type I and IIa) are suited for endurance and use oxidative metabolism to generate ATP whereas white glycolytic fibers (Type IIx and IIb) are suitable for bursts of activity and rely on glycolytic metabolism.[br]In the present study, we demonstrate that Tbx15 is most highly expressed in skeletal muscle compared to other tissues. Interestingly, Tbx15 expression is increased in the skeletal muscle of obese humans. Using a Lac-Z reporter construct [italic]in vivo[/italic], we find that Tbx15 is almost exclusively expressed in Type IIb glycolytic muscle fibers. Global knockout of Tbx15 results in severe musculoskeletal developmental malformations. Heterozygous Tbx15 knockout (Tbx15[sup]+/-[/sup]) mice are developmentally normal in appear, but histological examination of the quadriceps from 6 week old animals revealed a 2.5-fold increase in the density of oxidative Type IIa fibers compared to controls and a corresponding decrease in Type IIb fibers. Furthermore, muscle of Tbx15[sup]+/-[/sup] animals displayed a constitutive activation of adenosine monophosphate kinase (AMPK) signaling. Muscles of Tbx15[sup]+/-[/sup] animals also exhibited a decrease in expression of insulin-like growth factor 2 (IGF-2) during embryogenesis. C2C12 myoblast cell lines with a stable knockdown of Tbx15 also exhibited a robust increase in AMPK signaling, and a 70% decrease in the expression of IGF-2. On the other hand, stable over-expression of Tbx15 in C2C12 myoblasts leads to a seven-fold increase in IGF-2 expression.[br]Taken together, these data demonstrate that Tbx15 controls AMPK signal, IGF-2 expression, and fiber type in skeletal muscle. Thus, changes in Tbx15 expression in muscle contributes to regulation of energy metabolism and muscle function.[br][br]Nothing to Disclose: KYL, CV, SG, CRK 2012-06-23T12:30:00 320 2012-06-23T00:00:00 1899-12-30T12:30:00 1635 34 61 OR01-6 OR16-01 Saturday 6 2012


7 ENDO12L_OR02-1 ORAL SESSION: Development [amp] Modulation of Hypothalamic Neuropeptidergic Neurons (11:15 AM-12:45 PM) Speed of GnRH Neuronal Migration Is Controlled by Leading Process Actin and Microtubule Flow Bruce Ian Hutchins, Ulrike Klenke, Susan Wray National Institutes of Health, Bethesda, MD The GnRH system is essential for proper function of the HPG axis. Correct placement of migrating GnRH neurons and their projections within the CNS is essential for reproductive function and occurs early in development. Failure to establish an adult-like distribution can lead to reproductive disorders such as Kallman[apos]s syndrome. Migration from proliferative zones to the proper location in the brain is a challenging task for many neurons. This is especially true for GnRH neurons, which must first enter the brain from the nasal region before beginning their journey through the forebrain. Originating in the nasal placode, GnRH neurons follow olfactory axons into the brain, while extracellular signals regulate movement of GnRH neurons along this pathway. A major unresolved question in the field is how these signals effect the mechanical changes that underlie cellular movement. We hypothesized that dynamic actin/microtubule interactions form an intracellular engine that pulls GnRH neurons forward along their migratory pathway in response to signaling pathways initiated by extracellular cues. To investigate this hypothesis, migrating GnRH neurons were imaged in mouse nasal explants, a powerful in vitro model that recapitulates this stage of embryonic development. GFP-tagged actin was imaged in the leading process of GnRH neurons. Actin flow toward the distal leading process correlated with movement of the cell body. This actin flow could be evoked by pharmacologically stimulating calcium release from stores via phospholipase C, while blocking calcium release through IP3 receptors slowed actin flow and reduced movement of the cell body. A similar behavior was found after examination of the leading process microtubules. Based on these results, we hypothesized that actin flow draws the nucleus up the leading process by pulling forward a [apos]microtubule cage[apos] surrounding the nucleus. Depolymerizing microtubules was found to reduce migration rates, consistent with this hypothesis. Experiments are currently testing whether microtubule movement requires actin dynamics. These results, when taken together, integrate information from signaling pathways to the molecular engines pulling the cell body forward to reveal the precise mechanisms used by cues in the environment to set the speed of migrating GnRH neurons.[br][br]Sources of Research Support: The authors are supported by the Intramural Research Program of the National Institute of Neurological Disorders and Stroke, National Institutes of Health (B.I.H., U.K. and S.W.) and a fellowship from the Pharmacology Research Associate (PRAT) Prorgam, National Institute of General Medical Sciences, National Institutes of Health (B.I.H.).[br][br]Nothing to Disclose: BIH, UK, SW 2012-06-23T11:15:00 371 2012-06-23T00:00:00 1899-12-30T11:15:00 827 35 62 OR02-1 OR20-01 Saturday 7 2012


8 ENDO12L_OR02-2 ORAL SESSION: Development [amp] Modulation of Hypothalamic Neuropeptidergic Neurons (11:15 AM-12:45 PM) Alterations in Hypothalamic Gene Expression of Reproductive Neuropeptides during the Pubertal Transition in Mice Sheila J Semaan, Alexander S Kauffman University of California-San Diego, La Jolla, CA Puberty onset is induced by the secretion of gonadotropin releasing hormone (GnRH). However, the mechanisms controlling and timing puberty onset remain poorly-defined, especially regarding the role of neuropeptides that act upstream of GnRH neurons. In mammals, kisspeptin ([italic]Kiss1[/italic] gene) and neurokinin B (NKB; [italic]Tac2[/italic] gene), stimulate GnRH secretion, while RFRP-3 (the mammalian homologue of GnIH; [italic]Rfrp[/italic] gene) inhibits the GnRH axis. In the rodent hypothalamus, [italic]Kiss1[/italic] is expressed in the anteroventral periventricular nucleus (AVPV) and the arcuate (ARC) nucleus, with neurons in the ARC also co-expressing [italic]Tac2[/italic]. RFRP-3 neurons reside exclusively in the hypothalamic dorsomedial nucleus (DMN). Previous studies reported that [italic]Kiss1[/italic] expression in the brain is higher in adults than prepubertal animals, but the exact peripubertal age(s) when [italic]Kiss1[/italic] first increases is unknown, as is the precise developmental pattern of [italic]Kiss1[/italic] expression specifically in the ARC and AVPV during the pubertal transition. Likewise, the specific developmental pattern of [italic]Tac2[/italic] and [italic]Rfrp[/italic] expression just before and during puberty is not well-characterized. Lastly, the pattern of developmental changes in each of these reproductive neuropeptide systems in relation to one another during puberty has not been determined. Using [italic]in situ[/italic] hybridization, we determined the detailed pubertal developmental profile (from PND 15 to PND 30) of [italic]Kiss1, Tac2[/italic], and [italic]Rfrp[/italic] gene expression in the AVPV, ARC, and DMN, as well as [italic]cfos[/italic] induction in each of these regions, in female mice. The mean age of vaginal opening was postnatal day (PND) 27.5. [italic]Kiss1[/italic] expression in the AVPV of females increased steadily over the pubertal transition, reaching adult levels around PND 28. The largest daily increase was noted on PND 24, several days before the mean age of vaginal opening. In the ARC, however, Kiss1 and Tac2 cell numbers remained fairly steady over the pubertal transition, showing only a very minor increase. [italic]Kiss1[/italic] neurons were not highly activated in the AVPV at any pubertal age, as measured by cfos coexpression. Interestingly, [italic]Rfrp[/italic] expression decreased significantly around PND 21, which may relate to RFRP-3[apos]s possible role as an inhibitor of the GnRH axis prior to puberty. Additional data from current RFRP-3- and Tac2-cfos coexpression assays will provide a more detailed picture of how these key reproductive neuropeptide systems change during the pubertal transition.[br][br]Sources of Research Support: R00 HD056157, R01 HD065856, U54 HD012303, F32 HD066849, and DERC P[amp]F grant DK063491.[br][br]Nothing to Disclose: SJS, ASK 2012-06-23T11:30:00 371 2012-06-23T00:00:00 1899-12-30T11:30:00 532 35 63 OR02-2 OR20-01 Saturday 8 2012


9 ENDO12L_OR02-3 ORAL SESSION: Development [amp] Modulation of Hypothalamic Neuropeptidergic Neurons (11:15 AM-12:45 PM) Intrauterine Vitamin D[sub]3[/sub] Deficiency Disrupts the Female Reproductive Axis Joseph B Davis, Marlena Petti, Thalia Segal, Genevieve Neal-Perry Montefiore Medical Center, Bronx, NY; Albert Einstein College of Medicine, Bronx, NY; North Shore Long Island Jewish Medical Center, Hyde Park, NY Vitamin D3[sub] [/sub](VD3) receptors are localized in the hypothalamic-pituitary-ovarian axis (HPO) suggesting VD3 may be important in reproductive physiology. Consistent with this hypothesisVD3 deficiency is more prevalent in women with reproductive dysfunction. VD3 insufficiency and deficiency has reached near epidemic levels in reproductive aged women, thus raising the possibility that intrauterine and lactational (developmental) VD3 deficiency adversely affects the reproductive physiology of exposed female offspring. [bold]Objective[/bold]: To determine if developmental VD3 deficiency affects puberty or HPO function in mice. [bold]Method[/bold]: Females (n=14-24) born to dams fed a sufficient (control) or deficient VD3 diet during pregnancy and lactation were weaned onto aVD3 sufficient diet. The effect of developmental VD3 deficiency on puberty (vaginal opening and 1st estrus), estrous cycling, ovarian physiology, glucose tolerance, GnRH neuronal density and pituitary responsiveness to GnRH peptide was determined. Data was analyzed with 2-way ANOVA or non-parametric tests. [bold]Results[/bold]: Young adult mice exposed to developmental VD3 deficiency had a normal pubertal transition. However developmental VD3 deficiency resulted in extended estrous cycles (p[lt]0.01) characterized by a prolonged diestrus (p[lt]0.05) and oligoovulation (p[lt]0.05). Developmental VD3 deficiency was associated arrested ovarian follicular development characterized by more early stage primordial (p[lt]0.01) and primary (p[lt]0.01) follicles than controls. Developmental VD3 deficiency was not associated with glucose intolerance. Developmental VD3 deficiency did not affect E2 negative feedback effects on pituitary gonadotropin (LH and FSH) release or did not affect the number of GnRH neurons in the hypothalamus of adult females. However, developmental VD3 deficiency was associated with reduced pituitary FSH (P[lt]0.01) and tended to have less LH release following a GnRH peptide challenge under E2 positive feedback conditions.[bold]Conclusion[/bold]: These data suggest that developmental VD3 deficiency reduces pituitary responsiveness and gonadotropin release in response to GnRH peptide. This most likely disrupts ovarian physiology (arrested follicular development) and results in prolonged estrous cycles characterized by oligoovulation. These preliminary data suggest that maternal VD3 deficiency may adversely affects the reproductive phenotype of exposed adult female offspring through effects on the pituitary function and secondary effects on the ovary.[br][br]Sources of Research Support: HD066355.[br][br]Nothing to Disclose: JBD, MP, TS, GN-P 2012-06-23T11:45:00 371 2012-06-23T00:00:00 1899-12-30T11:45:00 918 35 64 OR02-3 OR20-01 Saturday 9 2012


10 ENDO12L_OR02-4 ORAL SESSION: Development [amp] Modulation of Hypothalamic Neuropeptidergic Neurons (11:15 AM-12:45 PM) Leptin Is Not Required for Fertility Oulu Wang, Satoru Sakihara, Kolbein Gudmundsson, Joseph Majzoub Children[apos]s Hospital, Boston, MA Stress responses to acute stressors improve the chances of immediate survival, even at the expense of immediate reproductive fitness. Stress-induced elevation of glucocorticoids is associated with inhibition of the hypothalamic-pituitary-gonadal reproductive axis. Leptin-deficient ob/ob mice are infertile and obese, and leptin is considered necessary for fertility. However, ob/ob mice also have significantly elevated glucocorticoid levels, and these elevated glucocorticoid levels may be responsible for infertility in leptin-deficient animals. Additionally, glucocorticoids stimulate appetite, and two common side effects of glucocorticoid therapy are hyperphagia and weight gain. We hypothesized that reducing glucocorticoid levels in leptin-deficient animals would restore fertility and ameliorate obesity.[br]To study this putative role of corticosterone in leptin deficiency, we generated mice with genetic deficits in the synthesis of glucocorticoids by deleting corticotropin-releasing hormone (CRH), leptin, or both. CRH- and leptin-deficient mice had hypotrophic adrenal glands, low corticosterone, and undetectable leptin levels. Compared to ob/ob mice, these males had higher testosterone levels, larger testes, and normal testicular histology. Females had higher luteinizing hormone and estradiol levels, larger ovaries, and larger uteri. Both males and females successfully generated litters, despite having global leptin deficiency, demonstrating that leptin is not required for reproduction. Leptin- and corticosterone-deficient mice had reduced hyperphagia and [gt]50% weight loss compared to ob/ob animals. Body fat composition, plasma triglycerides, and liver lipid content were all normalized.[br]To address whether the restoration of fertility and amelioration of body weight were the result of changes in glucocorticoids or CRH, we treated mice with exogenous corticosterone. Mice deficient for both CRH and leptin, but not either alone, were rendered infertile by glucocorticoid supplementation, without any induced elevation in CRH. Glucocorticoid supplementation caused increased body weight and plasma triglyceride levels in the CRH- and leptin-deficient group only, suggesting that glucocorticoids modulate fertility and appetite under conditions of insufficient leptin. These data indicate that the removal of glucocorticoid excess in the setting of low leptin is sufficient to restore fertility and ameliorate obesity.[br][br]Sources of Research Support: DoD NDSEG; NSF GRFP; NIH.[br][br]Nothing to Disclose: OW, SS, KG, JM 2012-06-23T12:00:00 371 2012-06-23T00:00:00 1899-12-30T12:00:00 1920 35 65 OR02-4 OR20-01 Saturday 10 2012


11 ENDO12L_OR02-5 ORAL SESSION: Development [amp] Modulation of Hypothalamic Neuropeptidergic Neurons (11:15 AM-12:45 PM) Deletion of Dopamine D2 Receptors in Pituitary Lactotropes Increases Food Intake, Body Weight and Adiposity Guillermina Maria Luque, Maria Ines Perez Millan, Damasia Becu-Villalobos, Marcelo Rubinstein IBYME-CONICET, Buenos Aires, Argentina; INGEBI-CONICET, Buenos Aires, Argentina Prolactin secretion is tonically inhibited by dopamine acting on lactotrope dopamine D2 receptors (D2Rs). Consequently, female mice lacking D2Rs ([italic]Drd2[sup]-/-[/sup][/italic]) exhibit chronic hyperprolactinemia and pituitary lactotrope hyperplasia. Although hyperprolactinemia has been associated with increased food intake and body weight, [italic]Drd2[sup]-/-[/sup][/italic] female mice display normal body weight probably because of multiple overlapping phenotypes derived from the absence of D2Rs from all brain areas and peripheral tissues. To dissect the importance of lactotrope D2Rs in food intake and body weight regulation we generated mutant mice carrying targeted [italic]Drd2 [/italic]deletions specifically in lactotropes (lac[italic]Drd2[sup]-/-[/sup][/italic]) using Cre/[italic]LoxP[/italic] technology. As expected, lac[italic]Drd2[sup]-/-[/sup][/italic] females are hyperprolactinemic whereas neuronal and peripheral D2Rs appear to function normally as determined by haloperidol-induced catalepsia and insulin serum levels. In addition, lac[italic]Drd2[sup]-/-[/sup][/italic] females showed increased body weight gain and adiposity in comparison with age-matched [italic]Drd2[sup]loxP/loxP[/sup][/italic] and [italic]Drd2[sup]-/-[/sup][/italic] female mice. Lac[italic]Drd2[sup]-/-[/sup][/italic] mice had similar body length than [italic]Drd2[sup]loxP/loxP[/sup][/italic] mice, but increased gonadal and retroperitoneal fat pads, as well as heavier livers (142%, 122% and 34% increase, respectively). Daily food intake was also increased in lac[italic]Drd2[sup]-/- [/sup][/italic]compared to [italic]Drd2[sup]loxP/loxP [/sup][/italic]mice (3.47[plusmn]0.10 g vs. 3.02[plusmn]0.17 g). Hypothalamic expression of genes associated to food intake showed that precursor of orexin ([italic]Ppo)[/italic] mRNA decreased in [italic]Drd2[sup]-/- [/sup][/italic]mice but not in lac[italic]Drd2[sup]-/-[/sup][/italic]. In contrast, hypothalamic [italic]Npy[/italic] mRNA was increased only in lac[italic]Drd2[sup]-/-[/sup][/italic] female mice. Thus, the increase in food intake, body weight and adiposity observed in lac[italic]Drd2[sup]-/-[/sup][/italic] females in comparison to [italic]Drd2[sup]-/-[/sup][/italic] mice highlights the importance of functional central D2Rs in the orexigenic effect promoted by high prolactin levels and suggest that the increased levels of [italic]Npy[/italic] mRNA found in lac[italic]Drd2[sup]-/-[/sup][/italic] female mice are probably involved in these phenotypes.[br][br]Nothing to Disclose: GML, MIPM, DB-V, MR 2012-06-23T12:15:00 371 2012-06-23T00:00:00 1899-12-30T12:15:00 442 35 66 OR02-5 OR20-01 Saturday 11 2012


12 ENDO12L_OR02-6 ORAL SESSION: Development [amp] Modulation of Hypothalamic Neuropeptidergic Neurons (11:15 AM-12:45 PM) Lateral Hypothalamic Neurotensin Neurons Act by Multiple Neural Pathways and Control Water Intake and Activity Gina M Leinninger, Paulette Goforth, Adriana Gata Garcia, Lyndsay M Christensen, Leslie Satin, Martin G Myers, Jr University of Michigan, Ann Arbor, MI; University of Michigan, Ann Arbor, MI The lateral hypothalamic area (LHA) integrates diverse physiologic stimuli with appropriate behavioral outputs (including feeding, drinking and locomotor behavior) to maintain homeostasis, but specific roles for individual populations of LHA neurons remain unresolved. We investigated the function of GABAergic LHA neurons that contain the neuropeptide Neurotensin (Nts). LHA Nts neurons are activated by dehydration, while the adjacent population of Orexin/hypocretin (OX)-containing neurons are inhibited by water deprivation. We injected an AAV that mediates the cre-dependent expression of the D3q Designer Receptor Activated Exclusively by Designer Drugs (DREADDs) into the LHA of Nts-cre mice, permitting the selective activation of LHA Nts neurons in response to the DREADD ligand, clozapine-N-oxide (CNO). Consistent with our previous data demonstrating the importance of LHA Nts neurons for the regulation of OX neurons by leptin, CNO-mediated activation of DREADD-expressing LHA Nts neurons inhibited a population of OX neurons. This DREADD-mediated activation of LHA Nts neurons promoted voracious drinking and increased locomotor activity, although food intake was not changed. While LHA Nts neurons synapse on LHA OX neurons and ventral tegmental area (VTA) dopamine (DA) neurons, numerous VTA DA neurons express Neurotensin Receptor-1 (NtsR1), but OX neurons do not. As VTA Nts promotes locomotor activity, our data suggest that LHA Nts neurons inhibit LHA OX neurons by GABA release and activate VTA dopamine neurons via Nts to modulate ingestive behaviors and locomotion.[br][br]Sources of Research Support: NIH R01 DK78056 (MGM) and NIH K99 DK090101 (GML).[br][br]Nothing to Disclose: GML, PG, AGG, LMC, LS, MGM 2012-06-23T12:30:00 371 2012-06-23T00:00:00 1899-12-30T12:30:00 1539 35 67 OR02-6 OR20-01 Saturday 12 2012


13 ENDO12L_OR03-1 ORAL SESSION: Cardiometabolic Risk (11:15 AM-12:45 PM) The Intake of Fiber Suppresses the High-Fat High-Carbohydrate Meal-Induced Endotoxemia, Oxidative Stress and Inflammation Husam Ghanim, Sandeep Dhindsa, Manav Batra, Kelly Green, Sanaa Abuaysheh, Ajay Chaudhuri, Paresh Dandona State University of New York at Buffalo, Buffalo, NY Our previous work has shown that the intake of a high fat high carbohydrate (HFHC) meal results in an increase in endotoxin (LPS) and LPS binding protein (LBP) concentrations concomitant with an increase in the expression of TLR-4, the receptor for LPS, and CD-14,which facilitates the binding of LPS to its receptor. In addition, there is an increase in ROS generation, NF[kappa]B binding and the expression of TNF[alpha] and IL-1[beta]. Our work has also demonstrated that an equicaloric meal rich in fiber and fruit (American Heart Association (AHA) recommended meal) does not induce these changes. We hypothesized, therefore, that the addition of fiber to a HFHC meal will prevent the changes induced by HFHC meal. Eight fasting normal subjects (BMI[lt]25Kg/m2) were given 900 Calorie meals of either HFHC or AHA or HFHC with additional fiber (30g) (Fiber One Original) on 3 sequential visits one week apart in a randomized crossover design. As expected, the HFHC meal induced an increase in LPS concentrations (by 69[plusmn]14%) with an increase in ROS generation (by 120[plusmn]24%) and the expression of TLR-4, CD14 and IL-1[beta] (by 71[plusmn]14%, 86[plusmn]14% and 127[plusmn]14%, respectively; p[lt]0.05 for all) in MNC. The AHA meal did not induce any of these changes. The addition of fiber to HFHC meal significantly reduced the increase in LPS concentrations (by 58[plusmn]10%; p[lt]0.05), ROS generation (by 47[plusmn]14%; p[lt]0.05) and the expression of TLR-4, CD14 and IL-1[beta] (by 37[plusmn]11%, 46[plusmn]12% and 52[plusmn]15%, respectively; p[lt]0,05 for all) observed with the HFHC meal alone. Additionally, the intake of fiber with the HFHC meal reduced postprandial glucose excursion and increased insulin concentrations compared to HFHC meal alone. We conclude that the addition of fiber to a pro-inflammatory HFHC meal has a beneficial anti-inflammatory and metabolic effect. Thus, the fiber content of the AHA meal may account for its non-inflammatory nature. Since ROS, TLR-4 and IL-1[beta] potentially interfere with insulin signal transduction and are known to be pro-inflammatory in the arterial wall, this action of dietary fiber may contribute to its benefits in the prevention of insulin resistance, type 2 diabetes and atherogenesis.[br][br]Disclosures: PD: Speaker, Novo Nordisk. Nothing to Disclose: HG, SD, MB, KG, SA, AC 2012-06-23T11:15:00 360 2012-06-23T00:00:00 1899-12-30T11:15:00 1997 36 68 OR03-1 OR10-01 Saturday 13 2012


14 ENDO12L_OR03-2 ORAL SESSION: Cardiometabolic Risk (11:15 AM-12:45 PM) Plasma 25-Hydroxyvitamin D Concentration and Prevalence of Metabolic Syndrome in the Diabetes Prevention Program Randomized Controlled Trial Joanna Mitri, Jason Nelson, Cheryl Garganta, David Nathan, Frank Hu, Bess Dawson-Hughes, Anastassios G Pittas Tufts Medical Center, Boston, MA; Institute for Clinical Research and Health Policy Studies, Boston, MA; Floating Hospital for Children at Tufts Medical Center, Boston, MA; Massachusetts General Hospital and Harvard Medical School, Boston, MA; Harvard School of Public Health Boston, MA and Channing Laboratory, Brigham and Women[apos]s Hospital, Boston, MA; Tufts University, Boston, MA Background: Low vitamin D status, assessed by blood 25-hydroxyvitamin D (25OHD) concentration, has been associated with cardiometabolic disease, including metabolic syndrome and type 2 diabetes but results are not consistent. The association of 25OHD and metabolic syndrome has not been studied in a multi-ethnic population at risk for diabetes and there are limited reports on the association between 25OHD concentration and non-traditional components of metabolic syndrome.[br]Objective: To investigate the association of plasma 25OHD concentration with metabolic syndrome and its individual components, and markers of insulin secretion and sensitivity in the at-risk-for-diabetes population of the Diabetes Prevention Program (DPP).[br]Design, setting and patients: Cross-sectional analysis using baseline data from the placebo and lifestyle intervention arms of the DPP (N=2040). The metformin arm was excluded to minimize the cost associated with measurement of 25OHD.[br]Outcomes: Metabolic syndrome as defined according to the modified criteria from the National Cholesterol Education Program[apos]s Adult Treatment Panel; individual components of metabolic syndrome (fasting plasma glucose, triglycerides and HDL cholesterol levels, waist circumference and blood pressure); insulin sensitivity, insulin secretion and disposition index (based on oral glucose tolerance test).[br]Results: After multivariate adjustment, including age, gender, race, site location, month of blood draw, body mass index, smoking, alcohol consumption, C-reactive protein, ultraviolet index and physical activity, participants in the highest tertile of vitamin D (median 25OHD concentration 30.6 ng/mL) had a lower risk of prevalent metabolic syndrome (odds ratio 0.62; 95%CI 0.45; 0.84) compared to participants in the lowest tertile (median 25OHD concentration 12.1 ng/mL). Those in the highest tertile of vitamin D status had smaller waist circumference, higher HDL, and lower fasting plasma glucose. Higher plasma 25OHD concentration was associated with higher insulin sensitivity and lower insulin secretion while there was no association with disposition index.[br]Conclusion: In a multi-ethnic population at high risk for diabetes, higher plasma 25OHD concentration was inversely associated with metabolic syndrome.[br][br]Sources of Research Support: Diabetes Program Precention Research Group. By research grants R01DK76092 and R01DK79003 (to AGP) from the National Institute of Diabetes and Digestive and Kidney Disease, the Office Of The Director - National Institutes of Health, and the National Institutes of Health Office of Dietary Supplements; UL1RR025752 (to Tufts Medical Center, Boston, MA) from the National Center for Research Resources; The US Department of Agriculture Agreement 58-1950-9001 (to BDH); The Marilyn Fishman Grant for Diabetes Research grant (to JM) from the Endocrine Fellows Foundation; research grant UO1DK48489 from the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) of the National Institutes of Health to the DPP clinical centers and the Coordinating Center for the design and conduct of the DPP study; The Southwestern American Indian Centers were supported directly by the NIDDK, including its Intramural Research Program, and the Indian Health Service; The General Clinical Research Center Program, National Center for Research Resources, supported data collection at many of the clinical centers; Funding was also provided by the National Institute of Child Health and Human Development, the National Institute on Aging, the National Eye Institute, the National Heart Lung and Blood Institute, the Office of Research on of Women[apos]s Health, the National Center for Minority Health and Human Disease, the Centers for Disease Control and Prevention, the Indian Health Service, and the American Diabetes Association; Lipha (Merck-Sante) provided medication. LifeScan Inc., Merck-Medco Managed Care, Inc., and Merck and Co. donated materials, equipment, or medicines for concomitant conditions.[br][br]Nothing to Disclose: JM, JN, CG, DN, FH, BD-H, AGP 2012-06-23T11:30:00 360 2012-06-23T00:00:00 1899-12-30T11:30:00 993 36 69 OR03-2 OR10-01 Saturday 14 2012


15 ENDO12L_OR03-3 ORAL SESSION: Cardiometabolic Risk (11:15 AM-12:45 PM) The Effects of Caloric Restriction on Fetuin-A and Cardiovascular Risk Factors in Rats and Humans: A Randomized Controlled Trial Ho Cheol Hong, Jae Hee Ahn, Nam Hoon Kim, Yoonjung Kim, Hae Yoon Choi, Joo Hyung Kim, Chai Ryoung Eun, Sae Jeong Yang, Hye Jin Yoo, Hee Young Kim, Ji A Seo, Nan Hee Kim, Sei Hyun Baik, Dong Seop Choi, Kyung Mook Choi Korea University, Seoul, Korea The secreted liver protein fetuin-A is associated with insulin resistance, metabolic syndrome, type 2 diabetes, and atherosclerosis. We examined the effect of caloric restriction (CR) on fetuin-A level and concomitant changes in hepatic steatosis and cardiovascular risk factors in rats and humans.[br]We performed a randomized controlled clinical trial to examine the circulating fetuin-A level and cardiovascular risk parameters including visceral fat area (VFA), atherogenic lipid profile, inflammatory markers, adipokines and brachial artery endothelial function in 76 overweight women with type 2 diabetes before and after 12 weeks of CR. In addition, the effects of CR on hepatic steatosis and fetuin-A mRNA expression were evaluated in obese OLETF rats, an animal model of obesity and type 2 diabetes.[br]Circulating fetuin-A levels were significantly decreased after 12 weeks of CR, accompanied by improvements in VFA, blood pressure, glucose, lipid profiles, and liver function. The CR group also showed significantly deceased apolipoprotein B, leptin, and insulin resistance compared to those in the control group, although endothelial function were not different. Multiple regression analysis showed that the changes in fetuin-A were independently associated with CR and changes in hsCRP, adiponectin, and HOMA-IR ([italic]R[/italic][sup]2[/sup] = 24.6%). Moreover, CR significantly reduced hepatic steatosis and fetuin-A expression, as well as weight, glucose, total cholesterol and triglyceride levels in OLETF rats.[br]CR significantly reduced the hepatic expression of fetuin-A and its circulating levels and improved several cardiovascular risk factors in obese rats and humans with type 2 diabetes.[br][br]Nothing to Disclose: HCH, JHA, NHK, YK, HYC, JHK, CRE, SJY, HJY, HYK, JAS, NHK, SHB, DC, KMC 2012-06-23T11:45:00 360 2012-06-23T00:00:00 1899-12-30T11:45:00 60 36 70 OR03-3 OR10-01 Saturday 15 2012


16 ENDO12L_OR03-4 ORAL SESSION: Cardiometabolic Risk (11:15 AM-12:45 PM) Low Serum Dehydroepiandrosterone Sulfate (DHEA-S) Is Independently Associated with Increased Risk of Cardiovascular Events in Elderly Men Asa Tivesten, Maria Nilsson, Daniel Carlzon, Liesbeth Vandenput, Magnus K Karlsson, Osten Ljunggren, Elizabeth Barrett-Connor, Dan Mellstrom, Claes Ohlsson University of Gothenburg, Gothenburg, Sweden; Lund University, Malm[ouml], Sweden; University of Uppsala, Uppsala, Sweden; University of California San Diego, La Jolla, CA [bold]Background:[/bold] Dehydroepiandrosterone (DHEA) is a sex hormone precursor from the adrenal cortex that circulates mainly as a sulfate ester (DHEA-S), which is desulfated within peripheral target tissues and subsequently exerts biological effects via conversion into active sex steroids (e.g., testosterone and estradiol). Several studies have addressed the association between DHEA-S levels and cardiovascular (CV) disease, but most are small cross-sectional or mortality studies and findings are inconsistent. The aim of the present study was to test the hypothesis that serum DHEA-S is an independent predictor of major fatal and nonfatal CV events in a large population-based cohort of elderly men followed for 5 years.[br][bold]Methods: [/bold]We used liquid/gas chromatography-mass spectrometry to analyze baseline levels of DHEA and DHEA-S in the prospective population-based MrOS Sweden study (2416 men, age 69-81 years). Complete CV clinical outcomes were available from central Swedish registers.[br][bold]Results: [/bold]During follow-up, 485 fatal and nonfatal CV events occurred. Both DHEA and DHEA-S levels were inversely associated with the risk of CV events. Men in the lowest quartile of DHEA-S had increased risk of major CV events compared with men in pooled quartiles 2-4 (age-adjusted hazard ratio [HR] 1.25, 95% confidence interval [CI] 1.03-1.53). This association remained significant after adjustment for traditional CV risk factors and serum testosterone and estradiol levels, and was not materially changed in analyses excluding 634 men with known CV disease at baseline or men whose CV events occurred during the first years of follow-up (follow-up 2.6 years or less; n=241). Men in the lowest quartile of both DHEA and DHEA-S, compared with men in quartile 2-4 of both, showed increased risk of any major CV event (HR 1.34, 95% CI 1.06-1.70), of coronary heart disease (HR 1.41, 95% CI 1.05-1.89) and cerebrovascular (HR 1.41, 95% CI 1.00-1.99) events.[br][bold]Conclusion:[/bold] Low serum DHEA-S independently predicted increased 5-year risk of CV events in older men.[br][br]Sources of Research Support: This study was supported by the Swedish Research Council, the Swedish Foundation for Strategic Research, The Avtal om L[auml]karutbildning och Forskning research grant in Gothenburg, the Swedish Heart-Lung Foundation, the Marianne and Marcus Wallenberg Foundation, the Lundberg Foundation, the Torsten and Ragnar S[ouml]derberg[acute]s Foundation, the [Aring]ke Wiberg Foundation and the Novo Nordisk Foundation.[br][br]Nothing to Disclose: AT, MN, DC, LV, MKK, OL, EB-C, DM, CO 2012-06-23T12:00:00 360 2012-06-23T00:00:00 1899-12-30T12:00:00 472 36 71 OR03-4 OR10-01 Saturday 16 2012


17 ENDO12L_OR03-5 ORAL SESSION: Cardiometabolic Risk (11:15 AM-12:45 PM) A Novel PPAR[gamma] Ligand, GQ16,decreases Inflammation and Atherosclerosis in a Mouse Model of the Metabolic Syndrome Alan Richard Collins, Laurie J Minze, Tuo Deng, Lin Jianxin, Maricela Ramirez, Rajagopalan Senapathy, Kevin Phillips, Joey Z Liu, Christopher J Lyon, Anders L Berkenstam, Paul Webb, Willa Ann Hsueh The Methodist Hospital Research Institute, Houston, TX; The Methodist Hospital Research Institute, Houston, TX Current ligands to peroxisomal proliferator activated receptor gamma (PPAR[gamma]) used to treat diabetes are highly useful, but have limiting detrimental side effects. New compounds are being sought that retain the beneficial anti-inflammatory and antioxidant effects without side effects. New ligands which specifically inhibit cdk5 phosporylation of PPAR[gamma] (SPPARMs) are being developed, but they have little anti-inflammatory effect compared to thiazolidinediones (TZDs). We previously characterized a mouse model of the metabolic syndrome, the male middle aged (12 months old) low density lipoprotein receptor deficient (Ldlr[sup]-/-[/sup]) mouse fed 3 months Western diet, in which the TZD rosiglitazone (RSG) decreases accelerated atherosclerosis. Using this model, we tested a novel compound, GQ16, which binds PPAR[gamma] in an altered conformation compared to TZDs, but still inhibits cdk5 phosporylation. We placed mice on: 1) chow control, 2) Western diet (WD), 3) WD with 1.2 g rosiglitazone/kg food or 4) WD with 0.2 g GQ16/kg food. At sacrifice, atherosclerosis was determined by the en face method. All metabolic parameters were significantly increased in the mice fed WD compared to chow. There were no differences in the body weights, %body fat, or fasted glucose in any of the WD treatments. Treatment with either RSG (1.4 ng/ml, p[lt]0.0001) or GQ16 (3.1 ng/ml, p[lt]0.01) decreased plasma insulin levels compared to WD (6.1 ng/ml). Neither RSG nor GQ16 altered the plasma lipids significantly. Atherosclerosis was increased by WD (16.4% vs 5.8% respectively, p[lt]0.01), which was substantially attenuated by both RSG (6.3%, p[lt]0.01 vs WD alone) and GQ16 (9.1%, p[lt]0.01 vs WD alone). Lesion histology indicated that vessels of mice treated with GQ16 had an abundance of chondrocyte-like cells, indicating a more stable lesion, suggesting wound healing compared to vessels from RSG and untreated mice. Adipocytes isolated from epidydimal fat pads were assayed for inflammatory markers. Adiponectin expression decreased with WD, while interleukin 1[alpha] and 1[beta] as well as the MHC class II transactivator (CIITA) increased. Both RSG and GQ-16 preserved adiponectin expression and attenuated pro-inflammatory genes, IL-1[alpha] and IL-1[beta], and others, as well as CIITA. Ligands such as GQ16 with have strong potential in the development of novel PPAR[gamma] therapeutics to treat inflammatory and metabolic disorders.[br][br]Nothing to Disclose: ARC, LJM, TD, LJ, MR, RS, KP, JZL, CJL, ALB, PW, WAH 2012-06-23T12:15:00 360 2012-06-23T00:00:00 1899-12-30T12:15:00 2208 36 72 OR03-5 OR10-01 Saturday 17 2012


18 ENDO12L_OR03-6 ORAL SESSION: Cardiometabolic Risk (11:15 AM-12:45 PM) Diabetes Aggravates Reperfusion Injury by Reducing [underline]A[/underline]poptosis [underline]R[/underline]epressor with [underline]C[/underline]aspase Recruitment Domain (ARC) Expression Anastasia Susie Mihailidou, Thi Yen Loan Le, Mahidi Mardini, Anthony W Ashton Royal North Shore Hospital, Sydney, Australia; University of Sydney, Sydney, Australia; Royal North Shore Hospital [amp] University of Sydney, Sydney, Australia; Westmead Hospital, Sydney, Australia; Royal North Shore Hospital, Sydney, Australia Diabetes is the fastest growing disease worldwide and myocardial infarction (MI) is a major cause of death. Acute hyperglycaemia at the time of MI also predicts adverse cardiovascular outcomes. Using an [italic]ex-vivo[/italic] experimental myocardial infarction model we have previously reported aggravated infarct size and apoptosis in both a Type 2 animal model of diabetes (Zucker rat) and during acute hyperglycemia, although the mechanism was not defined. Since myocardial apoptosis correlates with unfavourable post-infarction left ventricular remodelling with progression to heart failure, we now target whether there is a common mechanism between acute and prolonged hyperglycemia. ARC ([underline]A[/underline]poptosis [underline]R[/underline]epressor with [underline]C[/underline]aspase recruitment domain) is an endogenous apoptosis repressor protein, highly expressed in cardiac tissue and is unique to other proteins, by inhibiting both death receptor (extrinsic) and mitochondrial (intrinsic) death pathways. [bold][italic]Aim: [/italic][/bold]To determine the expression and activity of antiapoptotic repressor protein, ARC, during acute and prolonged hyperglycemia. [bold][italic]Methods: [/italic][/bold]Male Sprague Dawley rats (SD) and obese Zucker rats (ZDF) were anesthetised, blood glucose measured and hearts isolated and subjected to regional ischemia (30min) followed by reperfusion (2.5hr). For acute exposure studies, 22 mM glucose was perfused 15 mins. prior to inducing ischemia and throughout reperfusion. ARC expression was detected by immunostaining myocardial sections and quantified. Infarct area was measured relative to the area-at-risk and total area of left ventricle. [bold][italic]Results:[/italic][/bold] Glucose levels for ZDF rats (19 [plusmn] 1 mM, n=10) were comparable to acute hyperglycemia levels. Ischemia-reperfusion (I-R) markedly reduced ARC levels [0.99 x 10[sup]6[/sup] [plusmn] 0.1, N=7 vs 2.15 x 10[sup]6[/sup] [plusmn] 0.3, N=7 (sham), p[lt]0.05]. Reduced ARC expression persisted during acute hyperglycemia (Gluc 22 mM) [0.63 x 10[sup]6[/sup] [plusmn] 0.09, N=6 vs 2.15 x 10[sup]6[/sup] [plusmn] 0.3, N=7 (sham), p[lt]0.05], while prolonged hyperglycemia levels in the ZDF rats, aggravated reduction of ARC levels due to I-R [0.44 x 10[sup]6[/sup] [plusmn] 0.04, N=7 vs 0.99 x 10[sup]6[/sup] [plusmn] 0.1, N=7 (I-R), p[lt]0.05]. Preliminary studies to measure ARC activity show I-R induced loss of ARC expression. Both acute [58 [plusmn] 1.7%, N=8] and prolonged [49 [plusmn] 1.4%, N=7] hyperglycemia aggravated infarct area [39 [plusmn] 2%, N=8, (I-R alone), p[lt]0.05]. [bold][italic]Conclusion:[/italic][/bold] Our study identifies a novel mechanism in diabetes, whereby hyperglycemia aggravates reperfusion injury by degrading anti-apoptotic protein ARC, a master regulator of apoptosis.[br][br]Nothing to Disclose: ASM, TYLL, MM, AWA 2012-06-23T12:30:00 360 2012-06-23T00:00:00 1899-12-30T12:30:00 1180 36 73 OR03-6 OR10-01 Saturday 18 2012


37 ENDO12L_OR07-1 ORAL SESSION: New Insights into Clinical [amp] Molecular Diagnosis of Adrenal Tumors (11:15 AM-12:45 PM) Primary Pigmented Nodular Adrenocortical Disease (PPNAD) and Isolated Micronodular Adrenocortical Disease (iMAD) Are Not Only Genetically but Also Biochemically and Clinically Distinct Causes of Pediatric ACTH-Independent Cushing Syndrome Evgenia Gourgari, Anya Rothebuhler, Margaret Keil, Anelia Horvath, Constantine Stratakis National Institute of Child Health and Human Development, Bethesda, MD; National Institute of Child Health and Human Development, Bethesda, MD Endogenous Cushing syndrome (CS) is caused by excess adrenal glucocorticoid secretion that is adrenocorticotropin(ACTH)-dependent or -independent. In both adults and children, adrenocortical lesions causing CS include the cortisol-producing adenoma, the rare adrenocortical cancer, and bilateral adrenocortical hyperplasias (BAHs). BAHs can be further divided in micronodular or macronodular hyperplasiaswith adrenal nodules smaller or larger than 1 cm, respectively. Micronodular BAHs can be further divided in primary pigmented nodular adrenocortical disease (PPNAD) or various forms of isolated micronodular adrenocortical disease (iMAD). PPNAD is characterized by internodular atrophy and intense nodular pigment, mostly due to the deposition of lipofuscin, while iMAD has a variable picture of hyperplasia of zona fasciculata and is characterized by the relative absence of pigmentation. Patients with PPNAD show paradoxical stimulation of urinary 17-hydroxy steroids (17-OHS) and free cortisol (UFC) versus those with macronodular hyperplasias and/or cortisol-producing adenomas during the course of Liddle[apos]s test. It is unknown whether iMAD consistently shares this feature with PPNAD. Genetically, the two disorders are different, since most patients with PPNAD have germline mutations of PRKAR1A, whereas iMAD is due to mutations in mostly unknown genes, with the minority of the cases caused by germline defects of the phosphodiesterase (PDE) genes PDE11A and PDE8B. We studied patients that were seen at NIH during the last 30 years, all below the age of 21 years. We identified 10 patients with iMAD and 21 patients with PPNAD that had undergone Liddle[apos]s test before bilateral adrenalectomy for CS. We found statistically significant differences of iMAD versus PPNAD when comparing age at surgery (mean age 8.54 vs 14.07 years), BMI z score (2.43 vs 1.47, p=0.001), baseline 24hUFC/m2 (mean 351 vs 98mcg/m2/day, p=0.02). No difference was observed in sex distribution, baseline 24h urine17OH/gr cr and response to Liddles test. We conclude that both PPNAD and iMAD can be diagnosed by the [ldquo]paradoxical[rdquo] response to Liddle[apos]s test; however, they are genetically, histopathologically, and even clinically different disorders.[br][br]Nothing to Disclose: EG, AR, MK, AH, CS 2012-06-23T11:15:00 372 2012-06-23T00:00:00 1899-12-30T11:15:00 2085 40 92 OR07-1 OR01-01 Saturday 37 2012


38 ENDO12L_OR07-2 ORAL SESSION: New Insights into Clinical [amp] Molecular Diagnosis of Adrenal Tumors (11:15 AM-12:45 PM) Impact of Mutations in the Selectivity Filter of the KCNJ5 Channel on the Subtyping of Primary Aldosteronism with Adrenal Vein Sampling Teresa M Seccia, Maniselvan Kuppusamy, Giulio Ceolotto, Brasilina Caroccia, Maria V Cicala, Franco Mantero, GianPaolo Rossi University of Padua, Padua, Italy Background and aim. In primary aldosteronism (PA) mutations affecting the selectivity filter of the KCNJ5 K+ channel (Choi, Science 2011) were suggested to be associated with a more prominent aldosterone excess as assessed by peripheral plasma aldosterone concentrations (PAC)(Boulkroun, Hypertension 2012). Hence, we hypothesize that these mutations, being somatic, could determine a higher aldosterone secretion from the adrenal gland with the aldosterone-producing adenoma (APA) and therefore could affect the adrenal vein sampling (AVS) results. We therefore compared the lateralization index (LI) and the cortisol-corrected PAC in the adrenal vein draining the APA and the contralateral side between APA with (KCNJ5mut) and without (KCNJ5wt) the mutation.[br]Patients and Methods: Consecutive APA patients (n=86) undergoing AVS underwent sequencing of APA and germinal DNA to seek for the presence of the KCNJ5 mutations (Boulkroun, Hypertension 2012). We calculated the ratio of cortisol(PCC)-corrected PAC from the APA side (PAC/PCC)APA and of the PAC/PCC in the inferior vena cava (PAC/PCC)IVC and the lateralization index (LI) was calculated as the ratio of (PAC/PCC)APA and (PAC/PCC) at the contralateral side.[br]Results. The overall prevalence rate of G151R, L168R and T158A mutations was 34%. The KCNJ5mut and KCNJ5wt groups did not differ for gender, age, serum K+ levels, and ARR and blood pressure fall post-adrenalectomy. At baseline the KCNJ5mut group showed significantly higher (p[lt]0.03) PAC and ARR, and lower PRA (p[lt]0.03) than KCNJ5wt group, which suggested a more prominent aldosterone excess. This was reflected in a higher (PAC/PCC)APA/(PAC/PCC)IVC ratio and a higher LI in the KCNJ5mut than in the KCNJ5wt group. Accordingly, the LI was higher in the former than in the latter group (29.3[plusmn] 6.7 vs 16.7[plusmn]3.9, p[lt] 0.02).[br]Conclusions. The APA carrying the KCNJ5 mutations in the selectivity filter are characterized by a secretion of aldosterone from the tumor higher than APA wild type. As this translated into a higher LI at AVS, the presence of KCNJ5 mutations is likely to increase the accuracy of the subtyping of PA by means of AVS.[br][br]Nothing to Disclose: TMS, MK, GC, BC, MVC, FM, GR 2012-06-23T11:30:00 372 2012-06-23T00:00:00 1899-12-30T11:30:00 1886 40 93 OR07-2 OR01-01 Saturday 38 2012


39 ENDO12L_OR07-3 ORAL SESSION: New Insights into Clinical [amp] Molecular Diagnosis of Adrenal Tumors (11:15 AM-12:45 PM) Mitotane Induces a Mitochondrial Cytochrome c Oxidase Defect: A Novel Pharmacological Action for Adrenocortical Carcinoma Management Segolene Hescot, Abdelhamid Slama, Angelo Paci, Herve Remy, Rita Chadarevian, Severine Trabado, Anne Lombes, Jacques Young, Eric Baudin, Marc Lombes Faculty Medicine Paris SUd, Le Kremlin Bic[ecirc]tre, France; CHU Bicetre, Le Kremlin Bic[ecirc]tre, France; Institut Gustave Roussy, Villejuif, France; HRA-PHARMA France, Paris, France; Institut Cochin, Paris, France; CHU Bicetre, Le Kremlin Bic[ecirc]tre, France; Institut Gustave Roussy, Villejuif, France Mitotane (o,p[apos]DDD) is the most effective therapeutic strategy for adrenocortical carcinoma (ACC). Yet the molecular mechanisms of mitotane action remain unclear despite its well-established antisecretory and antiproliferative properties. Mitochondrial (mt) impact of mitotane has been previously suggested but not yet fully validated. We examined functional consequences of mitotane exposure on mitochondrial steroidogenesis, respiratory chain activity and biogenesis, using human adrenocortical secreting H295R and non-secreting SW13 cells. We first confirmed that mitotane exposure inhibits cell proliferation in a dose- and time-dependent manner. We were however unable to detect mitotane metabolites, o,p[apos]DDA and o[apos]p[apos]DDE, in cultured cells supernatant but could not rule out an intracellular metabolic transformation. We demonstrated that mitotane drastically reduces cortisol and 17-hydroxyprogesterone secretions by 70%, at a 50 [mu]M (14 mg/L) concentration, considered as the therapeutic plasma level threshold. This was accompanied by significantly decreased expression of genes encoding mitochondrial proteins involved in steroidogenesis (STAR, CYP11B1, CYP11B2). In both H295R and SW13 cells, 50 [mu]M mitotane induced a significant and reproducible 50% decrease of the maximum velocity of the respiratory chain complex IV (cytochrome c oxidase, COX) activity. This was associated with significantly decreased expression of both the mtDNA-encoded COX2 and the nuclear DNA encoded COX4 COX subunits, and with an important reduction of the whole COX steady state expression, as revealed by Blue Native PAGE. Enhanced mitochondrial biogenesis was shown by an increased mt DNA content and enhanced expression of the transcriptional regulator PGC1[alpha], consistent with an adaptive and compensatory mechanism against mitochondrial defect. Mitotane exposure also triggered a morphologic shift from filamentous to punctuate aspects of mitochondrial network, providing additional support for alteration in the mitochondrial compartment dynamics.[br]Collectively, our results provided direct evidence that mitotane induces mitochondrial respiratory chain defect. Better understanding of mitotane pharmacology should enable us to identify predictive factors of efficacy and toxicity.[br][br]Sources of Research Support: Grants from Inserm, Univ Paris-Sud, HRA-Pharma.[br][br]Nothing to Disclose: SH, AS, AP, HR, RC, ST, AL, JY, EB, ML 2012-06-23T11:45:00 372 2012-06-23T00:00:00 1899-12-30T11:45:00 913 40 94 OR07-3 OR01-01 Saturday 39 2012


40 ENDO12L_OR07-4 ORAL SESSION: New Insights into Clinical [amp] Molecular Diagnosis of Adrenal Tumors (11:15 AM-12:45 PM) Comparing Biochemical and Radiological Characteristics in Pheochromocytoma in Patients with Less Than Four Fold Elevated Metanephrines to Those with Higher Levels Subramanian Kannan, Andrei Purysko, Erick Remer, Allen Sipperstein, Eren Berber, Amr Fergany, Charles Faiman, Amir Hamrahian, Emmanuel Bravo Cleveland Clinic Foundation, Cleveland, OH; Cleveland Clinic Foundation, Cleveland, OH; Cleveland Clinic Foundation, Cleveland, OH; Cleveland Clinic Foundation, Cleveland, OH [bold]Background:[/bold] Pheochromocytoma (Pheo) needs to be ruled out in patients with suggestive symptomatology, adrenal incidentaloma and predisposing hereditary conditions. Tumor size shows a strong positive correlation with summed plasma (PL) or urinary (Ur) Metanephrines (METs). Correlation between CT attenuation (HU) and PL and/or Ur METs has not been described. A 4 fold elevation or higher of upper limits of normal (ULN) of PL or Ur METs is usually diagnostic for Pheo. A combination of PL or Ur METs and radiological features may be helpful in a timely diagnosis of Pheo in patients with METs [lt]4 fold ULN.[br][bold]Aim:[/bold] To compare Clinical and Radiological features in patients with Pheo who had PL and/or Ur METs [ge] 4 fold ULN (G1) and those with levels [lt] 4 fold ULN (G2).[br][bold]Methodology:[/bold] Among 252 patients with pathologically proven pheochromocytoma, we identified 105 patients who had images and at least one set of PL or Ur METs available for review (1997-2011). Metastatic Pheos were excluded. MET and NorMET were expressed as ratios above the ULN.[br][bold]Results: [/bold]The median (range) age and F/M sex ratios for G1 (n=83) and G2 (n=22) were 51 (19-84) years, 44/39; and 50 (24-73) years, 11/11 respectively. The percentage of patients with predisposing family history (G2 32% vs G1 12%, p 0.027) was different but similar for incidentally detected Pheo (G2 73% vs G1 48%, p 0.22). Extra-adrenal Pheo was exclusively seen in G1 (11%). There was a stronger correlation between radiological size of the tumor and summed ratios of PL METs (p [lt]0.001) and Ur METs (p[lt] 0.001) in G1 compared to G2 (p 0.14 and p 0.019). There was no correlation between PL or Ur METs and pre-contrast HU in each group. The radiological tumor size (cm) and the non-contrast CT HU for Gr 1 and Gr 2 was 4.9 (2-22) cm, 35.5 (17-59) HU; and 3 (0.9-5) cm;, 39 (25-52) HU, respectively (p [lt]0.001; p 0.65). The distribution of the radiological features in both groups are as follows (%): Heterogeneity (83 vs. 43 p 0.005), Necrosis (73 vs. 50 p 0.11), Hypervascularity (61 vs 56, p 0.99), Calcification (7 vs. 0, p 0.58), Hemorrhage (11 vs 0, p 0.58) and Cystic changes (69 vs. 50, p 0.22).[br][bold]Discussion:[/bold] We did not identify any tumor [gt] 5cm in size with PL or Ur METs [lt]4 fold ULN. None of the extr-adrenal tumors had elevations [lt] 4 fold ULN. Pheos in both groups had a pre-contrast HU [ge]17.[br][bold]Conclusion:[/bold] Patients with PL or Ur METs [lt] 4 fold ULN were more likely to have smaller tumors, predisposing family history and homogenous tumors on imaging.[br][br]Nothing to Disclose: SK, AP, ER, AS, EB, AF, CF, AH, EB 2012-06-23T12:00:00 372 2012-06-23T00:00:00 1899-12-30T12:00:00 1656 40 95 OR07-4 OR01-01 Saturday 40 2012


41 ENDO12L_OR07-5 ORAL SESSION: New Insights into Clinical [amp] Molecular Diagnosis of Adrenal Tumors (11:15 AM-12:45 PM) Genotype-Specific Correlations between Respiratory Chain Enzyme Activities, Succinate Accumulation and Catecholamine Content in Pheochromocytoma and Paraganglioma: An [italic]In Vitro[/italic] 1H-NMR Spectroscopy Study Jyotsna Upendra Rao, Udo F Engelke, Richard J Rodenburg, Ron A Wevers, Karel Pacak, Benno Kusters, Angelina G Goudswaard, Jacques W Lenders, Ad R Hermus, Arjen R Mensenkamp, Graeme Eisenhofer, Dirk H Kunst, Fred C Sweep, Henri J Timmers Radboud University Nijmegen Medical Centre, Nijmegen, Netherlands; Radboud University Nijmegen Medical Centre, Nijmegen, Netherlands; Radboud University Nijmegen Medical Centre, Nijmegen, Netherlands; Radboud University Nijmegen Medical Centre, Nijmegen, Netherlands; Radboud University Nijmegen Medical Centre, Nijmegen, Netherlands; Radboud University Nijmegen Medical Centre, Nijmegen, Netherlands; National Institute of Child Health and Human Development, Bethesda, MD; University Hospital Carl Gustav Carus, Dresden, Germany Background: Direct implication of mitochondrial enzymes in the tumorigenesis of pheochromocytoma and paraganglioma (PGL) came with the discovery of heterozygous germline mutations in succinate dehydrogenase (SDH) subunits causing familial paraganglioma syndromes. SDH catalyses the oxidation of succinate to fumarate in Krebs cycle and forms complex II of the mitochondrial respiratory chain. Reduced expression of components of respiratory chain complexes (RCC) and Krebs cycle has been observed in type I sporadics and patients with VHL (von Hippel Lindau) and SDHx mutations. Altered expression of components of RCCs, Krebs cycle and glycolysis in PGLs caused by SDHx and VHL mutations appears to be related to hypoxia/angiogenesis pathways of tumorigenesis through the stabilization of HIFs. Further, differences in energy metabolism in PGL may also translate into distinct profiles of catecholamine production, metabolism and secretion.[br]Objective: The aim of the study was to examine the relationship between RCC activities and succinate and catecholamine content of PPGLs across different genotypes.[br]Methods: 72 primary PGL tissues collected from patients with the following genotypes were investigated: SDHB (n=11), SDHD (n=7), SDHAF2 (n=2), VHL (n=7), NF1 (n=7), MAX (n=2), RET (n=14) and sporadic (n=22). Patients whose genotype was tested negative for SDHB, SDHC, SDHD, RET and VHL were considered as sporadics. The tumor tissue homogenates prepared in SEF buffer (10% w/v) were assayed for RCCs (complex I-IV) on Konelab20XT autoanalyzer using protocols reported before (1). Succinate and catecholamine levels in these tissues were measured using 1H-NMR spectroscopy. For this purpose, the tissues were homogenized in MilliQ water (10% w/v) and deproteinized by ultrafiltration using a 10 kDa filter. The ultrafiltrates were analyzed further using 500MHz 1H-NMR spectrometer.[br]Results: Complex II activity in general was barely above the detection limit in tissues with SDHx mutations. Low complex II activity was significantly associated with vast accumulation of succinate and with norepinephrine dominant tumor catecholamine content. Variations in the activities of RCCs I, III and IV were also observed among the different genotypes.[br]Conclusion: Decreased complex II activity in SDHx-related PPGL correlates with succinate accumulation and a noradrenergic phenotype. Genotype specific changes in the energy metabolism appear to have an impact on catecholamine metabolism.[br][br](1) Rodenburg R., J Inherit Metab Dis 2011; 34:283.[br][br]Nothing to Disclose: JUR, UFE, RJR, RAW, KP, BK, AGG, JWL, ARH, ARM, GE, DHK, FCS, HJT 2012-06-23T12:15:00 372 2012-06-23T00:00:00 1899-12-30T12:15:00 224 40 96 OR07-5 OR01-01 Saturday 41 2012


42 ENDO12L_OR07-6 ORAL SESSION: New Insights into Clinical [amp] Molecular Diagnosis of Adrenal Tumors (11:15 AM-12:45 PM) Definition of a New Algorithm of Genetic Testing in Paraganglioma and Pheochromocytoma Syndromes Julia Katharina Bickmann, Stefanie Sollfrank, Heidi Rossmann, Arno Schad, Racha Hassoun, Carolin Neukirch, Brigitte Schneider-Ratzke, Konstantinos Papaspyrou, Wolf J Mann, Karl J Lackner, Matthias M Weber, Christian Fottner University Medical Center Mainz, Mainz, Germany; University Medical Center Mainz, Mainz, Germany; University Medical Center Mainz, Mainz, Germany; University Medical Center Mainz, Mainz, Germany; University Medical Center Mainz, Mainz, Germany [bold]Objective[/bold]: We intended to optimise routine genetic testing of 116 pheochromocytoma and paraganglioma index patients by extending the usual panel of genes (SDHB, SDHC, SDHD, VHL, RET-protooncogene) by other genes that have been reported to be involved in the pathogenesis of endocrine tumor syndromes and by complementing genetic testing by prospective clinical follow-ups of all of these patients.[br][bold]Design and Method[/bold]: We designed sequencing assays of the genes MAX, SDHAF2/SDH5, TMEM127, PHD2 and SDHA. Large rearrangements of the SDH genes were detected by a commercial MLPA (Multiplex Ligation Dependant Probe Assay). Pyrosequencing was applied in the screening of the only known SDHAF2/SDH5 mutation and in evaluating various potential SNPs in a healthy control population.[br][bold]Results and Conclusions[/bold]: Out of 116 index patients 62% suffered from paragangliomas, 36% from pheochromocytomas and 2% from both tumor types. In 34% (39/116) of cases the underlying mutations were identified (familial tumor syndromes). Mutations were found in the following genes (given as percentage of all tested patients): 5% RET, 3% VHL, 1% TMEM127, 8% SDHB, 5% SDHC, 11% SDHD, and 1% SDHA. There were 6 novel SDH mutations including a large deletion in SDHB and an amino acid exchange in codon 1 of SDHA. We are currently complementing the genetic analysis by SDHB and SDHA immunohistochemistry of tumour samples. Familial cases of paragangliomas and pheochromocytomas are more common than assumed in the past. Genetic characterisation of paraganglioma patients including the detection of genomic rearrangements is essential for genetic counseling. According to our data all paraganglioma and pheochromocytoma patients who have undergone tumor surgery should also have genetic testing irrespective of their family histories. Even oligosymptomatic patients (e.g. isolated glomus tumors) may have more seriously affected relatives who would benefit from an early diagnosis.[br][br]Nothing to Disclose: JKB, SS, HR, AS, RH, CN, BS-R, KP, WJM, KJL, MMW, CF 2012-06-23T12:30:00 372 2012-06-23T00:00:00 1899-12-30T12:30:00 1037 40 97 OR07-6 OR01-01 Saturday 42 2012


43 ENDO12L_OR08-1 ORAL SESSION: Osteoporosis Physiology [amp] Clinical Trials (11:15 AM-12:45 PM) Bone Anabolic Efficacy and Safety of BA058, a Novel Analog of hPTHrP: Results from a Phase 2 Clinical Trial in Postmenopausal Women with Osteoporosis Gary Hattersley, John Bilezikian, Jonathan Guerriero, Prasanna Kumar, Jose Zanchetta, C Richard Lyttle, Louis St L O[apos]Dea Radius Health, Cambridge, MA; Columbia University College of Physicians and Surgeons, New York, NY; M S Ramaiah Memorial Hospital, Bangalore, India; Instituto de Investigaciones Metabolicas, Buenos Aires, Argentina BA058 is a synthetic analog of hPTHrP (1-34) undergoing clinical development for the treatment of postmenopausal osteoporosis. This was a randomized, double blind, placebo-controlled Phase 2 study to determine BA058 safety and effects on BMD and bone markers. Healthy postmenopausal women with severe osteoporosis (ages 55 to 85 years) were randomized to placebo, BA058 20, 40 or 80 ug, or teriparatide (TP) 20 ug for 24 weeks in an international, multicenter study. Available patients who completed 24 weeks could participate in an additional 24 weeks of treatment. 184 (83%) of 221 patients completed 6 months of treatment. The mean percent change in lumbar spine BMD at 24 weeks was 1.6% with placebo, 2.9%, 5.2%, and 6.7% with BA058 20, 40 and 80 ug respectively, and 5.5% with TP. The difference from placebo was significant (p[lt]0.001) for BA058 40 and 80 ug and for TP. Further increases in lumbar spine BMD were seen during the extension phase (n=55), with a mean percent change at 48 weeks of 0.7% with placebo, 5.1%, 9.8%, 12.9% with BA058 20, 40 and 80 ug respectively, and 8.6% with TP. A marked, dose dependent increase in total hip BMD was also seen at 24 weeks, where the mean change was 0.4% with placebo, 1.4%, 2.0%, and 2.6% with BA058 20, 40, and 80 ug respectively, and 0.5% with TP. At 24 weeks the change in serum and urine markers was significant (p[le]0.05) from baseline for BA058 40 and 80 ug and for TP for PINP, BSAP, osteocalcin and CTX, and with TP for NTX. BA058 was generally well tolerated with treatment-emergent AEs reported in 164 (74%) of 221 patients, with similar proportions across treatment groups. The most commonly reported events were influenza (10.4%) and headache (10.4%). Seven patients (3.1%) discontinued due to an adverse event, and serious adverse events were reported in 1.4% patients, none were treatment related. Mean serum calcium levels were consistently higher with TP compared to BA058, with mean values continuing to rise with TP over time. The safety profile of BA058 was consistent through an additional 6 months of treatment and no new safety signals were observed. In conclusion, treatment with BA058 80 ug resulted in significant BMD gains at all measured sites. BA058 was well tolerated, with safety events comparable with blinded placebo and consistent with medical events in this population. The safety and efficacy data support selection of BA058 80 ug for further clinical development for the treatment of postmenopausal osteoporosis.[br][br]Disclosures: GH: Employee, Radius Health. JG: Employee, Radius Health. CRL: Employee, Radius Health. LSLO: Employee, Radius Health. Nothing to Disclose: JB, PK, JZ 2012-06-23T11:15:00 Theater A 2012-06-23T00:00:00 1899-12-30T11:15:00 443 41 98 OR08-1 OR05-01 Saturday 43 2012


44 ENDO12L_OR08-2 ORAL SESSION: Osteoporosis Physiology [amp] Clinical Trials (11:15 AM-12:45 PM) BMD Response in the Odanacatib Phase 2 Trial: Prespecified Threshold and Subgroup Analyses Michael McClung, Neil Binkley, Henry Bone, Paul Miller, Nadia Verbruggen, Carolyn DaSilva, Elizabeth Rosenberg, Albert Leung Oregon Osteoporosis Center, Portland, OR; University of Wisconsin, Madison, WI; Michigan Bone and Mineral Clinic, Detroit, MI; Colorado Center for Bone Research, Lakewood, CO; MSD Europe, Brussels, Belgium; Merck Sharp [amp] Dohme Corp, Whitehouse Station, NJ [bold]Background:[/bold] Odanacatib (ODN) is a cathepsin K inhibitor in phase 3 development for osteoporosis. Treatment with ODN 50 mg once weekly for 2 years resulted in placebo (PBO)-subtracted mean increases in lumbar spine (LS) and total hip (TH) bone mineral density (BMD) of 5.7% and 4.1% respectively, P[le]0.001 vs. placebo. Five years of continuous treatment with ODN 50 mg resulted in BMD increases from baseline of 11.9% at LS and 8.5% at TH, and sustained reductions in bone resorption markers. In contrast, levels of bone formation markers were near baseline.[br][bold]Objectives:[/bold]To evaluate BMD response by thresholds and in pre-specified subgroups.[br][bold]Methods:[/bold] 399 postmenopausal women with low baseline BMD were randomized to ODN 3, 10, 25, 50 mg or PBO weekly, plus vitamin D and calcium. Percent change from baseline in LS BMD was the primary endpoint. Prespecified BMD-change threshold analyses and subgroup analyses by age, ethnicity, and baseline BMD and biomarkers are reported for women who received weekly placebo or ODN 50 mg (the dose being investigated in the phase 3 trial).[br][bold]Results:[/bold] At the LS, 0%, 95%, 76%, and 41% of participants treated with ODN 50 mg weekly for 2 years had BMD changes of [le]-3%, [gt]0%, [gt]3%, and [gt]6%, respectively, compared with 24%, 48%, 18%, and 7% in the PBO group. At the TH, the proportions were 0%, 85%, 48%, and 19% in the ODN group and 33%, 41%, 8%, and 0% in the PBO group. Subgroup analysis of % changes from baseline in LS BMD after 2 years revealed no statistically significant treatment-subgroup interactions between women who at baseline were [lt]65 vs. [ge]65 years old, Caucasian vs. non-Caucasian, or above or below the median values of baseline LS, hip trochanter, or femoral neck BMD or baseline bone resorption markers CTX or DPD. However, there were greater treatment effects in LS BMD in women with lower baseline TH BMD, higher baseline bone resorption marker NTX, or higher baseline bone formation markers BSAP and P1NP (7.2%, 6.6%, 7.5%, 6.9%, respectively) compared with women with the opposite baseline characteristics (4.3%, 4.6%, 4.2%, 4.4%, respectively). TH BMD subgroup analyses showed similar trends of smaller magnitude.[br][bold]Conclusions: [/bold]Treatment with ODN 50 mg was considerably more likely than treatment with PBO to increase LS and TH BMD to each threshold level. Subgroup analysis of BMD change suggests that participants with lower BMD or higher bone turnover at baseline may have a greater % BMD increase with ODN treatment.[br][br]Disclosures: MM: Investigator, Amgen, Merck & Co., Takeda; Consultant, Amgen, Lilly USA, LLC, Merck & Co., Novartis Pharmaceuticals, Warner Chilcott Pharmaceuticals, Takeda; Speaker, Amgen, Lilly USA, LLC, Merck & Co., Novartis Pharmaceuticals, Warner Chilcott Pharmaceuticals.NB: Consultant, Merck & Co., Lilly USA, LLC, Tarsa; Investigator, Merck & Co., Amgen; Researcher, Tarsa. HB: Consultant, Amgen, Merck & Co., Takeda, Tarsa; Investigator, Amgen, Merck & Co., Nordic Bioscience A/S, Novartis Pharmaceuticals, Takeda, Tarsa; Speaker Bureau Member, Amgen. PM: Consultant, Merck & Co.; Investigator, Merck & Co. NV: Employee, Merck & Co. CD: Employee, Merck & Co. ER: Employee, Merck & Co. AL: Employee, Merck & Co. 2012-06-23T11:30:00 Theater A 2012-06-23T00:00:00 1899-12-30T11:30:00 1763 41 99 OR08-2 OR05-01 Saturday 44 2012


45 ENDO12L_OR08-3 ORAL SESSION: Osteoporosis Physiology [amp] Clinical Trials (11:15 AM-12:45 PM) Distinctive Role of 6-Month Teriparatide on Advanced Bisphosphonate-Related Osteonecrosis of the Jaw (BRONJ); Case-Control Study Kyoung Min Kim, Wonse Park, Hyung-Jun Kim, Woong Nam, Sung-Kil Lim, In Ho Cha, Yumie Rhee Yonsei University College of Medicine, Seoul, Korea; Yonsei University College of Dentistry, Seoul, Korea; Yonsei University College of Dentistry, Seoul, Korea Management of bisphosphonate-related osteonecrosis of the jaw(BRONJ) is a problematic in subset of patients. The objective was to determine whether combining teriparatide(TPTD) could overcome the difficulties in managing advanced cases of BRONJ patients. Twenty four subjects of advanced BRONJ, who did not recover from osteonecrosis with previous dental care including sequestrectomy, out of 119 BRONJ patients were chosen for the case-control study. Fifteen patients received daily 20 ug of TPTD injection subcutaneously along with conventional therapy for 6 months[TPTD group] and the other 9 patients were managed only with conservative care[non-TPTD group]. Bone turnover markers and hormones related to bone metabolism were analyzed at baseline, 1 and 6 months of treatment. Stages of BRONJ were assessed every month by two separate dentists using stages established by American Association of Oral and Maxillofacial Surgery. Outcome of BRONJ treatment was defined as [apos]not improved[apos] with no changes in stages of BRONJ after 6 months of treatment, [apos]moderately improved[apos] with one stage improvement and [apos]markedly improved[apos] with two stages upgrade or complete healing. The initial stages of BRONJ were mostly stage 2, 93.3%(14/15) in TPTD group and 88.9%(8/9) cases in non-TPTD groups, and the remainders of both groups were stage 3. The increment of C-telopeptide(CTx) was significantly higher in TPTD group compared to the level of non-TPTD group at 6 months (475.9[plusmn]334.9% vs. 0.2[plusmn]11.9%, [italic]p[/italic][lt]0.05). Osteocalcin also showed significant increase at 6 months in TPTD-group (254.1[plusmn]29.5% vs. -2.5[plusmn]7.5%, [italic]p[/italic][lt]0.05). After 6 months of treatment, 62.5% of TPTD group was classified as [apos]markedly improved[apos], 37.5% [apos]moderately improved[apos] without any case that did not improve, whereas final status of non-TPTD group ended up as 60%, [apos]moderately improved[apos] and 40%, [apos]not improved[apos] ([italic]p[/italic][lt]0.05 vs. TPTD group). Interestingly, 25(OH)D level showed lower tendency in [apos]moderately improved[apos] patients compared to [apos]markedly improved[apos] ones in TPTD group (15.3[plusmn]9.3 ng/dL vs. 29.7[plusmn]6.3 ng/dL, [italic]p[/italic]=0.056). Initial 1 month change of CTx had a strong positive correlation with 25(OH)D (r=0.649, [italic]p[/italic][lt]0.05) and a negative correlation with PTH (r=-0.670, [italic]p[/italic][lt]0.05). In conclusion, 6-month use of TPTD was superior on the resolution of advanced BRONJ compared to the conventional therapy via accelerated recovery of bone remodeling. Moreover, vitamin D insufficiency could be a risk factor for lesser response to the TPTD in BRONJ patients.[br][br]Nothing to Disclose: KMK, WP, H-JK, WN, S-KL, IHC, YR 2012-06-23T11:45:00 Theater A 2012-06-23T00:00:00 1899-12-30T11:45:00 748 41 100 OR08-3 OR05-01 Saturday 45 2012


46 ENDO12L_OR08-4 ORAL SESSION: Osteoporosis Physiology [amp] Clinical Trials (11:15 AM-12:45 PM) Sclerostin Levels and Bone Turnover Markers in Adolescents with Anorexia Nervosa and Healthy Adolescent Girls Alexander T Faje, Pouneh Fazeli, Debra K Katzman, Karen K Miller, Anne Breggia, Clifford J Rosen, Nara Mendes, Anne Klibanski, Madhusmita Misra Massachusetts General Hospital and Harvard Medical School, Boston, MA; Hospital for Sick Children and University of Toronto, Toronto, Canada; Maine Medical Center Research Institute, Portland, ME; Massachusetts General Hospital for Children and Harvard Medical School, Boston, MA Sclerostin is an important determinant of bone formation and resorption. Adult studies have shown that sclerostin is negatively regulated by estradiol. Adolescents with anorexia nervosa (AN) are hypogonadal and have low bone mineral density (BMD) and decreased levels of bone turnover markers. Sclerostin has not been examined in AN as a potential mediator of impaired bone metabolism.[br]Our study objectives were to (i) compare sclerostin in girls with AN and controls, (ii) assess associations with bone turnover markers, and (iii) examine effects of transdermal estradiol on sclerostin in AN.[br]A subset of adolescent girls with AN and normal-weight controls 13-18 years old were selected from a prior study based on their lumbar spine BMD Z-scores. The subset included all subjects with AN with a baseline lumbar BMD Z-score [lt] -0.5 and controls with lumbar BMD Z-scores between +1.0 and -1.0. 69 girls (44 with AN and 25 normal-weight controls) were studied at baseline. 22 AN girls with a bone age of [ge] 15 years were randomized to 100mcg transdermal estradiol (n=13) or placebo (n=9) in a double-blind study for 12 months. Subjects randomized to transdermal estradiol received medroxyprogesterone 2.5mg daily for 10 days each month. All subjects also received calcium carbonate (1200mg) and vitamin D (400 IU) daily.[br]Sclerostin levels did not differ in AN versus controls (0.422[plusmn]0.016 vs. 0.408[plusmn]0.018 ng/ml, p=0.58). Sclerostin correlated positively with P1NP and CTX in controls (r=0.49 and 0.55, p=0.01 and 0.004, respectively) but not in AN (r=0.09 and -0.17, p=0.62 and 0.34). Treatment with transdermal estradiol significantly improved lumbar BMD compared with placebo ([Delta] lumbar BMD: 0.035[plusmn]0.007 vs. 0.005[plusmn]0.012 g/cm[sup]2[/sup], p=0.01). However, changes in sclerostin over twelve months did not differ in girls randomized to estradiol or placebo (-0.101[plusmn]0.031 vs. -0.128[plusmn]0.042 ng/ml, p = 0.61).[br]Levels of sclerostin do not differ in girls with AN compared with normal-weight adolescents. The relationship between sclerostin and markers of bone turnover in normal-weight girls is disrupted in adolescent girls with AN. Despite an increase in BMD with transdermal estradiol administration in AN, estrogen does not impact sclerostin levels in this group. These findings imply (i) the presence of additional unspecified factors that regulate sclerostin in adolescents with AN, and (ii) that changes in sclerostin do not mediate the beneficial effects of transdermal estradiol on BMD in this group.[br][br]Sources of Research Support: R01 DK062249 and UL1 RR025758-01.[br][br]Nothing to Disclose: ATF, PF, DKK, KKM, AB, CJR, NM, AK, MM 2012-06-23T12:00:00 Theater A 2012-06-23T00:00:00 1899-12-30T12:00:00 88 41 101 OR08-4 OR05-01 Saturday 46 2012


47 ENDO12L_OR08-5 ORAL SESSION: Osteoporosis Physiology [amp] Clinical Trials (11:15 AM-12:45 PM) Denosumab for the Treatment of Men with Low Bone Mineral Density Ugis Gruntmanis, Eric Orwoll, Christence S Teglbjaerg, Bente L Langdahl, Roland Chapurlat, Edward Czerwinski, David L Kendler, Jean-Yves Reginster, Alan Kivitz, E Michael Lewiecki, Paul D Miller, Michael A Bolognese, Michael R McClung, Henry G Bone, Osten Ljunggren, Bo Abrahamsen, Yu-Ching Yang, Rachel W Wagman, Suresh Siddhanti, Andreas Grauer, Jesse W Hall, Steven Boonen Dallas Veterans Affairs Medical Center and University of Texas Southwestern Medical Center, Dallas, TX; Oregon Health and Science University, Portland, OR; Clinical and Basic Research, Ballerup, Denmark; Aarhus University Hospital, Aarhus, Denmark; Hopital Edouard Herriot, Lyon, France; Karkow Medical Center, Krakow, Poland; University of British Columbia, Vancouver, Canada; University of Liege, Liege, Belgium; Altoona Center for Clinical Research, Duncansville, PA; New Mexico Clinical Research and Osteoporosis Center, Alburquerque, NM; University of Colorado Health Science Center, Lakewood, CO; Bethesda Health Research Center, Bethesda, MD; Oregon Osteoporosis Center, Portland, OR; Michigan Bone and Mineral Clinic, Detroit, MI; Uppsala University, Uppsala, Sweden ; Copenhagen University Hospital Gentofte, Hellerup, Denmark; Amgen Inc, Thousand Oaks, CA; Amgen Inc, Thousand Oaks, CA; Leuven University, Leuven, Belgium [bold]Purpose: [/bold]The objective of this study was to evaluate the effect of denosumab (DMAb) on bone mineral density (BMD) in men with low BMD.[br][bold]Method: [/bold]This was a multicenter, randomized, double-blind, placebo-controlled study where subjects were randomized 1:1 to receive either 60 mg DMAb or placebo administered subcutaneously once every 6 months over a 12-month period. Subjects were male, and were included if they were [ge]30 and [le]85 yrs of age; had a BMD T-score [le]-2.0 and [ge]-3.5 at the lumbar spine (LS) or femoral neck (FN), or had a prior major osteoporotic fracture and a T-score [le]-1.0 and [ge]-3.5 at the LS or FN; and had at least two lumbar vertebrae, one femur, and one forearm evaluable by DXA. Subjects received daily calcium ([ge]1000 mg) and vitamin D ([ge]800 IU) supplementation. The primary endpoint was to assess the percent change from baseline in LS BMD at 12 months. Secondary endpoints included the percent change from baseline in total hip (TH), FN, trochanter (TR), and forearm (1/3R) BMD at month 12. Sensitivity analyses for LS BMD were done by controlling for baseline covariates (testosterone levels, BMD T-scores, race, geographic region and 10-year major osteoporotic fracture risk). The efficacy of DMAb on LS BMD was assessed in different subgroup populations (grouped by testosterone levels, minimum BMD T-scores, and 10-year major osteoporotic fracture risk). Safety analysis included all randomized subjects who received [ge]1 dose of investigational product.[br][bold]Results: [/bold]A total of 242 subjects were randomized; 228 (94.2%) completed one year of treatment. The mean (SD) age was 65 (9.8) yrs and 15% of subjects had testosterone concentration [lt]250 mg/dL at baseline. After 12 months of treatment with DMAb, BMD increased from baseline by 5.7%, 2.4%, 2.1%, 3.1%, and 0.6% at the LS, TH, FN, TR, and 1/3R, respectively (all [italic]p[/italic][lt]0.01 compared with placebo and adjusted for multiplicity). Sensitivity analyses for LS BMD were consistent with the primary analysis (mean differences from placebo were significant, [italic]p[/italic][lt]0.01), indicating the primary analysis results were robust. DMAb treatment resulted in significant increases in LS BMD in all subgroups (mean differences from placebo were significant, [italic]p[/italic][lt]0.005), indicating DMAb is effective across different clinical situations. Adverse event incidence was similar between groups.[br][bold]Conclusions: [/bold]One year of DMAb therapy in men with low BMD was well tolerated and resulted in significant increases in BMD at all measured skeletal sites.[br][br]Disclosures: EO: Research Funding, Amgen, Eli Lilly & Company, Merck & Co.; Consultant, Eli Lilly & Company, Merck & Co., Amgen, Wright Medical Technology. CST: Coinvestigator, Amgen. BLL: Research Funding, Amgen, Eli Lilly & Company, Novartis Pharmaceuticals, MSD; Consultant, Amgen, Eli Lilly & Company, Novartis Pharmaceuticals, Servie, Nycomed, MSD. RC: Advisory Group Member, Amgen, Eli Lilly & Company, Servier, Merck & Co., Novartis Pharmaceuticals; Speaker, Amgen, Ipsen, Servier, Roche Pharmaceuticals. EC: Advisory Group Member, Amgen, Eli Lilly & Company, Servier, Merck & Co., Novartis Pharmaceuticals; Speaker, Amgen, Ipsen, Servier, Roche Pharmaceuticals. DLK: Coinvestigator, Eli Lilly & Company, GlaxoSmithKline, Roche Pharmaceuticals; Advisory Group Member, Merck & Co., Novartis Pharmaceuticals, Amgen, Pfizer Global R&D. J-YR: Advisory Group Member, Servier, Novartis Pharmaceuticals, Negma, Lilly USA, LLC, Wyeth Pharmaceuticals, Genzyme Corporation, Roche Diagnostics, Merck & Co., Nycomed, NPS, Theramex, UCB; Speaker, Merck Sharp and Dohme, Lilly USA, LLC, Rottapharm, IBSA, Genevrier, Novartis Pharmaceuticals, Roche Pharmaceuticals, GlaxoSmithKline, Teijin, Teva, Ebewee Pharma, Zodiac, Analis, Theramex, Nycomed, Novo Nordisk; Investigator, Bristol Myers Squibb, Merck Sharp and Dhome, Rottapharm, Teva, Lilly, Novartis, Roche, GlaxoSmithKline, Amgen, Servier. AK: Coinvestigator, Amgen. EML: Principal Investigator, Amgen, Eli Lilly & Company, Merck & Co., Novartis Pharmaceuticals, GlaxoSmithKline, Warner Chilcott Pharmaceuticals; Speaker Bureau Member, Amgen, Eli Lilly & Company, Novartis Pharmaceuticals, Warner Chilcott Pharmaceuticals; Scientific Board Member, Amgen, Eli Lilly & Company, Merck & Co., Novartis Pharmaceuticals; Consultant, GlaxoSmithKline. PDM: Research Funding, Amgen, Eli Lilly & Company, Genentech, Inc., Merck & Co., Novartis Pharmaceuticals, Warner Chilcott Pharmaceuticals; Speaker Bureau Member, Amgen, Eli Lilly & Company, Genentech, Inc., Merck & Co., Novartis Pharmaceuticals, Warner Chilcott Pharmaceuticals, GlaxoSmithKline. MAB: Research Funding, Amgen, Eli Lilly & Company, Merck & Co.; Speaker Bureau Member, Amgen, Eli Lilly & Company, Warner Chilcott Pharmaceuticals. MRM: Consultant, Merck & Co. HGB: Consultant, Amgen, Merck & Co., Takeda, Tarsa; Investigator, Amgen, Merck & Co., Nordic Bioscience A/S, Novartis Pharmaceuticals, Takeda, Tarsa; Speaker Bureau Member, Amgen. BA: Research Funding, Amgen, Novartis Pharmaceuticals; Consultant, Amgen, Novartis Pharmaceuticals, Eli Lilly & Company, Nycomed; Speaker Bureau Member, Merck & Co., Nycomed. Y-CY: Employee, Amgen. RWW: Employee, Amgen. SS: Employee, Amgen. AG: Employee, Amgen. JWH: Employee, Amgen. SB: Research Funding, Amgen; Consultant, Amgen. Nothing to Disclose: UG, OL 2012-06-23T12:15:00 Theater A 2012-06-23T00:00:00 1899-12-30T12:15:00 1789 41 102 OR08-5 OR05-01 Saturday 47 2012


48 ENDO12L_OR08-6 ORAL SESSION: Osteoporosis Physiology [amp] Clinical Trials (11:15 AM-12:45 PM) Behavior of Bone Turnover Markers in Infertile Women Undergoing Follicle-Stimulating Hormone Administration for [italic]In Vitro[/italic] Fertilization Procedures Umberto Omodei, Gherardo Mazziotti, Gloria Donarini, Monica Gola, Valentina Guella, Franca Pagani, Teresa Porcelli, Andrea Giustina Universit[agrave] degli Studi di Brescia, Brescia, Italy; Universit[agrave] degli Studi di Brescia, Brescia, Italy; Fondazione Poliambulanza, Brescia, Italy There is experimental but limited clinical evidence to suggest that the increase in follicle-stimulating hormone (FSH) secretion, as it occurs since the first phase of perimenopause, may be involved in determining post-menopausal bone loss. In this prospective study, we evaluated for the first time the sequential profile of serum bone turnover markers during acute stimulation with recombinant FSH in 29 infertile women (age ranging from 30 and 40 years) undergoing in vitro fertilization procedures. In this clinical model, the skeletal effects of exogenous FSH were evaluated after pharmacological suppression of endogenous gonadotropin and estrogen production. The women were evaluated for serum osteocalcin (Elecsys N-MID Osteocalcina, Roche Diagnostics, Monza, Italy; reference range: 11-32 [mu]g/L), [beta]-CTX (Elecsys [beta]-CrossLaps Roche Diagnostics, Monza, Italy; reference range: 0.10-0.62 [mu]g/L), FSH and estradiol at baseline (T0), 12-20 days after the first dose of GnRH analog (Leuprolide 1 mg subcutaneously each day) (T1), 3 days (T2), 10 days (T3) after recombinant FSH administration (Meropur 75 in the morning and Gonal-F 1050 in the evening at doses between 75 and 225 UI each day) and at pick-up (T4) and at the embryo-transfer day (T5). All women were taking also prednisolone between T4 and T5. At study entry, all patients had serum osteocalcin and [beta]-CTX values in the reference ranges for pre-menopausal women. At T1, the suppression of FSH and E2 secretion, as affect of GnRH analog administration, was accompanied by a significant increase in serum [beta]-CTX values as compared to T0 (p[lt]0.001), whereas no significant change in serum osteocalcin was observed at this time. After the administration of recombinant FSH, a rapid increase in serum FSH was observed, whereas serum E2 values increased more slowly. At this time, the increase in serum FSH values (with E2 in normal range) was not associated with any significant change in median serum [beta]-CTX and osteocalcin values as compared to T1. At T3 (when both FSH and E2 were high), serum [beta]-CTX values decreased significantly as compared to T1 (p[lt]0.001), whereas osteocalcin remained unchanged. At T4, serum [beta]-CTX and osteocalcin remained unchanged as compared to T3, whereas both markers decreased significantly (p[lt]0.001 for both parameters) at T5. In conclusion, our model suggests that FSH, at high-normal circulating values, does not seem to acutely exert relevant direct effects on bone.[br][br]Nothing to Disclose: UO, GM, GD, MG, VG, FP, TP, AG 2012-06-23T12:30:00 Theater A 2012-06-23T00:00:00 1899-12-30T12:30:00 2195 41 103 OR08-6 OR05-01 Saturday 48 2012


49 ENDO12L_OR09-1 ORAL SESSION: Pediatric Endocrinology, General (11:15 AM-12:45 PM) Mutations in the [italic]AAAS[/italic] Gene in 215 Patients with Triple A Syndrome [mdash] Analysis of a Genotype/Phenotype Relationship Angela Huebner, Daniela Klaus, Dana Landgraf, Petra Mitzscherling, Katrin Koehler Technical University Dresden, Dresden, Germany The triple A syndrome (MIM*231550) is a rare autosomal recessive disorder characterized by adrenocorticotropic hormone (ACTH) resistant adrenal failure, alacrima, achalasia and a variety of neurological and dermatological features. The syndrome is caused by mutations in the [italic]AAAS[/italic] gene encoding ALADIN, a protein of the nuclear pore complex. We investigated N=173 families including 215 patients with clinically suggested triple A syndrome. In 142 families we identified 62 different [italic]AAAS[/italic] mutations including 19 nonsense mutations, 15 deletions and 2 insertions leading to a frameshift and a truncated protein, 13 splice defects and 13 missense mutations which were present in a homozygous or compound heterozygous form. Although the mutations were scattered over the entire gene a few mutation hotspots became apparent including c.43C[gt]A (exon1), c.787T[gt]C (exon8) and c.1331+1G[gt]A (intron 14), each occurring in 20 and more families from different regions. In 31 families with triple A syndrome we could not detect [italic]AAAS[/italic] mutations indicating genetic heterogeneity. Two patients carry [italic]AAAS[/italic] mutations in a heterozygous form. The phenotype of patients who either lack a mutation or appear to be heterozygous does not differ from patients with a proven [italic]AAAS[/italic] mutation.[br]Genotype/phenotype analyses revealed a highly variable occurrence, age of onset and severity of all clinical symptoms between patients with the same [italic]AAAS[/italic] mutation. This is reflected by the considerable intrafamilial variability which may also concern the three main symptoms. Whereas alacrima appears to be the earliest and most consistent symptom (93 % of all patients), achalasia was observed in 86 %. Adrenal insufficiency (75 % of all patients) usually occurs in the first decade of life but may occur only in the second decade or even later. The severity and progression of neurological features resembling those of a mixed hereditary motor and sensory neuropathy with axonal degeneration cannot be correlated to the localization and the nature of the mutations in the [italic]AAAS[/italic] gene. Nasal speech and a marked generalized hyperreflexia appear to be pathognomonic signs.In conclusion, in view of the highly variable clinical course, the triple A syndrome seems to be an underdiagnosed disorder. [italic]AAAS[/italic] mutation screening should therefore be extended to patients who exhibit only single features of the disease. The obvious lack of a genotype/phenotype relationship is suggestive of modifying genes/factors which have to be determined.[br][br]Sources of Research Support: German Research Foundation HU/895 3-3,3-4,3-5 and 4-1 awarded to AH.[br][br]Nothing to Disclose: AH, DK, DL, PM, KK 2012-06-23T11:15:00 Theater C 2012-06-23T00:00:00 1899-12-30T11:15:00 1257 42 104 OR09-1 OR30-01 Saturday 49 2012


50 ENDO12L_OR09-2 ORAL SESSION: Pediatric Endocrinology, General (11:15 AM-12:45 PM) Finite Element and Extended Cortical Analysis in Young Amenorrheic Athletes, Eumenorrheic Athletes and Non-Athletes Kathryn E Ackerman, Melissa Putman, Gabriela Guereca, Alexander P Taylor, Lisa Pierce, David B Herzog, Anne Klibanski, Mary Bouxsein, Madhusmita Misra Harvard Medical School, Boston, MA; Children[apos]s Hospital Boston, Boston, MA; Massachusetts General Hospital, Boston, MA; Massachusetts General Hospital, Boston, MA; Massachusetts General Hospital for Children, Boston, MA CONTEXT: Lower bone mineral density (BMD) in young amenorrheic athletes (AA) compared to eumenorrheic athletes (EA) and non-athletes may increase fracture risk during a critical time of bone accrual. Finite element analysis (FEA) is a unique [italic]in vivo[/italic] tool to estimate bone strength, while extended cortical analysis (ECA) allows assessment of cortical parameters that may contribute to bone strength using data from high-resolution peripheral QCT (HRpQCT).[br]OBJECTIVE: We hypothesized that stiffness and failure load are impaired in AA at the distal radius and tibia compared to EA and non-athletes despite weight-bearing exercise.[br]DESIGN AND SETTING: Cross-sectional study at the Clinical Research Center[br]SUBJECTS: Subjects included 34 female endurance athletes 14-21 years old involved in weight-bearing sports (17 AA, 17 EA) and 16 non-athletes of normal-weight (BMI 10th-90th percentiles) and comparable age and maturity. All subjects had body composition assessment using dual energy x-ray absorptiometry (DXA).[br]OUTCOME MEASURES: We used ECA to assess cortical parameters including porosity, and FEA to assess stiffness and failure load, using images obtained from HRpQCT.[br]RESULTS: At the weight-bearing tibia, both groups of athletes had greater cortical perimeter and porosity (p=0.02 and 0.01 respectively), and greater trabecular area than non-athletes (p=0.01), while percent cortical area was lowest in AA (14.6[plusmn]3.2% vs. 16.4[plusmn]3.0% in EA and 18.6[plusmn]4.9% in non-athletes; p=0.01). Despite greater cortical porosity in EA than non-athletes, stiffness and failure load were higher in EA (p=0.01 for both). This advantage, however, was lost in AA. At the non-weight-bearing radius, failure load and stiffness were lower in AA than non-athletes [failure load (kN): 3.60 [plusmn] 0.77 in AA, 4.05 [plusmn] 0.60 in EA and 4.20 [plusmn] 0.74 in non-athletes, p=0.048; stiffness (kN/m): 70.5[plusmn]14.8 in AA, 79.5[plusmn]12.1 in EA and 83.0[plusmn]15.6 in non-athletes, p=0.04]. After controlling for lean mass and menarchal age, athletic status accounted for 5-9% of the variability in stiffness and failure load, while menarchal age accounted for 8-23%, and lean mass for 12-37%.[br]CONCLUSION: AA have decreased bone strength at the distal radius (non-weight-bearing site) compared with non-athletes, and lose the advantage of weight-bearing exercise seen in EA at the distal tibia.[br][br]Sources of Research Support: NIH grants 1 UL1 RR025758-01 and 1 R01 HD060827-01A1.[br][br]Nothing to Disclose: KEA, MP, GG, APT, LP, DBH, AK, MB, MM 2012-06-23T11:30:00 Theater C 2012-06-23T00:00:00 1899-12-30T11:30:00 51 42 105 OR09-2 OR30-01 Saturday 50 2012


51 ENDO12L_OR09-3 ORAL SESSION: Pediatric Endocrinology, General (11:15 AM-12:45 PM) Inhibition of Pref-1 by Estradiol in Adolescent Girls with Anorexia Nervosa Is Associated with Improvement in Lumbar Bone Mineral Density Alexander T Faje, Pouneh Fazeli, Debra K Katzman, Karen K Miller, Anne Breggia, Clifford J Rosen, Nara Mendes, Madhusmita Misra, Anne Klibanski Massachusetts General Hospital and Harvard Medical School, Boston, MA; Hospital for Sick Children and University of Toronto, Toronto, Canada; Maine Medical Center Research Institute, Portland, ME; Massachusetts General Hospital for Children and Harvard Medical School, Boston, MA Regulation of mesenchymal stem cell (MSC) differentiation into the osteoblast or the adipocyte lineage may influence bone formation. Adolescents with anorexia nervosa (AN) have low bone mineral density (BMD) and decreased bone formation. Preadipocyte factor 1 (Pref-1), an inhibitor of adipocyte and osteoblast differentiation, is elevated in states of estrogen deficiency and may negatively affect BMD in adolescent girls with AN.[br][bold]Objective:[/bold] To (i) compare levels of Pref-1 in adolescent girls with AN and controls, (ii) investigate the effects of transdermal estradiol on Pref-1 in AN, and (iii) examine associations of changes in Pref-1 with changes in lumbar BMD and markers of bone turnover.[br][bold]Design and Methods:[/bold] Sixty-nine girls (44 with AN and 25 normal-weight controls) 13-18 years of age were studied at baseline. Subjects were selected from a prior study based on their lumbar spine BMD Z-scores. The subset included all subjects with AN who had a baseline lumbar BMD Z-score less than -0.5 and normal-weight controls with lumbar BMD Z-scores between +1.0 and -1.0. Twenty-two AN girls with a bone age of [ge] 15 years were randomized to 100mcg transdermal estradiol (n = 13) or placebo (n = 9) in a double-blind study for 12 months. Subjects randomized to transdermal estradiol received medroxyprogesterone 2.5mg daily for 10 days each month. All subjects also received calcium carbonate (1200mg) and vitamin D (400 IU) daily.[br][bold]Results:[/bold] Although Pref-1 levels did not differ in AN versus controls (0.246 [plusmn] 0.015 vs. 0.267 [plusmn] 0.022 ng/ml, p = 0.46) at baseline, levels significantly decreased in girls with AN after treatment with transdermal estradiol compared with placebo (-0.015 [plusmn] 0.016 vs. 0.060 [plusmn] 0.026 ng/ml, p = 0.02). Changes in Pref-1 over twelve months correlated inversely with changes in lumbar BMD (r = -0.48, p = 0.02) and positively with changes in CTX (r = 0.73, p = 0.01). Levels of Pref-1 were inversely associated with estradiol levels in subjects with AN (r = -0.67, p = 0.004) after 12 months of treatment with transdermal estradiol or placebo.[br][bold]Conclusions:[/bold] One year of estrogen replacement suppresses Pref-1 in girls with AN. Additionally, Pref-1 levels correlate inversely with changes in BMD and positively with changes in CTX, a marker of bone resorption. Inhibition of Pref-1 may in part mediate the beneficial effects of transdermal estradiol replacement on bone mass in adolescent girls with AN.[br][br]Nothing to Disclose: ATF, PF, DKK, KKM, AB, CJR, NM, MM, AK 2012-06-23T11:45:00 Theater C 2012-06-23T00:00:00 1899-12-30T11:45:00 87 42 106 OR09-3 OR30-01 Saturday 51 2012


52 ENDO12L_OR09-4 ORAL SESSION: Pediatric Endocrinology, General (11:15 AM-12:45 PM) Effect of Oxandrolone Versus Placebo on Body Composition and Bone Density in Klinefelter Syndrome: Results of a Two-Year, Double-Blind, Placebo-Controlled Trial Martha Zeger Bardsley, Karen Kowal, Judith Levine Ross Thomas Jefferson University/Nemours, Philadelphia, PA [bold]Introduction:[/bold] Klinefelter[apos]s syndrome (KS) is a relatively common genetic disorder defined by the karyotype 47,XXY. Males with KS often have decreased muscle tone, increased central adiposity, and are at increased risk for metabolic syndrome. The KS phenotype may be the result of androgen deficiency in utero, infancy and childhood. Androgen replacement is standard in adolescent and adult KS males but has not been used earlier in childhood. The results presented are from a randomized clinical trial to study the effects of childhood androgen replacement on the KS neurocognitive and physical phenotypes.[br][bold]Objective:[/bold] To determine the effects of androgen treatment for two years in boys with KS on body composition, including muscle mass, body fat, and bone density.[br][bold]Study design:[/bold] This double-blind, placebo-controlled clinical trial (2005-2011, NCT00348946) randomized 93 boys, ages 4-12 yrs, to 2 groups: oxandrolone (Ox)(N=46) and placebo (Pl)(N=47). Protocol specified Ox dose was 0.06 mg/kg daily. Study visits occurred every 6 months for 2 years. Measurements included limb circumference, body fat percent by skin fold, BMI and weight. All measurements were converted to standard deviation scores. Bone mass was determined by use of the bone health index (BHI), using automated radiogrammetry (1). Statistical analysis included repeated measures ANCOVA and Fishers Exact test.[br][bold]Results:[/bold] 93 patients enrolled, and the 80 (86%) subjects who completed the 2 year study were compared. Their mean ages at 2 years were (Ox) 9.1[plusmn]2.1 years, n=39 vs. (Pl) 10.2 [plusmn]2.6 years, n=41, (P=0.03). Weight SDS increased in the Ox (0.3[plusmn]0.6) vs. Pl (-0.07[plusmn]0.4) (P[lt].05), but the change in BMI SDS did not differ (P=0.7). Fewer boys had body fat [gt] 25% in the Ox (23%) vs. Pl (49%) group (P=0.03). Increased muscle mass was demonstrated by increased lower leg circumference SDS in Ox group (change from baseline: 0.5[plusmn]0.7 Ox vs. 0.1[plusmn]0.5 Pl, P[lt]0.01). Mean BHI SDS change from baseline was 0.4[plusmn]0.7 (Ox) vs. -0.2 [plusmn]0.6 (Pl) (P [lt]0.01), indicating increased bone mass (cortical thickness) in the Ox group.[br][bold]Conclusions: [/bold]This unique double-blind, randomized clinical trial demonstrates that Ox increases weight SDS but not BMI SDS, and is associated with increased muscle mass, lower percent body fat, and increased bone cortical thickness. Therefore, gradual age-appropriate androgen replacement should be considered to optimize body composition in KS.[br][br](1)Thodberg HH, van Rign RR, Tanaka T, Martin DD, Kreiborg S, A paediatric bone index derived by automated radiogrammetry. Osteoporosis International 2009.[br][br]Sources of Research Support: This study was sponsored by NIH: Study #R01NS050597.[br][br]Disclosures: JLR: Scientific Board Member, Eli Lilly & Company; Speaker, Eli Lilly & Company. Nothing to Disclose: MZB, KK 2012-06-23T12:00:00 Theater C 2012-06-23T00:00:00 1899-12-30T12:00:00 1991 42 107 OR09-4 OR30-01 Saturday 52 2012


53 ENDO12L_OR09-5 ORAL SESSION: Pediatric Endocrinology, General (11:15 AM-12:45 PM) Optimum Macronutrient Content of the Diet for Adolescents with Pre-Diabetes; RESIST a Randomized Control Trial ACTRN12608000416392 Sarah P Garnett, Megan Dunkley, Mandy Ho, Louise A Baur, Manny Noakes, Kate Steinbeck, Helen J Woodhead, Kerryn Chisholm, Susie Burrell, Carolyn R Broderick, Robert Parker, Sukanya De, Shubha Shrinivasan, Geoffrey R Ambler, Michael R Kohn, Chris T Cowell The Children[apos]s Hospital at Westmead, Sydney, Australia; The Children[apos]s Hospital at Westmead, Sydney, Australia; University of Sydney, Sydney, Australia; CSIRO, Adelaide, Australia; Sydney Medical School, University of Sydney, Sydney, Australia; Sydney Children[apos]s Hospital at Campbelltown, Sydney, Australia; The Children[apos]s Hospital at Westmead, Sydney, Australia; The Children[apos]s Hospital at Westmead, Sydney, Australia; The Children[apos]s Hospital at Westmead, Sydney, Australia Rationale: Pre-diabetes and clinical insulin resistance in adolescents are rapidly emerging clinical problems with serious health outcomes. With appropriate management these conditions are potentially reversible. Yet there are no randomised trials of lifestyle interventions in this group to inform clinical practice.[br]Aim: To determine the efficacy and effectiveness of two structured lifestyle interventions, differing only in diet composition, on insulin sensitivity in obese adolescents with pre-diabetes and/or clinical features of insulin resistance.[br]Design: A 12 month RCT taking place at two tertiary hospitals in Sydney, Australia. At baseline adolescents are prescribed metformin and randomised to either a high carbohydrate low fat diet or a moderate carbohydrate increased protein diet. The program commences with an intensive 3 month dietary intervention, followed by a 3 month intensive exercise intervention and a 6 month maintenance phase (1).[br]Inclusion criteria: 10 to 17 year olds who are overweight or obese, with either pre- diabetes and/or clinical features of insulin resistance.[br]Clinical endpoints: Insulin sensitivity index (ISI) (2), adiposity, cardiometabolic profile and fitness. Differences in outcome measures overtime and between trial arms will be examined at 3, 6, 12 and 24 months.[br]Preliminary data: Recruitment is complete, n=111 (66 girls) and 101 have completed 3 months (4 dropped out and the remaining 6 will complete in February 2012). At baseline, most had a family history of obesity (87%), type 2 diabetes (75%) and/or a high risk ethnic background (58%) including Aboriginal, Maori and Pacific Islander. 94 (85%) had acanthosis nigricans and 14 (13%) had pre-diabetes. The mean BMI z-score was 2.3[plusmn]0.3, the median ISI was 1.3 [0.3 to 3.4 and 58 (52%) had SBP and/or DPB [ge] 90th centile, 56 (50%) had ALT and/or GTT [ge] 30 U/L and 70 (63%) had HDL cholesterol [le]1.03mmol/L and/or triglycerides [ge]1.7mmol/L.[br]Discussion: We have recruited an obese population with clinical and biochemical abnormalities placing them at high risk of developing type 2 diabetes. The three month outcomes including differences between dietary groups will be presented.[br][br](1)Garnett SP et al., BMC Public Health 2010; 10:575. (2)Matsuda M et al., Diabetes Care 1999; 22:1462.[br][br]Sources of Research Support: Diabetes Australia Research Trust 2008, BUPA Health, Foundation Australia Pty Limited (formerly MBF Foundation)2008 to 2012; Heart Foundation (#G08S3758)2009 to 2010. SG was supported by a National Health and Medical Research Council Australian Clinical, Research Fellowship(#457225)2007 to 2010 and is supported by a Cancer Institute NSW ECR Fellowship 2011 to 2013.[br][br]Nothing to Disclose: SPG, MD, MH, LAB, MN, KS, HJW, KC, SB, CRB, RP, SD, SS, GRA, MRK , CTC 2012-06-23T12:15:00 Theater C 2012-06-23T00:00:00 1899-12-30T12:15:00 729 42 108 OR09-5 OR30-01 Saturday 53 2012


54 ENDO12L_OR09-6 ORAL SESSION: Pediatric Endocrinology, General (11:15 AM-12:45 PM) Metabolic Syndrome Features in Children Born Post-Term Ahila Ayyavoo, Paul Hofman, Jose Derraik, Sarah Mathai, Peter Stone, Lynn Sadler, Wayne Cutfield University of Auckland, Auckland, New Zealand; National Research Centre for Growth and Development, Auckland, New Zealand; Auckland District Health Board, Auckland, New Zealand [bold]Introduction [/bold][br]We have recently shown from a large Swedish cohort that nearly half of the boys born post-term were overweight or obese at 16 years of age. In contrast, post-term girls were leaner at birth, but had similar weights to control girls at 16 years (Beltrand et al., J Pediatr 2011).[br][bold]Hypothesis[/bold][br]Pre-pubertal children born post-term have features of the metabolic syndrome.[br][bold]Methods[/bold][br]Two groups of healthy pre-pubertal children aged 4[ndash]11 years were studied: post-terms (n=36; born at [ge] 42 weeks gestation) and controls (n=56; 38-40 weeks). Following an overnight fast, we performed a frequently sampled intravenous glucose tolerance test with insulin. Insulin sensitivity (Si) was calculated using Bergman[apos]s minimal model. Biochemical markers of metabolic syndrome were examined. Auxological and DEXA-derived body composition data were obtained. Children also underwent 24-hour blood pressure (BP) monitoring. Data were analysed separately for boys and girls using linear mixed models, controlling for appropriate confounders. Data are expressed as mean[plusmn]SEM.[br][bold]Results[/bold][br][bold]Si[/bold] was reduced in both post-term boys (n=18; 9.7[plusmn]1.21 vs 14.2[plusmn]1.1x10[sup]-4[/sup] min[sup][minus]1[/sup][middot](mU/L); p=0.014) and girls (n=18; 7.3[plusmn]0.7 vs 10.4[plusmn]1.2 x 10[sup]-4[/sup] min[sup][minus]1[/sup][middot](mU/L)); p=0.046) compared to controls. There was an appropriate compensatory increase in [bold]acute insulin response[/bold] in post-term boys (549 [plusmn]101 vs 269[plusmn]25 mU/L; p=0.009), but not in post-term girls (378.5[plusmn]53.3 vs 277[plusmn]29.8 mU/L; p=0.65). [bold]IGFBP1[/bold] was lower in post-term boys (9.9ng/ml[plusmn]1.57 vs 18.31[plusmn]2.14 ng/ml; p=0.027) and girls (9.23[plusmn]1.47 vs 17.44[plusmn]2.14 ng/ml; p=0.08).[br]There were further changes suggestive of metabolic syndrome in post-term boys, including elevated [bold]IGFBP3[/bold] (3815 ng/ml [plusmn]241: 3313[plusmn]110 ng/ml; p=0.027), and reductions in both [bold]nocturnal systolic [/bold](8.2[plusmn]1.0 vs 13.8[plusmn]1.0 mmHg; p[lt]0.0001) and [bold]nocturnal diastolic [/bold](14.9[plusmn]0.9 vs 19.8[plusmn]1.3 mmHg; p 0.004) [bold]BP dip[/bold]. Post-term boys were also dyslipidemic, with higher [bold]total cholesterol[/bold] (4.4 [plusmn]0.3 vs 3.9[plusmn]0.1 mmol/l; p=0.058), [bold]LDL[/bold] (2.6[pusmn]0.2 vs 2.2[plusmn]0.1 mmol/l; p=0.069) and [bold]triglycerides[/bold] (0.84[plusmn]0.07 vs 0.7[plusmn]0.05 mmol/l; p=0.088) than boys born at term. Further, post-term boys had higher serum [bold]leptin[/bold] concentrations (5.9 [plusmn]1.4 vs 3.2[plusmn]0.4 ng/ml; p=0.056) despite similar body composition as boys born at term (21.8[plusmn]2.4 vs 17.5[plusmn]1.1 %; p=0.47).[br][bold]Conclusion[/bold]s[br]Post-term boys and girls have insulin resistance. Post-term boys have additional features of metabolic syndrome. We speculate that insulin resistance precedes obesity in post-term children.[br][br]Sources of Research Support: National Research Centre for Growth and Development; APEG Research Grants 2010 [amp] 2011.[br][br]Nothing to Disclose: AA, PH, JD, SM, PS, LS, WC 2012-06-23T12:30:00 Theater C 2012-06-23T00:00:00 1899-12-30T12:30:00 518 42 109 OR09-6 OR30-01 Saturday 54 2012


55 ENDO12L_S18-1 ORAL SESSION: Clinical Trial Symposium (3:45 PM-5:15 PM) Weight Loss, Exercise, or Both and Cardiometabolic Risk Factors in Obese Older Adults: Results of a Randomized Controlled Trial Matthew F Bouchonville, Krupa Shah, Reina Armamento-Villareal, David R Sinacore, Clifford Qualls, Dennis T Villareal University of New Mexico School of Medicine, Albuquerque, NM; University of Rochester Medical Center, Rochester, NY; Washington University School of Medicine, St Louis, MO; New Mexico Veterans Affairs Health Care System, Albuquerque, NM; University of New Mexico School of Medicine, Albuquerque, NM; Washington University School of Medicine, St Louis, MO; Washington University School of Medicine, St Louis, MO; New Mexico Veterans Affairs Health Care System, Albuquerque, NM; University of New Mexico School of Medicine, Albuquerque, NM [bold]Background: [/bold]In older adults, obesity exacerbates the age-related decline in physical and cardiometabolic health; however, the appropriate treatment for obese older adults is controversial. We recently reported that the combination of weight loss and exercise provides greater improvement in physical function than either intervention alone ([italic]N Engl J Med[/italic] 2011). We now report the results of the independent and combined effects of weight loss and exercise on cardiometabolic risk factors in this population.[br][bold]Methods: [/bold]In this 1-year randomized controlled trial, 107 obese (BMI [ge] 30 mg/kg[sup]2[/sup]) older (age [ge] 65 years) adults were randomly assigned to control, diet, exercise, and combined diet-exercise. The primary outcome was the change in insulin sensitivity index (ISI), measured from the oral glucose tolerance test (OGTT). Secondary outcomes included glucose and insulin response to the OGTT, waist circumference and abdominal adiposity, systolic and diastolic blood pressure (BP), and serum concentration of lipids and adipocytokines.[br][bold]Results: [/bold]Ninety-three participants (87%) completed the trial. In the intention to treat analyses, the ISI improved in the diet group (70%) and in the diet-exercise group (86%), but not in the exercise group or the control group (between-group [italic]P[/italic][lt]0.05). Accordingly, both the glucose and the insulin area under the curves during the OGTT improved in the diet group (10%) and in the diet-exercise group (15-20%), but not in the exercise or the control group (between-group [italic]P[/italic][lt]0.05). Moreover, systolic BP (-9% and -10%), diastolic BP (-8% and -6%), waist circumference (-7% and -7%), abdominal visceral fat (-25% and -33%), serum triglycerides (-26% and -27%), C-reactive protein (-27% and -27%), and TNF-R (-7% and -9%) decreased while adiponectin (21% and 33%) increased in the diet group and in the diet-exercise group, but not in the exercise group or the control group (all between-group P[lt]0.05). Body weight decreased in the diet group (10%) and in the diet-exercise group (9%) but did not decrease in the exercise group or the control group (between-group [italic]P[/italic][lt]0.05).[br][bold]Conclusions: [/bold]Diet-induced weight loss but not exercise training improves insulin sensitivity and multiple other cardiometabolic risk factors simultaneously in obese older adults. Therefore, in addition to additive effects of weight loss with exercise on physical function, weight loss is particularly important in obese older adults to decrease the risk for cardiometabolic syndrome.[br][br]Villareal DT et al., N Engl J Med 2011; 364:1218-29.[br][br]Sources of Research Support: RO1 Grant; National Institute of Aging; NIH Grant AG025501 awarted to DTV.[br][br]Nothing to Disclose: MFB, KS, RA-V, DRS, CQ, DTV 2012-06-23T15:45:00 362 2012-06-23T00:00:00 1899-12-30T15:45:00 2288 101 896 S18-1 OR300-01 Saturday 55 2012


56 ENDO12L_S18-2 ORAL SESSION: Clinical Trial Symposium (3:45 PM-5:15 PM) The Effect of Six Years of Denosumab Treatment on New Vertebral and Nonvertebral Fractures in Postmenopausal Women with Osteoporosis: Results from the FREEDOM Extension Trial Henry G Bone, Jacques P Brown, Roland Chapurlat, Nathalie Franchimont, Maria L Brandi, Edward Czerwinski, Marc-Antoine Krieg, Zulema Man, Dan Mellstrom, Sebastiao C Radominski, Jean Yves Reginster, Heinrich Resch, Jose A Roman, Nadia S Daizadeh, Andrea Wang, Michelle L Geller, Rachel B Wagman, Socrates Papapoulos Michigan Bone and Mineral Clinic, Detroit, MI; Laval University and CHUQ Research Centre, Quebec City, Canada; Universit[eacute] de Lyon, Lyon, France; Amgen Inc, Thousand Oaks, CA; University of Florence, Florence, Italy; Krakow Medical Center, Krakow, Poland; University Hospital of Lausanne, Lausanne, Switzerland; Centro TIEMPO, Buenos Aires, Argentina; Sahlgrenska University Hospital, G[ouml]teborg, Sweden; Universidade Federal do Paran[aacute], Curitiba, Brazil; University of Li[egrave]ge, Li[egrave]ge, Belgium; St Vincent Hospital, Vienna, Austria; Hospital Universitario La Fe, Valencia, Spain; Amgen Inc, Thousand Oaks, CA; Amgen Inc, Thousand Oaks, CA ; Leiden University Medical Center, Leiden, Netherlands The FREEDOM trial open-label extension is evaluating the long-term safety and efficacy of denosumab (DMAb) for up to 10 years. We report the intention-to-treat (ITT) results for women who participated in the first 3 years of the extension, representing up to 6 years of DMAb exposure. We also report the results for the pre-specified per protocol (PP) and modified per protocol (MPP) subsets.[br]During the extension, each woman is scheduled to receive 60 mg DMAb every 6 months and supplemental calcium and vitamin D daily. For the analyses here, women from the FREEDOM placebo group received 3 years of DMAb (cross-over group) and women from the FREEDOM DMAb group received 3 more years of DMAb for a total of 6 years (long-term group). The PP and MPP subsets excluded subjects who were non-compliant with the protocol and the MPP subset further excluded subjects who missed [ge] 2 doses of DMAb during the extension.[br]Of the 5928 women eligible for the extension, 4550 (77%) enrolled (N = 2207 cross-over; N = 2343 long-term). During the first 3 years of DMAb treatment during the extension, the cross-over group had significant gains in bone mineral density (BMD) at the lumbar spine and total hip, and further significant increases in BMD occurred over years 4 to 6 in the long-term group. Serum CTX was rapidly and similarly reduced after the 1[sup]st[/sup] (cross-over) or 7[sup]th[/sup] (long-term) DMAb dose with the characteristic attenuation observed at the end of the dosing period.[br]In FREEDOM, DMAb reduced the risk of new vertebral (2.3% DMAb vs 7.2% placebo) and nonvertebral (6.5% DMAb vs 8.0% placebo) fractures over 3 years. In the first 3 years of the extension, for the cross-over group, incidences of new vertebral (2.8%) and nonvertebral (5.6%) fractures were similar to the FREEDOM DMAb group. In the long-term group, fracture incidences remained low (3.5% for new vertebral and 3.8% for nonvertebral fractures). The fracture efficacy results for the PP and MPP subsets were consistent with the ITT analysis. Incidences of adverse events (AEs) and serious AEs did not increase over time with DMAb. No subjects developed neutralizing antibodies to DMAb.[br]DMAb treatment for 3 years in the cross-over group reproduced FREEDOM efficacy observations. DMAb treatment for 6 years (long-term group) remained well tolerated, maintained reduced bone turnover, and continued to significantly increase BMD. Fracture incidence remained low in the ITT, PP, and MPP populations.[br][br]Sources of Research Support: Amgen Inc. sponsored this study.[br][br]Disclosures: HGB: Consultant, Amgen, Merck & Co., Takeda, Tarsa; Investigator, Amgen, Merck & Co., Nordic Bioscience A/S, Novartis Pharmaceuticals, Takeda, Tarsa; Speaker Bureau Member, Amgen. JPB: Investigator, Amgen, Eli Lilly & Company, Merck & Co., Novartis Pharmaceuticals, Warner Chilcott Pharmaceuticals; Advisory Group Member, Amgen, Eli Lilly & Company, Merck & Co., Novartis Pharmaceuticals, Warner Chilcott Pharmaceuticals; Speaker, Amgen, Eli Lilly & Company, Novartis Pharmaceuticals. RC: Advisory Group Member, Amgen, Eli Lilly & Company, Servier, Merck & Co., Novartis Pharmaceuticals; Speaker, Amgen, Ipsen, Servier, Roche Pharmaceuticals. NF: Employee, Amgen. EC: Investigator, Eli Lilly & Company, Novartis Pharmaceuticals, Roche Pharmaceuticals, Amgen, Pfizer, Inc., Servier, Merck Serono S.A., Astra Zeneca, Ardea Biosciences, INC Research, Shire, Biotest AG, Andromeda Biotech Ltd., Johnson & Johnson; Speaker, Amgen, Roche Pharmaceuticals, Servier, Zentiva. SCR: Advisory Group Member, Novartis Pharmaceuticals, Aventis Pharmaceuticals; Study Investigator, Bristol-Myers Squibb, Amgen, Roche Pharmaceuticals, Pfizer, Inc. JYR: Advisory Group Member, Servier, Novartis Pharmaceuticals, Negma, Lilly USA, LLC, Wyeth Pharmaceuticals, Genzyme Corporation, Roche Diagnostics, Merck & Co., Nycomed, NPS, Theramex, UCB; Speaker, Merck Sharp and Dohme, Lilly USA, LLC, Rottapharm , IBSA, Genevrier, Novartis Pharmaceuticals, Roche Pharmaceuticals, GlaxoSmithKline, Teijin, Teva, Ebewee Pharma, Zodiac, Analis, Theramex, Nycomed, Novo Nordisk; Investigator, Bristol Myers Squibb, Merck Sharp and Dhome, Rottapharm, Teva, Lilly, Novartis, Roche, GlaxoSmithKline, Amgen, Servier. NSD: Employee, Amgen. AW: Employee, Amgen. MLG: Employee, Amgen. RBW: Employee, Amgen. SP: Medical Advisory Board Member, Amgen, Eli Lilly & Company, GlaxoSmithKline, Merck & Co., Novartis Pharmaceuticals; Speaker, Amgen, GlaxoSmithKline; Scientific Board Member, Merck & Co. Nothing to Disclose: MLB, M-AK, ZM, DM, HR, JAR 2012-06-23T16:00:00 362 2012-06-23T00:00:00 1899-12-30T16:00:00 1686 101 897 S18-2 OR300-01 Saturday 56 2012


57 ENDO12L_S18-3 ORAL SESSION: Clinical Trial Symposium (3:45 PM-5:15 PM) Efficacy of Recombinant Human Parathyroid Hormone (rhPTH[1[ndash]84]) for the Treatment of Adults with Hypoparathyroidism: A Randomized, Double-Blind, Placebo-Controlled Study John Bilezikian, Leif Mosekilde, Dolores Shoback, Tamara Vokes, Michael Mannstadt, Bart Clarke, Hjalmar Lagast, Roger Garceau Columbia University, College of Physicians [amp] Surgeons, New York, NY; Aarhus University Hospital, Aarhus, Denmark; San Francisco VA Medical Center, University of California, San Francisco, CA; University of Chicago, Chicago, IL; Massachusetts General Hospital and Harvard Medical School, Boston, MA; Mayo Clinic Rochester, Rochester, MN; NPS Pharmaceuticals, Bedminster, NJ Therapy for hypoparathyroidism (HypoPARA) is limited to symptomatic management, often with large amounts of calcium (Ca) and vitamin (Vit) D metabolites leading to concern for possible long-term complications such as end-organ damage. We conducted a randomized, double-blind, placebo (PBO)-controlled study to assess whether replacement therapy with recombinant human parathyroid hormone (rhPTH[1[ndash]84]) reduced Ca and Vit D needs, while normalizing or maintaining albumin-corrected total serum Ca concentration.[br]During a 2[ndash]16 week optimization period, oral Ca and active Vit D doses were adjusted to achieve and maintain serum Ca concentrations of 8.0[ndash]9.0 mg/dL for 2 weeks. Subjects received daily 50 [mu]g subcutaneous injections of rhPTH(1[ndash]84) or PBO for 24 weeks. The rhPTH(1[ndash]84) dose could be increased to 75 or 100 [mu]g if oral Ca and active Vit D doses were sufficiently high to allow for reduction. Regular serum Ca measurements influenced further adjustments in Ca and Vit D requirements in order to maintain serum Ca levels of 8.0[ndash]9.0 mg/dL. A [ldquo]responder[rdquo] was defined as subject whose need for oral Ca and active Vit D could be reduced by 50% at Week 24 while maintaining a serum Ca level of greater than or equal to baseline. Adverse events were monitored throughout treatment and during a 4-week follow-up period.[br]The total number of study subjects was 134 randomized (2:1) to rhPTH(1[ndash]84) or PBO (n=90 and n=44, respectively). A total of 84 subjects in the rhPTH(1[ndash]84) group and 37 in the PBO group completed the study. At the end of treatment, 48/90 (53.3%) subjects in the rhPTH(1[ndash]84) group and 1/44 (2.3%) in the PBO group were responders (treatment difference, 51.1; 95% confidence interval 39.9[ndash]62.3; [italic]P [/italic][lt] 0.001). The mean dose of oral Ca increased from baseline by 5.7% in the PBO group but decreased by 52.0% in the rhPTH(1[ndash]84) group ([italic]P [/italic][lt] 0.001). Active Vit D was reduced by 30.5% in the PBO group and 78.4% in the rhPTH(1[ndash]84) group ([italic]P [/italic][lt] 0.001). At study end, 37 (41.1%) subjects in the rhPTH(1[ndash]84) group and 1 (2.3%) in the PBO group achieved independence from Vit D and could be maintained on an oral Ca dose of [le]500 mg/day. Treatment with rhPTH(1[ndash]84) was generally well tolerated.[br]Treatment of HypoPARA with rhPTH(1[ndash]84) significantly reduces the need for large quantities of Ca and active Vit D, with a majority of responders becoming independent of Vit D and high-dose Ca.[br][br]Sources of Research Support: NPS Pharmaceuticals.[br][br]Disclosures: JB: Advisory Group Member, NPS; Principal Investigator, NPS. DS: Advisory Group Member, NPS; Investigator, NPS. TV: Advisory Group Member, NPS. MM: Advisory Group Member, NPS. BC: Advisory Group Member, NPS. HL: Employee, NPS. RG: Employee, NPS. Nothing to Disclose: LM 2012-06-23T16:15:00 362 2012-06-23T00:00:00 1899-12-30T16:15:00 1682 101 898 S18-3 OR300-01 Saturday 57 2012


58 ENDO12L_S18-4 ORAL SESSION: Clinical Trial Symposium (3:45 PM-5:15 PM) Achievement of Composite of Recommended Goals in Obese Subjects with Type 2 Diabetes Mellitus Using Extended-Release Phentermine/Topiramate in a 56-Week Weight Loss Intervention Donna H Ryan, W Timothy Garvey, Wesley W Day Pennington Biomedical Research Center, Baton Rouge, LA; University of Alabama at Birmingham, Birmingham, AL; Vivus, Inc, Mountain View, CA Current goals for management of type 2 diabetes mellitus (T2DM) include weight loss (WL) and reduction of HbA1c to [lt]6.5% and systolic blood pressure (SBP) to [lt]130 mg/dL. The 56-week, randomised CONQUER study in overweight/obese subjects with [ge]2 weight-related comorbidities showed significant WL with extended-release phentermine/topiramate (PHEN/TPM ER) vs placebo (PBO). This post-hoc analysis assessed percentage of subjects in CONQUER with T2DM at baseline who achieved the following composite goals (CG) by Week 56: CG-A, WL [gt]5%, HbA1c [lt]6.5%, SBP [lt]130 mm Hg; and CG-B, WL [gt]10%, HbA1c [lt]6.5%, SBP [lt]130 mm Hg. Percentage of subjects with net changes (% increase minus % decrease) in concomitant antidiabetic and antihypertensive medications at Week 56 was also captured. Subjects were managed to T2DM standards of care (diet, exercise; metformin monotherapy was permitted at baseline; addition of oral hypoglycaemics by physicians blinded to assignment was permitted to achieve established HbA1c targets [ADA]). In total, 357 subjects with T2DM at baseline with [ge]1 postbaseline assessment of WL, HbA1c, and SBP were included: lifestyle intervention plus PBO (n=144), PHEN 7.5 mg/TPM ER 46 mg (7.5/46; n=63), or PHEN 15 mg/TPM ER 92 mg (15/92; n=150). At baseline, 66% were female, mean age was 52.6 years, mean weight was 100.9 kg, mean HbA1c was 6.8%, and 70.8% (n=253) were using metformin. Mean SBP was 125.9 mm Hg (40.3% of subjects had SBP [ge]130 mm Hg). At 56 weeks, CG-A was achieved by 11.8%, 27.0%, and 39.3% and CG-B by 4.2%, 14.3%, and 31.3% of the PBO, 7.5/46, and 15/92 groups, respectively ([italic]P[/italic][lt].0001 between treatment groups). For subjects achieving CG-A or CG-B, percentage of subjects with a net change in concomitant antidiabetic and antihypertensive medications was neutral or increased for PBO and decreased in PHEN/TPM ER[ndash]treated subjects ([italic]P[/italic]=NS between treatment groups). In these subjects with T2DM, there were 6 reports of hypoglycaemia: 4 (3 mild, 1 severe) in PBO, 1 (mild) in 7.5/46, and 1 (mild) in the 15/92 group; none led to study-drug discontinuation. Discontinuation due to adverse events (AEs) in this population was 8%, 9%, and 19% for PBO, 7.5/46, and 15/92, respectively, with the most common AEs being constipation and paraesthesia. These data indicate that the WL achieved by PHEN/TPM ER[ndash]treated subjects can drive improvements in HbA1c and SBP, significantly increasing the achievement of composite goals recommended for the management of T2DM.[br][br]Sources of Research Support: VIVUS, Inc.[br][br]Disclosures: DHR: Consultant, Vivus USA, Novo Nordisk, Takeda. WTG: Medical Advisory Board Member, Daiichi-Sankyo, Johnson & Johnson, Vivus USA, Abbott Laboratories; Clinical Researcher, Merck & Co., Amylin Pharmaceuticals, Vivus USA, Abbott Laboratories, Daiichi-Sankyo; Stockholder, Merck & Co., Genzyme Corporation, Vivus USA, Bristol-Myers Squibb, Amylin Pharmaceuticals; Speaker Bureau Member, Merck & Co., Abbott Laboratories. WWD: Employee, Vivus USA. 2012-06-23T16:30:00 362 2012-06-23T00:00:00 1899-12-30T16:30:00 535 101 899 S18-4 OR300-01 Saturday 58 2012


59 ENDO12L_S18-5 ORAL SESSION: Clinical Trial Symposium (3:45 PM-5:15 PM) Testosterone Dose-Response Relationships in Surgically Menopausal Women: Effects on Body Composition and Muscle Strength in a Randomized Trial Grace Huang, Shalender Bhasin, Thomas G Travison, Maithili Davda, Matthew Ho, Thomas W Storer, Renee Miciek, Phillip E Knapp, Lauren Collins, Monica Ursino, Connie Dzekov, Shehzad Basaria Boston University School of Medicine, Boston, MA; Charles R Drew University of Medicine and Science, Los Angeles, CA [bold]Background:[/bold] Menopause has been associated with a loss of muscle mass and an increase in fat mass. Testosterone therapy is being considered to improve body composition in menopausal women. However, the dose-response relationships between testosterone and the changes in body composition and muscle strength in women have not been established.[br][bold]Objective: [/bold]To determine the dose-dependent effects of graded doses of testosterone on body composition and muscle strength in surgically menopausal women.[br][bold]Methods: [/bold]71 surgically menopausal women received a standardized estrogen regimen during the 12-week run-in period, and were then randomized to one of 5 groups to receive weekly IM injections of placebo (n=15), 3 (n=14), 6.25 (n=14), 12.5 (n=15) or 25 mg (n=13) testosterone enanthate for 24 weeks. Total and free testosterone levels were measured by LC-MS/MS and equilibrium dialysis, respectively. Lean body mass (LBM) and fat mass were measured using DXA scan and muscle strength was assessed by one-repetition maximum method at baseline and week 24.[br][bold]Results: [/bold]71 women were randomized. At baseline, mean age was 53 yrs, BMI 30 kg/m[sup2], total testosterone 13.7 ng/dl and free testosterone 2.2 pg/ml. On-treatment nadir testosterone concentrations were 14, 87, 106, 164 and 211 ng/dl at the 0, 3, 6.25, 12.5 and 25-mg doses, respectively. Changes in LBM were highly correlated with changes in testosterone concentrations; the estimated between-person difference in LBM was 0.67 kg per 100 ng/dl change in testosterone (95% CI: 0.23, 1.11; p = 0.003). There was a significant increase in LBM in the 25-mg dose group (average increase=1.7 kg, p= 0.01). No significant changes in fat mass and leg press strength were seen.[br][bold]Conclusion: [/bold]Testosterone administration in surgically menopausal women was associated with dose and concentration-dependent gains in LBM. Long-term randomized trials are needed to determine whether clinically meaningful improvements in other outcomes can be achieved safely with testosterone doses that do not induce virilization or worsen cardiovascular risk.[br][br]Nothing to Disclose: GH, SB, TGT, MD, MH, TWS, RM, PEK, LC, MU, CD, SB 2012-06-23T16:45:00 362 2012-06-23T00:00:00 1899-12-30T16:45:00 1982 101 900 S18-5 OR300-01 Saturday 59 2012


60 ENDO12L_S18-6 ORAL SESSION: Clinical Trial Symposium (3:45 PM-5:15 PM) Carbohydrate Modified Diet and Insulin Sensitizers Reduce Fasting Insulin and Body Weight, Modulate Measures of the Metabolic Syndrome, and Improve Adipokines in EMPOWIR (Enhance the Metabolic Profile of Women with Insulin Resistance) a Randomized Clinical Trial (NCT00618071) of Normoglycemic, Hyperinsulinemic Women with Midlife Weight Gain Harriette Rosen Mogul, Ruth Freeman, Philipp E Scherer, Michael Frey, Feras M Hantash, Ron Greenbaum, Sheila Jozak, LeeAnn Klein, Karen Tanenbaum New York Medical College, Valhalla, NY; Einstein, Bronx, NY; University of Texas Southwestern Medical Center, Dallas, TX; Quest Diagnostics, San Juan Capistrano, CA; Westchester Medical Center, Valhalla, NY [bold]BACKGROUND [/bold]Progressive midlife weight gain is associated with diabetes[sup]1 [/sup]and all-cause mortality[sup]2[/sup] and preventive interventions targeting high risk populations are critically needed. [bold]EMPOWIR[/bold], a double blind, placebo controlled, randomized clinical trial of metformin (MF) 2000mg [amp] MF plus 4 mg (low dose) rosiglitazone (RSG) was conducted to evaluate prior research on Syndrome [bold]W,[/bold] an early manifestation of insulin resistance, defined by the triad of [bold]W[/bold]eight gain ([ge]20 lbs after the 20[apos]s), [bold]W[/bold]aist gain, and [bold]W[/bold]hite Coat hypertension and normoglycemic hyperinsulinemia[sup]3-6 [/sup]and test the [bold]hypothesis [/bold]that insulin sensitizers plus carbohydrate modified diet could reduce BW and attenuate risks of diabetes and related comorbidities. [bold]METHODS [/bold]We compared fasting insulin (FIn) [amp] glucose (FBS), BW, HOMA, blood pressure (BP), lipids, adiponectin (Adp), [amp] leptin (Lp) at baseline [amp] 6-months in 46 subjects (mean age 46.6, BMI 30.5kg/m[sup]2[/sup], A1C 5.4%) meeting inclusion criteria: age 35-55; 20 lb wt gain; normal GTT with [uarr]AUC-insulin. Paired t-tests [amp] multivariate models were used (SPSS 19.0). Participants attended 4 weekly nutrition workshops (lead-in phase) at 2 study sites to introduce the EMPOWIR dietary intervention: a food exchange program (40% carbohydrates-40% protein-20% fat) promoting increased intake of vegetables, low-glycemic index fruits, low-fat protein [amp] dairy products, elimination of free sugars, [amp] restriction of 3 allowable additional carbohydrates (starches) to after 4PM. [bold]RESULTS[/bold] Significant reductions in mean BW were observed in all 3 study arms ([italic]P[apos]s[/italic]=.049,.005,.017). FBS declined significantly only in the MF arm (89.4[rarr]84.0mg/dl, [italic]P[/italic]=.034) FIn reduction was significant in the MF arm (12.5[rarr]8.0[micro]U/ml, [italic]P[/italic]= .011). HOMA decreased in MF [amp] MF+RSG arms (2.5[rarr]1.6 [amp]1.9[rarr]1.3, [italic]P[apos]s=[/italic].054 [amp].013). SBP decreased in the MF arm (114.3[rarr]107.2, [italic]P[/italic]=.001). HDL increased in placebo [amp] MF arms (49.3[rarr]56.3 [amp] 57.4[rarr]67.7 mg/dl, [italic]P[apos]s[/italic]=.016 [amp].026) without other significant lipid changes. Adp increased only in the MF+RSG arm (11.2[rarr]18.5[micro]g/ml, [italic]P[/italic][lt].001). Lp did not decline in subjects with BW loss (mean difference -0.17, [italic]P[/italic]=.907). MF was well tolerated with no reported adverse events. [bold]CONCLUSIONS [/bold]The EMPOWIR diet independently, and in combination with insulin sensitizers, reduced BW in a diverse cohort of normoglycemic, hyperinsulinemic midlife women and merits additional study. Measures of glucose homeostasis and metabolic syndrome improved significantly with MF, whereas Adp increased only in the MF + RSG study arm.[br][br]1. Colditz GA, Willett WC, Rotnitzky A, Manson JE. Weight gain as a risk factor for clinical diabetes mellitus in women. Ann Intern Med 1995; 122(7):481-486. 2. Manson JE, Willett WC, Stampfer MJ et al. Body weight and mortality among women. N Engl J Med 1995; 333(11):677-685. 3. Mogul HR, Peterson SJ, Weinstein BI, Zhang S, Southren AL. Metformin and carbohydrate-modified diet: a novel obesity treatment protocol: preliminary findings from a case series of nondiabetic women with midlife weight gain and hyperinsulinemia. Heart Dis 2001; 3(5):285-292. 4. Mogul HR, Peterson SJ, Weinstein BI, Li J, Southren AL. Long-term (2-4 year) weight reduction with metformin plus carbohydrate-modified diet in euglycemic, hyperinsulinemic, midlife women (Syndrome W). Heart Dis 2003; 5(6):384-392. 5. Mogul HR, Weinstein BI, Mogul DB et al. Syndrome W: a new model of hyperinsulinemia, hypertension and midlife weight gain in healthy women with normal glucose tolerance. Heart Dis 2002; 4(2):78-85. 6. Mogul HR, Freeman R, Hantash FX. Long-term hormonal adaptations to weight loss. N Engl J Med 2012; 366.[br][br]Sources of Research Support: EMPOWIR (ClinicalTrials.gov:NCT00618071) is an unsolicited, investigator-initiated study supported by Glaxo Smith Kline, with additional support from a Clinical and Translational Science Award (CTSA) from the National Institutes of Health(UL11RR025740)awarded to the GCRC at Einstein of Yeshiva University.[br][br]Disclosures: HRM: Principal Investigator, GlaxoSmithKline. FMH: Study Investigator, Quest Diagnostics. Nothing to Disclose: RF, PES, MF, RG, SJ, LK, KT 2012-06-23T17:00:00 362 2012-06-23T00:00:00 1899-12-30T17:00:00 778 101 901 S18-6 OR300-01 Saturday 60 2012


61 ENDO12L_SAT-1 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Primary Amenorrhea Due to Endometrial Hypoplasia in Three Consanguineous Sisters Cicilia LR Santos, Larissa S Nazareno, Angelica A Amato, Adriana L Porto University of Bras[iacute]lia, Bras[iacute]lia, Brazil Introduction: primary amenorrhea (PA) comprises a broad range of possible clinical conditions. We describe herein 3 consanguineous sisters with PA associated with endometrial hypoplasia (EmH), a rare condition of unknown etiology. Case reports: three sisters were referred to the Endocrinology Unit because of primary amenorrhea. They had normal pubertal development, were born from consanguineous healthy parents, and had no past history of systemic or inflammatory pelvic disease. General physical and gynecological examination were unremarkable. Serum TSH and prolactin levels were all normal, and their karyotype was 46,XX. Pelvic ultrasonography showed a slightly diminished uterus ([lt]25 cm3). Serial hormones profile and pelvic ultrasonografic imaging (USG) over 4 weeks showed: Case 1:33 years-old(yo), sparse, irregular vaginal bleeding after past use of estrogen-progestagen oral contraceptive. Hormones profile: First weed(W1): FSH 10,07mIU/ml; LH 4,9mIU/ml; Estradiol(E) 7,0pg/mL; Progesteron(P) 0,24 ng/mL; USG: endometrium(Em) 1mm, ovarian follicles (OFs) [lt] 10mm. Second week(W2): FSH 7,54; LH 6,6; E 22,88; P [lt] 0,21; USG: Em 1mm, maximum follicular diameter(OFmax)= 12mm. Third week(W3): FSH 16,47; LH 37,6; E 46,82; P 0,75; USG: Em 3mm, corpus luteum (CL) of 13mm. Fourth week(W4): FSH 5,2; LH 4,3; E 54,61; P 8,77; USG: Em 3mm, OFs of 2 and 3mm. Case 2:30yo, denied past hormonal treatment. W1: FSH 9,56; LH 21,0; E 87,88; P [lt] 0,21; USG: thin Em and OFmax= 12mm. W2: FSH 8,57; LH 15,6; E 63,74; P 7,25; USG: Em 1mm, OFs [lt]5mm. W3: FSH 3,26; LH 3,9; E 138,18; P 5,69; USG: Em not detected, OFs [lt]6mm. W4: FSH 12,34; LH 10,20; E 53,74; P 0,58; USG: Em 2.2mm, OFs [lt]6mm. Case 3:31yo, denied past hormonal treatment, had a unilateral salpingectomy due to a tubal pregnancy. W1:FSH 7,92; LH 3,9; E 10,11; P 0,23; USG:thin Em and OFs [lt]1mm. W2: FSH 10,07; LH 8,0; E 7,0; P [lt]0,21; USG: Em 1mm and OFs[lt]7mm. W3: FSH 7,89; LH 7,9; E 35,78; P [lt]0,21; USG:Em 1mm and OFmax =10mm. W4: FSH 6,13; LH 14,9; E 436,32; P 0,34; TV USG:Em 2.4mm, image compatible with cystic CL. Conclusions: We observed a hormonal cyclic pattern with P levels on the ovulatory range and USG image of CL in 2 of the 3 cases, whereas thin Em appearance remained unchanged. Altogether, these findings suggest EmH, probably associated with a monogenic disturbance. Possible candidates would be PAX8, a transcriptional factor involved in mullerian duct development, and WNT4, which has been associated with Rokitansky syndrome.[br][br]Nothing to Disclose: CLRS, LSN, AAA, ALP 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1544 49 122 SAT-1 PO32-01 Saturday 61 2012


62 ENDO12L_SAT-2 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Fetal Sulfate Supply in Human Gestation Paul A Dawson, Scott Petersen, Robyn L Rodwell, Phillip Johnson, Joanna Rakoczy, Francesca Grace, David G Simmons, H David McIntyre Mater Medical Research Institute, South Brisbane, Australia; Mater Mothers Hospital, South Brisbane, Australia; Mater Health Services, South Brisbane, Australia; University of Queensland, St Lucia, Australia; University of Queensland, South Brisbane, Australia Sulfate is an important nutrient for numerous cellular and metabolic processes in fetal development, including biotransformation of iodothyronines and steroids. In human gestation, maternal serum sulfate levels increase by approximately 2-fold, which enhances sulfate availability to the growing fetus. Sulfate is supplied from maternal circulation to the fetus via placental sulfate transporters. In this study, we measured venous cord plasma sulfate levels from healthy term pregnancies (altruistic cord blood donors), as well as maternal plasma sulfate levels in early ([lt]20wks) and late ([gt]30wks) gestation. Maternal plasma sulfate levels increased from (394[plusmn]12uM, n=43) in early pregnancy to (429[plusmn]12uM, n=33) in late gestation. Cord blood sulfate levels were lower in male babies (325[plusmn]16uM, n=31) when compared to female babies (371[plusmn]13uM, n=42). To build a model of directional sulfate transport in the placenta, we determined the relative abundance and spatial expression of all 10 sulfate transporter mRNAs in placentae (n= 10 male and 6 female babies) from uncomplicated healthy pregnancies at elective caesarean section. Quantitative real time PCR was used to determine the relative abundance of sulfate transporter mRNAs, and the spatial expression of mRNAs was determined by [italic]in situ[/italic] hybridisation with gene-specific DIG-labelled riboprobes. We detected very strong mRNA expression of [italic]SLC13A4[/italic] and [italic]SLC26A2[/italic], and intermediate levels of [italic]SLC26A1[/italic], [italic]SLC26A6[/italic] and [italic]SLC26A11[/italic]. Placental [italic]SLC13A4[/italic] mRNA was localised specifically to syncytiotrophoblasts, whereas [italic]SLC26A2[/italic] was detected in the cytotrophoblast cells. In summary, these data show high maternal plasma sulfate levels in late gestation when high sulfate levels are required for fetal development. In addition we show lower cord plasma sulfate levels from male babies when compared to female babies at birth, suggesting gender differences in sulfate requirements or placental sulfate transport. We identified [italic]SLC13A4[/italic] and [italic]SLC26A2[/italic] as the most abundant sulfate transporter mRNAs expressed in human placentae, suggesting a role for these 2 transporters in supplying sulfate from mother to fetus.[br][br]Sources of Research Support: Mater Medical Research Institute. Mater Mothers[apos] Hospital Golden Casket Research Grant.[br][br]Nothing to Disclose: PAD, SP, RLR, PJ, JR, FG, DGS, HDM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 86 49 123 SAT-2 PO32-01 Saturday 62 2012


63 ENDO12L_SAT-3 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Prolonged Pregnancy in Women Is Associated with Attenuated Myometrial Expression of Progesterone Receptor Co-Regulator Kr[uuml]ppel-Like Factor 9 John Mark P Pabona, Daying Zhang, David S Ginsburg, Frank A Simmen, Rosalia CM Simmen University of Arkansas for Medical Sciences, Little Rock, AR; Crozer Chester Medical Center, Upland, PA; Arkansas Children[apos]s Hospital Research Institute, Little Rock, AR Post-term pregnancy represents 3-5% of all complications during delivery and leads to both maternal and neonatal morbidity and mortality. The signaling mechanism(s) that maintains uterine quiescence during pregnancy and initiates the timely induction of uterine contractility at term remains unclear. Kr[uuml]ppel-like factor 9 (KLF9), an Sp-family member, is a progesterone receptor (PGR) interacting protein in uterine cells. In mice, KLF9 is robustly expressed in myometrial circular and longitudinal layers during late pregnancy. Further, mice null for [italic]Klf9[/italic] exhibit delayed parturition with increased gestational length and incidence of neonatal deaths. To examine the contribution of KLF9 in human parturition, we evaluated myometrial biopsies obtained from women during delivery, following procedures approved by the Institutional Board of Crozer Chester Medical Center and with signed consent. Participants with gestational age of 37 to [lt] 41 weeks were considered to have term pregnancy, while those with [ge]41 weeks were considered to exhibit prolonged pregnancy. Gene expression analysis showed that myometria from women with prolonged pregnancy (n=5) had decreased levels of KLF9, PGR-B and myosin light chain kinase transcripts, when compared to those of women with term pregnancy (n=8). The reduction in transcript levels was confirmed at the level of nuclear KLF9, PGR-A and PGR-B proteins and was accompanied by lower PGR-A/PGR-B ratio (P[lt]0.05). The loss of total PGR expression in myometria of patients with prolonged, relative to those with term pregnancies was also confirmed by immunohistochemistry. Estrogen receptor (ER) [alpha] expression, both at the level of mRNA and nuclear protein, was also lower for myometria of women with prolonged vs. term pregnancies. Myometrial levels of total and phosphorylated NF-[kappa]B (p65) proteins as well as of PGF2[alpha] receptor transcripts were similarly attenuated in women with prolonged relative to those with term pregnancies. In contrast, transcript levels for oxytocin receptor and inducible nitric oxide synthase did not differ between the two groups. Loss of myometrial KLF9 may aberrantly influence PGR, ER[alpha] and NF- [kappa]B signaling networks to promote uterine quiescence and delay labor. Our findings suggest that prolonged pregnancy associated with a deficiency in KLF9 expression may be a conserved phenomenon in mice and humans and importantly, that KLF9 has a physiological and pathological role in the timely onset of parturition.[br][br]Sources of Research Support: NIH-HD21961.[br][br]Nothing to Disclose: JMPP, DZ, DSG, FAS, RCMS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 240 49 124 SAT-3 PO32-01 Saturday 63 2012


64 ENDO12L_SAT-4 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Incident Metabolic Syndrome Is Not Increased in Women with Hyperandrogenemic Oligomenorrhea across Menopausal Transition: Longitudinal Follow-Up from the Study of Women[apos]s Health Across the Nation (SWAN) Alex J Polotsky, Amanda Allshouse, Sybil L Crawford, Sioban D Harlow, Naila Khalil, Rasa Kazlauskaite, Nanette Santoro, Richard Legro University of Colorado, Aurora, CO; University of Massachusetts, Worcester, MA; University of Michigan, Ann Arbor, MI; Boonshoft School of Medicine, Wright State University, Dayton, OH; Rush University Medical Center, Chicago, IL; Pennsylvania State College of Medicine, Hershey, PA Reproductive-age women with oligomenorrhea (Oligo) and hyperandrogenemia (HA), features of Polycystic Ovary Syndrome (PCOS), exhibit adverse cardiovascular risks as compared to age-matched controls. Recent data suggest that menstrual irregularity(1), HA(2) and insulin resistance(3) may improve with aging in PCOS women approaching menopause. Cross-sectional analysis of 2543 SWAN women at baseline showed that HA but not history of Oligo was independently associated with the risk of prevalent metabolic syndrome (MetS) (4). Persistence of metabolic risks after menopause is not well understood.[br][bold]Objective[/bold]: To evaluate metabolic health in women with history of HA and Oligo across menopausal transition.[br][bold]Methods[/bold]: SWAN is a multi-ethnic cohort of over 3000 US women as they traverse menopause with mean age at baseline of 45.8 years. Current analysis includes 8-10 years of follow-up. History of [ge] 2 instances of non-gestational/non-lactational amenorrhea for [ge] 3 months was classified as Oligo. The highest tertile of serum testosterone was set as HA. MetS was defined by NCEP ATP III criteria. Cox Hazard Ratios (HR) of MetS were estimated, with adjustment for age, ethnicity, BMI, smoking, menopausal stage [amp] study site.[br][bold]Results[/bold]: Among 1932 SWAN subjects who were MetS-free at baseline, 409 new cases of MetS were identified during 12,363 woman-years of follow-up over 8 years. Mean age at natural menopause was 51.3 years among 734 women who were at least 12 months past their final menstrual period. Women with HA and Oligo developed incident cases of MetS at a similar rate compared to their counterparts: HR of 1.1 (0.9-2.3) vs. eumenorrhea and normal androgens; HR of 1.2 (0.7-2.0) vs. Oligo and normal androgens; HR of 1.2 (0.7-1.9) vs. HA and eumenorrhea. Women with HA and Oligo had a similar trajectory of change for all individual components comprising MetS. Smoking and obesity were the strongest predictors of incident MetS: HR of 1.5 (1.1-1.9) for current smoking [amp] HR of 1.8 (1.7-1.9) per each 5 units of BMI.[br][bold]Conclusions[/bold]: While women acquire MetS at a more rapid rate after menopause, SWAN participants with history of HA and Oligo did not develop incident MetS more frequently than those without these characteristics. This suggests that history of androgen excess and menstrual irregularity in early life does not remain an independent risk for metabolic health after menopause. Attention to modifiable risks of obesity and smoking is warranted regardless of the PCOS diagnosis.[br][br](1)Elting MW et al., Human Reproduction 2000; 15:24. (2)Winters SJ et al., Fertil Steril 2000. 73: 724. (3)Brown ZA et al., Fertil Steril 2011; 96: 1259. (4)Polotsky AJ et al., Fertil Steril 2011; 96: S42.[br][br]Sources of Research Support: SWAN has grant support from the NIH, DHHS, through the NIA, the NINR and the ORWH (Grants NR004061; AG012505, AG012535, AG012531, AG012539, AG012546, AG012553, AG012554, AG012495). The content of this abstract is solely the responsibility of the authors and does not necessarily represent the official views of the NIA, NINR, ORWH or the NIH.[br][br]Nothing to Disclose: AJP, AA, SLC, SDH, NK, RK, NS, RL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 276 49 125 SAT-4 PO32-01 Saturday 64 2012


65 ENDO12L_SAT-5 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Aromatase Inhibition Normalizes Luteal Function but Dramatically Reduces Estrone Conjugate Excretion in Obese Women Lauren A Ross, Alex J Polotsky, Alexander Kucherov, Andrew P Bradford, Jennifer Lesh, Justin Chosich, Nanette Santoro University of Colorado Denver, Aurora, CO; Albert Einstein College of Medicine, Bronx, NY Adult female obesity is associated with menstrual cycle irregularities, subfertility and a hypogonadotropic phenotype. We have previously shown that aromatase inhibitor (AI)-treated obese women exhibit increased gonadotropin production similar to untreated normal weight women (1).[br][bold]Objective:[/bold] To assess the mechanism of action of AI in obesity, we examined reproductive hormone excretion across the menstrual cycle in obese women compared to normal weight women, after treatment with an AI.[br][bold]Methods:[/bold] We administered a 7-day, weight-adjusted dose of letrozole to 22 regularly cycling women: 12 obese (BMI, 37.1[plusmn]7.0kg/m2) and 10 normal weight (BMI, 21.2[plusmn]1.3kg/m2). Urine was collected daily over the course of the treatment cycle and assayed for luteinizing hormone (LH) [amp] follicle stimulating hormone (FSH; using the DELFIA immunofluorometric assay platform), and estrone conjugates (E1c) [amp] pregnanediol glucuronide (Pdg; using an in-house ELISA). All hormones were normalized to creatinine, and the day of ovulation was determined using a validated algorithm (2).[br]LH, FSH, E1c and Pdg were estimated for the whole cycle as well as follicular and luteal phases by computing sums of individual hormones. Groups were compared by t and Mann-Whitney tests as appropriate.[br][bold]Results[/bold]: Age did not differ between obese and normal weight women (30.5[plusmn]4.3 years vs. 30.9[plusmn]1.0 years).[br]Whole cycle E1c was half the level of controls in the obese women [331(259-465) mIU/mg Cr vs. 775(695-2180) mIU/mg Cr for obese and normal weight groups, respectively, p[lt]0.01] as was average E1c (p[lt]0.01) and peak E1c (p[lt]0.01).[br]Whole cycle Pdg (12.3 (5.2-16.2) ug/mgCr vs. 12.5 (9.2-33.8) ug/mgCr, p=0.47); LH (242 [plusmn]144 mIU/mg Cr vs. 335[plusmn]166 mIU/mg Cr, p=0.18); FSH (179[plusmn]95 mIU/mg Cr vs. 137[plusmn]59 mIU/mg Cr, p=0.23) did not differ between obese and normal weight women, respectively, nor did they differ by follicular or luteal phase means for these hormones.[br][bold]Conclusions[/bold]: Normalization of gonadotropin output and luteal function occurs at the expense of drastically reduced E1c excretion in AI-treated obese women as compared to normal weight AI-treated controls. These findings suggest that the adiposity-related reproductive phenotype is receptive to modulation of estrogen dynamics. Obese women demonstrate increased sensitivity to interruption of estrogen feedback with AI, implying that female obesity is a state of enhanced estrogen negative feedback at the hypothalamic-pituitary level.[br][br]1. Polotsky AJ et al. Endocr Rev 2011;32:S12. 2. Santoro N et al. JCEM 1996;81:1495.[br][br]Sources of Research Support: NIH U54 HD058155 Center for the Study of Reproductive Biology; K24 HD041978 to NS; 1UL1 RR025780 (University of Colorado CTRC).[br][br]Nothing to Disclose: LAR, AJP, AK, APB, JL, JC, NS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 902 49 126 SAT-5 PO32-01 Saturday 65 2012


66 ENDO12L_SAT-6 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Effects of Vaginal Estriol Cream on Circulating Hormones in Normal Cycling Women K-Y Francis Pau, Heidi Jones, Ilana Dzuba, Susan A Ballagh, Tracy Irwin, Bryna Harwood, Wesley H Clark, Yael Swica, Beverly Winikoff Oregon National Primate Research Center, Oregon Health [amp] Sciences University, Beaverton, OR; CUNY School of Public Health, Hunter College, New York, NY; Gynuity Health Projects, New York, NY; Los Angeles County Harbor-UCLA, Torrance, CA; University of Illinois at Chicago, Chicago, IL; New York Presbyterian Hospital, New York, NY; Columbia University College of Physicians and Surgeons, New York, NY [bold]Background:[/bold] Studies in ovariectomized non-human primates suggest that estriol (E3) administered topically protects against SIV transmission (1). In humans, HIV transmission is higher in premenopausal women during the luteal phase and pregnancy when progesterone (P4) levels are high (2). As part of a Phase 1 randomized, placebo-controlled, double-blinded trial to evaluate the effect of a topical E3 cream on vaginal health, this report examines the cream[apos]s effect on systemic hormones.[br][bold]Methods:[/bold] Of the 102 eligible consenting women aged 18-40 years, 82 completed 4 study visits in both follicular and luteal phases before and after randomization to 4.0mg estriol cream (1mg estriol/1ml cream) (n=38) or matching placebo (n=44). A single blood sample was taken at each visit for detection of estrone (E1), estradiol (E2), E3, P4, luteinizing hormone (LH) and follicle-stimulating hormone (FSH). Data were analyzed with SPSS and compared by paired rank sum test.[br][bold]Results:[/bold] Serum E3 was non-detectable ([lt]0.07 ng/ml) in all subjects during baseline and among those assigned to placebo. E3 cream increased serum E3 level in 10 subjects (26.3%) during the follicular phase (0.09 +/- 0.01 ng/ml, range 0.08-0.23 ng/ml) and 11 (28.92%) during the luteal phase (0.09 +/- 0.01, range 0.08-0.30 ng/ml). None of the other 5 hormonal levels changed (p[gt]0.05) from baseline in the follicular phase. During the luteal phase, P4 levels were lower with E3 cream compared to placebo (median: 1.24 vs 8.69 ng/ml, p[lt]0.009; mean: 3.92 vs 7.17 ng/ml, p[lt]0.004). Of the 11 subjects with increased E3, 6 had P4 levels below the mean. Baseline P4 levels during the luteal phase did not differ between the two groups (median: 5.64 vs 6.60 ng/ml, p= 0.61).[br][bold]Conclusions:[/bold] Consistent with findings in non-human primates, these results show that a small amount of E3 can be found in the circulation of less than 30% of users in either the follicular or luteal phase. While vaginal E3 cream has little effect on circulating E1, E2, LH and FSH, serum P4 levels are reduced during the luteal but not the follicular phase. The relationship of E3 and P4 levels and their association with menstrual cycle changes remain to be explored.[br][br](1)Smith SM, Mefford M, Sodora D, Klase Z, Singh M, Alexander N, Hess D, Marx PA. Topical estrogen protects against SIV vaginal transmission without evidence of systemic effect. AIDS. 2004 Aug 20;18(12):1637-43. (2)Mishell DR, Davajan V. Infertility, contraception, and reproductive endocrinology. Montvale, NJ: Medical Economics, 1986.[br][br]Sources of Research Support: The United States Agency for International Development (USAID) through the International Partnership for Microbicides.[br][br]Nothing to Disclose: K-YFP, HJ, ID, SAB, TI, BH, WHC, YS, BW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 931 49 127 SAT-6 PO32-01 Saturday 66 2012


67 ENDO12L_SAT-7 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Factors within the Intravascular Compartment Associate with Progression of Brain White Matter Hyperintensity in Women of the Kronos Early Estrogen Prevention Study (KEEPS) Limor Raz, Kejal Kantarci, Nirubol Tosakulwong, Timothy G Lesnick, Muthuvel Jayachandran, Samantha M Wille, Matthew C Murphy, Matthew L Senjem, Clifford R Jack, Virginia M Miller Mayo Clinic, Rochester, MN; Mayo Clinic, Rochester, MN; Mayo Clinic, Rochester, MN; Mayo Clinic, Rochester, MN White matter hyperintensities (WMH) on brain MRI correlate with a future risk for mild cognitive impairment in elderly populations of men and women. Relationships of WMH with vascular disease in younger but vulnerable populations such as menopausal women remain unexplored. Our objective was to investigate relationships among conventional and novel cardiovascular (CV) risk factors regarding the presence and progression of WMH in recently menopausal women (within 6 months [ndash] 3 yrs of their last menses) enrolled in the Kronos Early Estrogen Prevention Study (KEEPS) at Mayo Clinic (n=95). Conventional CV risk factors (systolic blood pressure, plasma lipids, glucose, triglycerides and body mass index) and activated cell membrane-derived microvesicles (MVs) were assessed in each woman prior to brain MRI at baseline (prior to randomization to treatment groups), and at 36 and 48 months (mo) after randomization. MRI images included a Fluid Attenuated Inversion Recovery (FLAIR) sequence from which WMH volumes were calculated using a semi-automated segmentation algorithm. Changes in WMH volume were quantified from baseline to 36 and 48 mo. Linear regression models adjusted for age, time past menopause and APOE [epsilon]4 status were used to test for association of CV factors with baseline WMH and changes in WMH. The median [inter-quartile range (IQR)] of WMH at baseline was 1909 (1538, 2655) mm[sup]3[/sup]. Volumes of WMH were not associated with conventional CV risk factors at baseline. Volumes of WMH tended to increase over time; median (IQR) volume changes were 101 (-165, 510) mm[sup]3[/sup] and 155 (-89, 567) mm[sup]3[/sup] at 36 and 48 mo, respectively. These increases in WMH associated with numbers of platelet-derived and thrombogenic MVs, (those positive for phosphatidylserine) at baseline, but not with conventional CV risk factors. These results suggest that activated platelets and cell membrane-derived thrombogenic MVs within the vascular compartment may affect development of WMH within the brain as has been suggested for their involvement in anatomical changes in other organs such as arterial intimal medial thickening and arterial calcification in the heart.[br][br]Sources of Research Support: The Aurora Fnd, NIH HL90639, NS066147, HD65987 and the Mayo Fnd.[br][br]Nothing to Disclose: LR, KK, NT, TGL, MJ, SMW, MCM, MLS, CRJ, VMM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 568 49 128 SAT-7 PO32-01 Saturday 67 2012


68 ENDO12L_SAT-8 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) The Pathogenic Role of Novel Steroid Receptor Coactivator-1 C-Terminal Isoform in Endometriosis Progression Sang Jun Han, Shannon M Hawkins, Khurshida Begum, Sung Yun Jung, Ertug Kovanci, Jun Qin, John P Lydon, Francesco J DeMayo, Bert W O[apos]Malley Baylor College of Medicine, Houston, TX; Baylor College of Medicine, Houston, TX; Baylor College of Medicine, Houston, TX Approximately 10% of women of reproductive age in the United States suffer from the severe symptoms of endometriosis. However, almost nothing is known of its molecular etiology. Although endometriosis is considered as an estrogen-dependent inflammatory disease, the role of coactivators (SRCs), key regulators of estrogen receptor signaling, has not been elucidated in endometriosis progression. Here, we reveal that a previously undiscovered isoform of SRC-1 works as a [apos]causative molecular regulator[apos] in endometriosis progression and has the following unique molecular properties:[bold]1) [/bold]A novel 70-kDa SRC-1 proteolytic isoform has been discovered that migrates to the cytoplasm and is highly elevated in endometriotic tissue of mice with surgical induced endometriosis - and also in human endometriotic stromal cells isolated from ectopic lesions of women with endometriosis; [bold]2)[/bold] TNFalpha activates the MMP9 protease to process SRC-1 to this 70-kDa C-terminal isoform (791-1441 aa.) within endometriotic tissue; [bold]3)[/bold] To promote endometriotic tissue growth, the processed 70-kDa SRC-1 C-terminal isoform then acts mechanistically by protecting human endometrial cells from TNFalpha-induced [apos]apoptosis[apos] by directly interacting with, and inhibiting, procaspase 8 and importantly, it also enhances the epithelial-mesenchymal transition of human endometrial cells to promote their invasive capacity. Thus, this TNFalpha/MMP9/SRC-1-isoform axis functions as a newly discovered pathogenic pathway for endometriosis progression; the isoform could be employed as a molecular marker and/or therapeutic target for endometriosis.[br][br]Sources of Research Support: R01 HD08188 to B. W. O[apos]Malley and U54HD007495 pilot grant to S.J. Han.[br][br]Nothing to Disclose: SJH, SMH, KB, SYJ, EK, JQ, JPL, FJD, BWO 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 588 49 129 SAT-8 PO32-01 Saturday 68 2012


69 ENDO12L_SAT-9 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Racial/Ethnic Differences in Sex Hormone Levels among Postmenopausal Women in the Diabetes Prevention Program (DPP) Catherine Kim, Sherita Golden, Kieren Mather, Gail A Laughlin, Shengchun Kong, Bin Nan, Elizabeth Barrett-Connor, John Randolph, Diabetes Prevention Program (DPP) University of Michigan, Ann Arbor, MI; Johns Hopkins University, Baltimore, MD; Indiana University, Indianapolis, IN; University of California, San Diego, La Jolla, CA; George Washington University, Washington, DC Context: Studies in mid-life women conflict regarding racial/ethnic differences in sex hormones: sex hormone binding globulin (SHBG), total and bioavailable estradiol (E2), total and bioavailable testosterone (T), and dehydroepiandrosterone (DHEA). Such differences may contribute to racial/ethnic variation in the incidence and consequences of sex hormone-sensitive comorbidities in older women, such as gynecological malignancies.[br]Objectives: We compared sex hormones by race/ethnicity in postmenopausal non-Hispanic white (NHW), Hispanic, and African-American (AA) women in the Diabetes Prevention Program (DPP).[br]Methods: The DPP was a randomized controlled trial of intensive lifestyle modification vs. metformin (850 mg twice a day) vs. placebo for diabetes prevention. We examined postmenopausal glucose-intolerant women participating in the DPP who did (n=310) or did not (n=370) use oral estrogen, with and without progestin, at baseline and 1-year follow-up. Postmenopause was defined as the absence of menses at baseline for at least one year without gynecologic surgery, bilateral oophorectomy, or age [gt]= 55 years and hysterectomy. Main outcome measures were baseline and 1-year change in serum levels of SHBG, total and bioavailable E2 (bioE2), total and bioavailable T, and DHEA.[br]Results: Among women not using estrogen, NHW had higher baseline total and bioavailable E2 and T levels than Hispanic women independent of age, type of menopause, waist circumference, alcohol intake, and current smoking. NHW also had higher levels of baseline bioE2 and lower levels of SHBG than AA. Among oral estrogen users, NHW had higher baseline total E2 and bioE2 than Hispanics, and higher levels of SHBG than AA independent of the same covariates. At 1-year, after adjustment for randomization arm, change in waist circumference, and the above covariates, NHW women not using estrogen had larger declines in total E2 and bioE2 levels than AA. Changes in sex hormones at 1-year did not differ by race/ethnicity among women using estrogen.[br]Conclusions: Among postmenopausal glucose-intolerant women, there were significant race/ethnicity differences in sex hormones and changes in sex hormones. Higher sex hormone levels in NHW women are consistent with reports of a higher incidence of hormone-sensitive cancers in NHW women compared to Hispanics and AA. Whether the greater reduction in E2 at 1-year in NHW results in a preferential reduction in risk in this group warrants further study.[br][br]Sources of Research Support: NIH (NIDDK).[br][br]Nothing to Disclose: CK, SG, KM, GAL, SK, BN, EB-C, JR 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 614 49 130 SAT-9 PO32-01 Saturday 69 2012


70 ENDO12L_SAT-10 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Relations between Soluble P-Selectin and Sex Hormones in Postmenopausal Women in the Framingham Offspring Study Pornpoj Pramyothin, Thomas G Travison, Shalender Bhasin, Andrea D Coviello Boston University School of Medicine and Boston Medical Center, Boston, MA; Boston University School of Medicine and Boston Medical Center, Boston, MA Context: Soluble P-selectin (sP-sel), a cell-adhesion molecule with key roles in inflammation and coagulation, may contribute to cardiovascular disease (CVD) risk. sP-sel levels are higher in postmenopausal women but lower in women taking hormone therapy (HT). The relations between sP-sel and sex hormone levels in post-menopausal women is not understood.[br]Objective: To determine the relations between circulating levels of sP-Sel and endogenous sex hormones in postmenopausal women not taking HT: total (TT) and free (FT) testosterone, total (E2) and free (FE2) estradiol, total (E1) and free (FE1) estrone and their primary binding protein sex-hormone binding globulin (SHBG).[br]Methods: Multivariable linear regression models were used to determine the cross-sectional associations between circulating sP-sel and endogenous sex hormones in postmenopausal women who attended exam 7 (1998-2001) of the Framingham Offspring Study (FOS) who were not taking HT (n=875). Models were adjusted for factors associated with sex hormones and CVD risk including age, body mass index (BMI), smoking, insulin resistance, hypertension, cholesterol levels and statin use. Due to the inflammation associated with diabetes (DM), subgroup analyses were performed in women without DM (n=763). sP-sel was measured by R[amp]D enzyme-linked immunoassay; SHBG by immunoassay(Delfia-Wallac); TT, E2, and E1 by LC-MSMS; and FT, FE2, and FE1 were calculated.[br]Results: Mean[plusmn]SD age was 65[plusmn]8 yrs and mean[plusmn]SD BMI 28.0[plusmn]5.7 kg/m2. The prevalence of hypertension was 38%, hyperlipidemia 22%, metabolic syndrome 40%, diabetes 12.8%, and CVD 4%. In adjusted analyses, a 10% increase in sP-sel was associated with a 0.9% decrease in SHBG (p=0.03) and a 1.8% increase in E2 (p[lt]0.01) but not with changes in E1 (p=0.1). Further adjustment for SHBG did not alter results. A 10% increase in sP-sel was associated with a 2.4% increase in FE2 (p[lt]0.01) and 1.2% increase in FE1 (p=0.02). Adjustment for SHBG attenuated these associations to 1.9% (p[lt]0.01) for FE2 and 1% (p=0.04) for FE1. sP-sel was not associated with TT or FT in adjusted models. Exclusion of women with DM yielded similar results.[br]Conclusions: Circulating sP-sel levels were negatively associated with SHBG and positively associated with total and free E2 as well as FE1 in postmenopausal women not on HT. Defining the connection between inflammation and coagulation pathways and estrogens will lead to a better understanding of CVD in post-menopausal women.[br][br]Sources of Research Support: NIH Grant RO1 HL094755 (to A.D.C., S.B., and R.S.V.).[br][br]Nothing to Disclose: PP, TGT, SB, ADC 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 660 49 131 SAT-10 PO32-01 Saturday 70 2012


71 ENDO12L_SAT-11 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Anti-Mullerian Hormone and Its Specific Receptor Are Differentially Regulated by Gonadotropins in Normo and Dysovulatory Women with the Polycystic Ovarian Syndrome Alice Pierre, Maeliss Peigne, Michael Grynberg, Nassim Arouche, Joelle Taieb, Laetitia Hesters, Jean-Yves Picard, Didier Dewailly, Renato Fanchin, Sophie Catteau-Jonard, Nathalie di Clemente INSERM, Clamart, France, Metropolitan; Universit[eacute] de Lille II, Lille, France, Metropolitan Anti-Mullerian hormone (AMH) is a member of the transforming growth factor family well known for its role during male sexual differentiation. However, AMH and its specific receptor AMHR-II are also expressed by granulosa cells of growing follicles. In rodents, AMH exerts a repressive role on folliculogenesis, but up to now, this effect has not been confirmed in human. These last years, serum AMH has been recognized as a useful tool in reproductive endocrinology, as a reliable marker of the ovarian follicular status and predictor of the ovarian response to controlled ovarian hyperstimulation. In particular, serum AMH is elevated in women with the polycystic ovarian syndrome (PCOS), the most common cause of female infertility, which is characterized by an increased number of small follicles. We had previously shown that AMH and AMHR-II expression are up-regulated in granulosa cells from PCOS women undergoing in vitro fertilization treatment, in keeping with a repressive role of AMH on follicle growth in these women. In this work, using real-time RT-PCR and ELISA experiments, we compared the regulation by gonadotropins of AMH and AMHR-II expression in primary culture of granulosa cells from control and normo and dysovulatory PCOS women. We show that AMH mRNAs are up-regulated by FSH in the three groups of women (+70% p=0.0068 and p=0.0469 for control and dysovulatory PCOS women respectively, and +40% p=0.0313 for normo ovulatory PCOS women), whereas there are stimulated by LH only in granulosa cells from dysovulatory PCOS women (+120% p=0.0391). Similarly, AMHR-II expression is unaffected by FSH in the three groups of women, whereas it is down-regulated by LH in control and normo ovulatory PCOS women (-30% p=0.0049 and -40% p=0.0469 respectively), but not in dysovulatory ones. Because LH levels are often elevated in PCOS women, our results could explain the increase of AMH expression by their granulosa cells. The reason why LH does not repress AMHR-II expression in dysovulatory PCOS cells is unknown. Altogether, our results demonstrate that LH regulation of the AMH/AMHR-II system is altered in PCOS granulosa cells, maintaining them in an immature stage. Because this dysregulation is associated with dysovulation in PCOS women, our results are in keeping with a role of AMH in this process.[br][br]Nothing to Disclose: AP, MP, MG, NA, JT, LH, J-YP, DD, RF, SC-J, NdC 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 926 49 132 SAT-11 PO32-01 Saturday 71 2012


72 ENDO12L_SAT-12 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Impact of [italic]MTHFR[/italic] Polymorphisms C677T and A1298C in the Synthesis of Homocysteine and Follicular Estradiol Aline Amaro Santos, Bianca Bianco, Priscila Queiroz, Juliana Christofolini, Rene Busso, Denise Maria Christofolini, Caio Parente Barbosa Faculdade de Medicina do ABC, Santo Andre, Brazil [bold]Introduction: [/bold][italic]MTHFR[/italic] gene is essential in the folate metabolism, a major factor in the methylation reactions. Hypo or hypermethylation have been associated with dietary deficiencies of folate and genetic changes as polymorphisms of [italic]MTHFR[/italic] gene. The most common polymorphisms for this gene are C677T and A1298C and can also be linked to changes in the concentrations of estrogens. It was suggested that patients undergoing assisted reproduction techniques that carry the polymorphism had significantly lower concentrations of serum estrogen and follicle producing significantly fewer oocytes retrieved since the hormonal and biochemical content has great influence on the development and quality oocyte. We aimed to correlate levels of estradiol in blood and follicular fluid of infertile women with endometriosis and unexplained Infertility and polymorphism in [italic]MTHFR [/italic]gene, and to correlate the data with indicators of embryo quality. [bold]Material[/bold] [bold]and methods[/bold]: Eleven women that had unexplained infertility and forty patients that present endometriosis were studied. These patients underwent assisted reproduction treatment. The analysis of blood and follicular fluid was performed by measuring the concentrations of homocist[iacute]ne and estradiol and by the evaluation of molecular genetic polymorphisms C677T and A1298C [italic]MTHFR[/italic] gene from peripheral blood DNA. We used a control group composed by couples with male cause of infertility for comparison of molecular genetic analysis and serum and follicular homociste[iacute]ne and estradiol concentration. [bold]Results[/bold][bold]:[/bold] The groups were defined through the molecular genetic testing and the results were related to the findings serum and follicle estradiol. The distribution of genotypes revealed no differences between groups of infertile patients and controls (p=0.930 for serum and p=0.237 for follicle) for the C677T polymorphism and (p=0.603 for serum and p=0.550 for follicle) for polymorphism A1298C. Also, no differences were found concerning the concentration of homoc[iacute]steine between the three different genotypes. Embryo quality and embryo development showed no difference. [bold]Conclusion:[/bold] In this study was not observed a significant difference in the synthesis of serum estradiol and follicle in the presence of polymorphisms for C677T and A1298C of [italic]MTHFR[/italic] gene. There is no negative impact of these polymorphisms in the parameters of oocyte quality and embryonic development suggesting that the polymorphisms studied are not correlated with lower rates of pregnancy.[br][br]Sources of Research Support: FAPESP No. 2011/16274-4.[br][br]Nothing to Disclose: AAS, BB, PQ, JC, RB, DMC, CPB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1040 49 133 SAT-12 PO32-01 Saturday 72 2012


73 ENDO12L_SAT-13 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Pregnancy Loss and Neonatal Outcomes in Women with Thyroid Dysfunction in the First Trimester of Pregnancy Kewal Aditya, Varun Gupta, Shweta Achint, Tapasya Dhar, Jubbin Jagan Jacob, Kumkum Awasthi Christian Medical College and Hospital, Ludhiana, India; Christian Medical College and Hospital, Ludhiana, India Context: Universal screening of thyroid functions using both TSH assays and FT4 estimation in pregnancy leads to at least five different common diagnostic entities. Pregnancy and foetal outcomes in overt hypothyroidism and thyrotoxicosis is well documented, while outcomes with subclinical hypothyroidism and isolated hypothyroxemia are less clear.[br]Objective: To evaluate the pregnancy and neonatal outcomes in women with first trimester maternal thyroid dysfunction.[br]Design: The study is part of a prospective study evaluating routine screening of thyroid function tests among 1000 pregnant women in the first trimester (Thyroid Screening in Urban Ludhiana in Pregnancy Study: TULIPS). Normal first trimester values for TSH were considered to be between 0.03 to 2.3 mIU/L based on data from Chinese patients using the same automated chemiluminescent assay.(1) Normal non-pregnant range for free T4 was used for classification. The patients were classified based on the above ranges and compared to the reference group (antibody negative euthyroid patients).[br]Setting: The study was conducted in the antenatal clinic of a university affiliated teaching hospital.[br]Outcomes: The outcomes evaluated included early and late pregnancy losses, preterm delivery, birth weights and five neonatal complications.[br]Results: Of the 951 patients, whose data was available, 21 were antibody positive euthyroid patients (GpA), 376 had subclinical hypothyroidism (GpB), 98 had overt hypothyroidism (GpC), 2 had overt hyperthyroidism (GpD) and 31 had isolated hypothyroxemia (GpE). The rest 423 were included as the reference group (GpR). GpA, GpD and GpE did not show any statistical difference in outcomes when compared to the GpR. Subclinical hypothyroidism (GpB) was associated with significantly increased foetal loss, preterm delivery, increased low birth weight(LBW)rates and increased small for gestation age(SGA)rates, the odds ratios (95% CI) for these being 2.87(1.67-4.95), 1.91(1.06-3.44), 1.82(1.27-2.61) and 2.11(1.40-3.18) respectively. Overt hypothyroidism (GpC) was also associated significantly increased foetal loss, increased LBW rates, increased SGA rates and increased incidence of neonatal hyperbilirubinemia, the odds ratios (95% CI) for these being 2.54(1.17-5.51), 1.83(1.06-3.15), 2.12(1.15-3.89) and 3.14(1.57-6.25) respectively.[br]Conclusions: Subclinical hypothyroidism is associated with a similar magnitude of risk as overt hypothyroidism with regards to pregnancy loss and birth weights.[br][br](1) Panesar et al., Ann Clin Biochem 2001;38:329.[br][br]Nothing to Disclose: KA, VG, SA, TD, JJJ, KA 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1080 49 134 SAT-13 PO32-01 Saturday 73 2012


74 ENDO12L_SAT-14 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Newly Diagnosed Hyperthyroidism in the Third Trimester of Pregnancy Manige Konig, Rana Malek, Eleanor Grimm Hutchens University of Maryland School of Medicine, Baltimore, MD Background: Hyperthyroidism in the third trimester of pregnancy is an uncommon phenomenon and should lead to further investigation. Choriocarcinoma is one of the possible causes of hyperthyroidism.[br]Clinical case: A 34 year old Asian female presented with increasing shortness of breath, weakness, hemoptysis to the hospital. Patient was 31 weeks gravid (G4P1A2). She had no other past medical history and had received most of her prenatal care in China.[br]Lab test results showed hyperthyroidism with a TSH of [lt] 0.02 uIU/ml (0.50-4.50 uIU/ml), free T4 of 4.8 ng/dL (0.7-1.8 ng/dL) and total T3 of 2.86 ng/ml (0.80-2.00 ng/ml).[br]She was started on methimazole 30 mg oral every 24 hours.[br]After 1 week of treatment her free T4 remained elevated with 3.4 ng/dl (0.7-1.8 ng/dl).[br]Further work up for possible autoimmune causes for hemoptysis, shortness of breath remained negative.[br]One week after admission, her respiratory status declined requiring intubation. CT-angio was negative for a pulmonary embolism but revealed multiple bilateral lung masses concerning for metastasis or granulomatous disease.[br]A b-HCG was obtained which showed a high level of 1,433,740 mIU/ml confirming the diagnosis of metastatic choriocarcinoma. The fetus was delivered emergently via C-section and survived without complications.[br]The surgical pathology report of the placenta confirmed the diagnosis of a choriocarcinoma.[br]The patient was started emergently on chemotherapy with EMA/CO (etoposide, methotrexate, actinomycin D, cyclophosphamide) and with a downwards trend in b-HCG the thyroid function normalized by day 8 of chemotherapy with a normal fT4 of 1.2 ng/dl (0.7-1.8ng/dl). At this point methimazole was discontinued.[br]Unfortunately the patient was pronounced brain dead after an acute bilateral parenchymal hemorrhage with left to right midline shift and ventricular entrapment.[br]Conclusion: This is a rare case in which a viable pregnancy co-existed with metastatic choriocarcinoma. HCG is composed of an a and b subunit. Hyperthyroidism can be induced via the homology of the a subunit of HCG to TSH; however, the lower potency of HCG to the TSH receptor requires HCG levels [gt] 100,000 mIU/mL to cause hyperthyroidism. The treatment is chemotherapy and methimazole until thyroid function tests normalize. Choriocarcinoma must be in the differential for all pregnant patients presenting with new hyperthyroidism in the third trimester.[br][br]Nothing to Disclose: MK, RM, EGH 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 203 49 135 SAT-14 PO32-01 Saturday 74 2012


75 ENDO12L_SAT-15 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Re-Evaluation of Estrogen Receptor Expression in Endometriotic Cells Masao Izawa, Fuminori Taniguchi, Tomoiki Kiyama, Naoki Terakawa, Tasuku Harada Tottori University School of Medicine, Yonago, Japan; Tottori University School of Medicine, Yonago, Japan [bold]Background:[/bold] Endometriosis is an estrogen-dependent disease. The role of estrogen is obvious because the symptoms associated with endometriosis disappear after menopause, and the administration of GnRH agonists or progestin relieves the pelvic disease and the associated pain. A marked up-regulation of aromatase gene leads to a high estrogen environment in endometriotic tissues (1, 2). The urgent issue to be clarified is to understand the molecular basis of estrogen action. So far, a higher estrogen receptor [beta] (ER[beta]) and a lower estrogen receptor [alpha] (ER[alpha]) in endometriotic tissues have been documented to explain the pathophysiology of endometriosis. However, the recent discovery of multiple ER[beta] isoforms with distinct functions prompted us to re-evaluate the molecular basis of estrogen receptor (ER) expression in endometriotic tissues.[br][bold]Objective:[/bold] We re-evaluated the molecular basis of ER expression in stromal cells from chocolate cysts of patients with endometriosis.[br][bold]Patients:[/bold] The chocolate cyst lining in ovaries of patients with endometriosis was the source of endometriotic tissue. As control, endometrial tissues were obtained from uteri of cycling premenopausal women who had uterine leiomyoma. These patients had received no hormonal treatment before surgery. We obtained the informed consent from all patients.[br][bold]Methods:[/bold] Stromal cells were purified from endometriotic and endometrial tissues. ER[alpha] and ER[beta] mRNAs were assayed by real-time RT-PCR. Specific primer sets of unique 5[apos]-untraslated exons/exon2 in [italic]ESR1[/italic] were used for ER[alpha] promoter assay. Specific primer sets of unique 5[apos]-untraslated exons/exon1 in [italic]ESR2[/italic] were used for ER[beta] promoter assay.[br][bold]Results:[/bold] The expression of ER[alpha] mRNA in endometrial cells was 7-fold higher than that in endometriotic cells. A single cDNA sequence predicting a 66KDa ER[alpha] was amplified. 5[apos]-untranslated exons, C, D and F, were expressed. The expression of ER[beta] mRNA in endometriotic cells was higher than that in endometrial cells. Transcripts for ER[beta] isoforms ([beta]1, [beta]2, [beta]4 and [beta]5) were detectable. Among them, the ER[beta]5 was the most dominant. The expression was 4-fold higher than the ER[beta]1 in endometriotic cells. The 5[apos]-untranslated exon ON was expressed.[br][bold]Conclusion:[/bold] We re-evaluated the molecular basis of ER expression in endometriotic cells. Along with a marginal level of ER[alpha] mRNA, transcripts for 4 ER[beta] isoforms were demonstrated. These findings may provide a new facet in understanding the pathophysiological role of estrogen in endometriosis.[br][br]1. Izawa M, Harada T et al., Fertil Steril 2008; 89: 1390. 2. Izawa M, Taniguchi F et al., Fertil Steril 2011; 95: 33.[br][br]Nothing to Disclose: MI, FT, TK, NT, TH 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1178 49 136 SAT-15 PO32-01 Saturday 75 2012


76 ENDO12L_SAT-16 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Estradiol Metabolites and Changes in Insulin Sensitivity during Oral Contraceptive Use Kai I Cheang, Xia Xu, Timothy D Veenstra, John E Nestler Virginia Commonwealth University, Richmond, VA; National Cancer Institute, Frederick, MD; Virginia Commonwealth University, Richmond, VA BACKGROUND: Deterioration in insulin sensitivity has been reported in some women taking combined hormonal oral contraceptives (OC). Mouse models suggest the estradiol metabolites 2-hydroxyestradiol (2OHE2) and 2-hydroxyestrone (2OHE1) regulate glucose metabolism. We prospectively evaluated changes in estradiol metabolites during OC use and their effect on insulin sensitivity.[br]METHODS: In 25 women during administration of ethinyl estradiol 35mcg and norgestimate 0.18/0.215/0.25 mg, insulin sensitivity (Si) was evaluated by frequently sampled IV glucose tolerance test, and serum endogenous estrogens and metabolites were quantified by LC-MS2 at baseline, 3 and 6 months. Pearson[apos]s correlations were used to evaluate associations between [Delta]Si and [Delta]estrogen metabolites. Subjects were also stratified into those whose Si worsened and those whose Si remained unchanged during the 6 months. Trends of 2OHE1 and 2OHE2 levels during 6 months of OC use between the 2 groups were evaluated by repeated measures analyses.[br]RESULTS: Ratios of 2OHE1/Total estrogens (-2.5[plusmn]2.0, p=0.003) and 2OHE2/Total estrogens (-0.11[plusmn]0.05, p=0.002) decreased during OC use. Although Si did not change significantly (+0.62[plusmn]0.4 min-1/mu/L, p=0.4198) overall after 6 months of OC, substantial inter-individual variability exist, and 50% of subjects had worsened Si. [Delta]2OHE2 (p=0.043) and [Delta]2OHE2/Total estrogens (p=0.045) were directly correlated with [Delta]Si. When participants were divided into whether their Si deteriorated or remained unchanged during the 6 months, divergent patterns of 2OHE2 were observed. Women whose Si deteriorated exhibited a more pronounced decrease in 2OHE2/Total than women whose Si remained changed (-0.38[plusmn] 0.29 vs. -0.08 [plusmn] 0.30 [x10-2], p=0.0166).[br]CONCLUSIONS: Changes in endogenous estradiol metabolism may determine OC[apos]s effects on insulin sensitivity.[br][br]Sources of Research Support: NIH K23HD049454; UL1RR031990; American Heart Association 11GRNT7730026 awarded to KIC.[br][br]Nothing to Disclose: KIC, XX, TDV, JEN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1303 49 137 SAT-16 PO32-01 Saturday 76 2012


77 ENDO12L_SAT-17 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Use of a Quality of Life (QOL) Menopause Rating Scale (MRS) To Define Hormone and Aging Symptoms during the Menopause and Post-Menopause (Geripause) Bernard A Eskin, Jay Tank, Owen Montgomery, Christopher Sell Drexel University College of Medicine, Philadelphia, PA; Drexel University College of Medicine, Philadelphia, PA [bold]Introduction.[/bold] The menopausal transitions are distinct in th lives of women, with certain common physiologic events thaat occur, related to the imbalance of the hypothalaamic-pituitary-ovarian axis. The symptoms experienced are myriad, and include all domains of a woman[apos]s life - psychological, physiological, physical, and sexual. While each woman is unique in terms of the specific symptoms experienced, including their severity and frequency, it is evident that they affect their lives to a great extent. They can lead to a substantial decrease of the QOL. The MRS was developed in response to the lack of standardized scales to measure the aseverity of aging symptoms and their impact on the health-related quality of QOL.[br][bold]Materials and Methods.[/bold] 60 women (20 in each group 40-49, 50-64, 65+) were randomly recruited. The patients were consented on the research study, given the MRS questionnaire (MQ) to fill out on their own, and a blood sample was taken for hormonal assays. The MQ has been standardized to include the 11 most common symptoms a woman might experience during the pre-menopausal, menopausal, and post-menopausal transition period. It consists of a list of 11 symptoms grouped into 3 domains: psychological, physiological, somato-vegetative, and urogenital. The respondents havee 5 choices for each symptom listed: no symptoms, mild, moderate, marked, and severe, with a score of 0-4, respectively. The composite scores for each of the dimensions are based on adding up the scores of these symptoms.[br][bold]Results.[/bold] The results from the MQ were stratified but stable in all age groups. The changes in symptoms are determined by many factors. In this study we looked at the results from the MQ to more clearly delineate the effects of aging versus hormone loss. They showed a significant difference between the pre-menopausal and early post-menopausal groups, particularly the psychological symptoms. Several theoies have been suggested for random changes seen in the statistics, but there is an high degree of varaibility.[br][bold]Conclusion.[/bold] The MRS questionnaire has been shown to have substantial validity and reliability, and should be considered as a potential diagnostic tool. One advantage of using the MRS is its complete reliance on the patient for the answers, leading to more accurate representation of the most concerning symptoms. This can lead to more defined therapeutic modalities that will benefit the patient.[br][br]Nothing to Disclose: BAE, JT, OM, CS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1548 49 138 SAT-17 PO32-01 Saturday 77 2012


78 ENDO12L_SAT-18 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) The Effect of a Tissue-Specific Estrogen Complex (TSEC) on Normal Mammary Gland and Breast Cancer Xenografts in Mice: Rationale for Use of a Tissue-Selective Estrogen Complex Yan Song, Wei Yue, Jiping Wang, Richard J Santen University of Virginia, Charlottesville, VA The Women[apos]s Health Initiative (WHI) studies showed that the combination of an estrogen plus a progestin increased the risk of breast cancer in postmenopausal women while estrogen alone did not. As a novel means of eliminating the need for a progestin as menopausal hormone therapy (MHT), use of a tissue selective estrogen complex (TSEC) has been proposed. The most studied TSEC in women involves the pairing of a SERM, bazedoxifene (BZA) with conjugated estrogens (CE). Our hypothesis is that this combination may prevent the growth of occult breast cancers in women. To systematically examine the effects of this TSEC on breast, we quantified its effects on mouse mammary gland and occult human breast cancer xenografts. We initially used computerized morphometry to examine the effects of estradiol (E[sub]2[/sub]) and CE alone or in combination with BZA on ductal development in immature castrate mice. BZA completely blocked CE or E[sub]2[/sub] stimulated ductal and terminal end bud growth (P[lt]0.05). We then examined human breast tumor proliferation. Occult, undiagnosed breast cancer is present at autopsy in 7% of women between the ages of 40 and 80 (Santen et al JCEM 95:S1-66, 2010). Using a MCF-7 mouse xenograft model to mimic occult tumors in women, we demonstrated that plasma levels of 80 pg/ml of E[sub]2[/sub] (estrogen clamp method) stimulated growth of small tumors. This effect was completely blocked with BZA (2 mg/kg) (p[lt]0.05). BZA also inhibited E[sub]2[/sub] stimulation of PR, pS2, cMyC and AREG; the enhancement of Ki67 and PCNA as proliferation markers; and the anti-apoptotic effect of estrogen. CE was much less potent than E[sub]2[/sub] on reducing apoptosis and on stimulation of Ki67 expression. In addition, gene expression analysis showed less stimulation of proliferation related genes (AREG, cyclin D1, and cMyc) by CE compared to E[sub]2[/sub] but all effects were blocked by BZA. Surprisingly CE alone did not stimulate tumor growth but did cause a 6-fold increase in uterine weight. This uterine weight bioassay confirmed that CE were effectively absorbed, de-conjugated in vivo, and capable of exerting an estrogenic effect on uterus but not on breast cancer. In summary, these results demonstrate that BZA can abrogate the growth of E[sub]2[/sub] stimulated occult human breast tumors. The estrogenic effects of CE on cellular markers could be blocked by BZA. These data support our hypothesis that the CE/BZA TSEC exerts anti-estrogenic effects on benign and malignant breast tissue.[br][br]Santen et al., JCEM 2010; 95:S1.[br][br]Sources of Research Support: Pfizer Inc.[br][br]Disclosures: RJS: Advisory Group Member, Pfizer, Inc., Novo Nordisk; Consultant, Teva. Nothing to Disclose: YS, WY, JW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1584 49 139 SAT-18 PO32-01 Saturday 78 2012


79 ENDO12L_SAT-19 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Development and Characterization of a Hyperglycosylated VEGF Analogue That Is Superior to Recombinant VEGF for Stimulating Angiogenesis during Ovarian Folliculogenesis in Rats Rhonda K Trousdale, Martin A Julius, Susan V Pollak, Joyce W Lustbader Columbia University, New York, NY; Columbia University, New York, NY; Harlem Hospital, New York, NY Physiological post-natal angiogenesis occurs cyclically in reproductive age women. Within the ovary, follicles rapidly expand during folliculogenesis. The independent vascular network surrounding each individual follicle must also expand rapidly to provide adequate oxygen, food, and nutrients to the growing structure. As women age, poor ovarian follicle vascular development is associated with reduced fertility.[br]A unique aspect of evaluating angiogenesis in rat ovarian follicles is the ability to accurately quantify vascular area for a well-defined structure as opposed to only evaluating vascular density. We previously reported that during non-stimulated folliculogenesis, as follicles increase in size the vasculature develops in a reproducible ratio(1). However, treatment with a long-acting follicle stimulating hormone analogue (FSH-CTP) promotes rapid follicle development with a lag in vascular development. Co-administration of FSH-CTP with recombinant vascular endothelial growth factor (rVEGF) did not enhance enhance follicle vascular development(1). We hypothesized rVEGF therapy was ineffective due to the inherently short half-life of VEGF(30 min).[br]We developed a hyper-glycosylated VEGF protein(VC). Hyper-glycosylation increased the half-life 3-fold to 80 min with decline to baseline at 200 min.[br]To evaluate the effect of VEGF monotherapy on follicle vasculature, 21 day female Sprague Dawley rats were given saline; rVEGF(1[mu]g); or VC(1[mu]g) at 0 and 24 hours. At 48 hours ovaries were evaluated. There was no difference in follicle vasculature development between these 3 groups.[br]However, when rats were stressed by administration of FSH-CTP, co-administration with VC significantly enhanced all parameters measured including mean antral follicle number, mean follicle size, and mean follicle vasculature. Furthermore, VC co-administration restored the ratio of vasculature to inner area back to the level of non-FSH-CTP treated rats.[br]This study demonstrates hyperglycosylation of VEGF-A[sub]165[/sub] does not interfere with protein activity and enhances the half-life without loss of biological activity. While administration of rVEGF during the stress of FSH-CTP ovarian hyperstimulation was unable to improve vasculogenesis, VC co-administered with FSH-CTP was able to enhance ovarian follicle vascular development leading to an increase in the number of large, healthy antral follicles. Short-term administration of VC during [italic]in vitro[/italic] fertilization may improve fertility in older women.[br][br](1)Trousdale RK et al., Endocrinology 2007; 148:1296.[br][br]Sources of Research Support: Juvenile Diabetes Foundation Innovative Grant; NIH Grant DK063224; NIH Grant HD055126 awarded to JWL.[br][br]Nothing to Disclose: RKT, MAJ, SVP, JWL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1658 49 140 SAT-19 PO32-01 Saturday 79 2012


80 ENDO12L_SAT-20 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Development of a Well-Characterized PAPP-A2 ELISA To Measure Bioactive PAPP-A2 in Maternal Biological Fluids Bhanu Kalra, Ajay Kumar, Anthony Tran, Claus Oxvig, Shivani Shah AnshLabs, Webster, TX; University of Aarhus, Gustav Wieds, Denmark Pregnancy-associated plasma protein-A2 (PAPP-A2) is a novel metalloproteinase identified as a homolog of PAPP-A in the metzincin superfamily of pappalysins. PAPP-A2 shares 46% sequence identity with PAPP-A. PAPP-A2 is a noncovalently linked dimer of two 220-kDa subunits. It exhibits roboust proteolytic activity against IGFBP-5 and possibly also IGFBP-3. PAPP-A2 is expressed in a wide range of tissues and is abundant in placental syncytiotrophoblasts and the pregnant uterus. The physiological importance of PAPP-A2 is not fully known.[br][bold]Methodology:[/bold] We have developed a well characterized two-step sandwich-type enzymatic microplate ELISA to measure PAPP-A2 levels in the maternal serum and other biological fluids. The assay measures PAPP-A2 in 50 [micro]L of sample (diluted 20 folds in sample diluent) against recombinant PAPP-A2 calibrators (0.1-10 ng/mL). The antibody pair used in the PAPP-A2 ELISA measures bioactive PAPP-A2 and does not detect PAPP-A2 missing the C-terminal region (residues 1397-1791), dimeric PAPP-A, PAPP-A proMBP complex and MMP-9.[br][bold]Validation [/bold]Total imprecision calculated on 4 samples over 12 runs, 4 replicates per run, using CLSI EP5-A guidelines was 4.7% at 1.03ng/mL, 4.% at 1.8ng/mL, 4.6% at 2.6ng/mL and 4.4% at 3.13ng/mL. The limit of detection calculated using six serum samples in the range of 0.091-3.085ng/mL over 12 runs is 0.071ng/mL.[br]The functional sensitivity of the assay at 20% CV was 0.08ng/mL. Dilution studies showed an average recovery of 100-110%. The median PAPP-A2 value on a first trimester samples (n=65) and second trimester samples (n=65) were 30.76ng/mL and 42.14ng/mL, respectively. When potential interferents (hemoglobin, triglycerides and bilirubin) were added at two times their physiological concentration, PAPP-A2 concentrations were within [plusmn]10% of the control.[br][bold]Conclusions:[/bold] A quantitative, robust and fully characterized microplate PAPP-A2 ELISA has been developed to measure PAPP-A2 in maternal serum. The approximate median PAPP-A2 levels found in a first and second trimester maternal serum can be measured with [lt] 5 % CV using this assay. The performance of the assay is acceptable for investigation of clinical utility in a variety of maternity related disorder.[br]* For Research Use Only. Not for use in diagnostic procedures.[br][br]Nothing to Disclose: BK, AK, AT, CO, SS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1690 49 141 SAT-20 PO32-01 Saturday 80 2012


81 ENDO12L_SAT-21 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Evaluation of Vascular Aging Using Myometrial Artery Calcification and Intima Thickness Sarah Catherine Hessler, Gerson Weiss, Debra S Heller, Peter G McGovern, Catherine Susan Delia, Laura T Goldsmith UMDNJ-New Jersey Medical School, Newark, NJ Although the incidence of cardiovascular disease increases with advancing age in women, studies have not successfully distinguished the effects of changing hormonal status on vascular aging from the effects of chronological age. Currently, models of human vascular aging that are generally used involve either imaging techniques or invasive sampling to assess markers of vascular aging, such as calcification and intimal hyperplasia. Occurrence of these markers has been shown to increase with advancing age and correlates with the presence of coronary heart disease. In the United States, greater than 500,000 hysterectomies are performed each year. No studies have demonstrated whether myometrial vessels can be used for the study of vascular aging. We hypothesized that myometrial arteries are a useful model for assessment of markers of vascular aging and that occurrence of these markers increases with age.[br]To test this hypothesis, myometrial arteries were assessed for calcifications utilizing hematoxylin and eosin-stained slides, and for intimal thickness utilizing orcein-stained slides for elastin. Tissues from 45-70 year old subjects who had a benign indication for hysterectomy were studied. Exclusion criteria were diagnosis of hypertension or diabetes, or treatment with sex steroids within 3 months prior to surgery. Subjects were categorized as premenopausal if they had their last menstrual period within 6 months. Calcifications were determined to be present or absent in tissues from all subjects (n=50). Intimal thickness was assessed in premenopausal women (n=16). For each subject, to assess intimal thickness, three myometrial arteries at least 0.5mm diameter were scored in a blinded manner according to the percent occlusion of the vessel lumen using the following scale: 0, less than 5%; 1, 5-20%; 2, 21-50%; or 3, greater than 50%. Data were assessed by linear regression.[br]In premenopausal women age 45-55 years, a significant positive relationship between age and intimal thickness was observed (p =0.035), intimal thickness increased with increasing age. Myometrial artery calcifications were infrequently observed, only in 2 post-menopausal subjects.[br]Myometrial arteries in hysterectomy specimens are an appropriate model for the study of vascular aging. The abundance of hysterectomy specimens may allow better differentiation of the effects of age and hormonal status on vascular aging.[br][br]Nothing to Disclose: SCH, GW, DSH, PGM, CSD, LTG 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1697 49 142 SAT-21 PO32-01 Saturday 81 2012


82 ENDO12L_SAT-22 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Androstenediol May Interact with Estradiol To Modulate Vasomotor Symptoms during the Menopausal Transition Sybil Crawford, Daniel S McConnell, Sara R Pittenger, Marcelle I Cedars, Frank Z Stanczyk, Ellen B Gold, Nancy A Gee, Bill L Lasley University of Massachusetts, Worcester, MA; University of Michigan, Ann Arbor, MI; University of California, San Francisco, CA; University of Southern California, Los Angeles, CA; University of California, Davis, CA; University of California, Davis, CA; University of California, Davis, CA Background: During the menopausal transition (MT) circulating estradiol (E2) concentrations decrease modestly in all women while circulating adrenal delta five steroids can increase up to ten fold. Androstenediol (Adiol) ranges in concentration from 100 to 1,000 pg/mL at or near the final menstrual period (FMP), while circulating E2 levels may decline to 10 pg/mL or less. E2 is primarily an estrogen receptor (ER) alpha ligand while Adiol can activate both the ER alpha and ER beta receptor-mediated pathways and possesses both estrogenic and androgenic properties. Circulating E2 concentrations are negatively related to the frequency of vasomotor symptoms (VMS) and intervention with E2 is effective in attenuating hot flushes. However, direct measurements of circulating E2 do not accurately predict the occurrence or severity of hot flushes such that inter-woman differences in the VMS experience have not been adequately explained. The wider range of circulating Adiol, compared to E2, may help explain between women differences in VMS symptoms and the lack of correlation with circulating E2.[br]Aim: To characterize the relationship of E2, Adiol and the E2/Adiol ratio in relation to the occurrence and frequency of hot flushes in women during the MT.[br]Methods: Logistic regression analysis was conducted for mid-aged women (n=142 women, 998 observations) during the MT for VMS frequency (infrequent = 1-5 days in the past 2-weeks) and frequent (6+ days in the past two weeks) versus none.[br]Results: Circulating E2 was negatively related to VMS, in the setting of both low Adiol (Odds ratio (OR) =0.37, 95% CI 0.24 [ndash] 0.59, p[lt]0.0001) and high Adiol (OR=0.63, 95% Confidence interval (CI) 0.39 [ndash] 1.03, p=0.065). Adiol was negatively related to VMS at low E2 (OR 0.72, 95% CI 0.39 [ndash] 1.33, p=0.296) but positively related to VMS at high E2 (OR 1.22, 95% CI 0.59 [ndash] 2.50, p=0.590), although the interaction was not statistically significant (p=0.137).[br]Conclusion: These data suggest that while VMS frequency has some relation to circulating E2 concentrations, the protective effect of E2 on VMS may be modulated by the antagonistic effects of the relatively higher circulating Adiol levels.[br][br]Sources of Research Support: The Study of Women[apos]s Health Across the Nation (SWAN) has grant support from the National Institutes of Health (NIH), DHHS, through the National Institute on Aging (NIA), the National Institute of Nursing Research (NINR) and the NIH Office of Research on Women[apos]s Health (ORWH) (Grants NR004061; AG012505, AG012535, AG012531, AG012539, AG012546, AG012553, AG012554, AG012495, AG017719). The content of this abstract is solely the responsibility of the authors and does not necessarily represent the official views of the NIA, NINR, ORWH or the NIH.[br][br]Nothing to Disclose: SC, DSM, SRP, MIC, FZS, EBG, NAG, BLL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1738 49 143 SAT-22 PO32-01 Saturday 82 2012


83 ENDO12L_SAT-23 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Development of a Fully Characterized Total PAPP-A Chemiluminescence Assay for Maternal Serum Evaluation* Ajay Kumar, Bhanu Kalra, Amita S Patel, Claus Oxvig AnshLabs, Webster, TX; University of Aarhus, Gustav Wieds, Denmark Maternal serum concentrations of pregnancy-associated plasma protein A (PAPP-A, EC 3.4.24.79, papalysin-1) are used to predict occurrence of Down syndrome. During pregnancy, PAPP-A is produced in high concentrations by the trophoblast and released into maternal circulation. In pregnancy, PAPP-A primarily circulates as 500-kDa heterotetrameric 2:2 complex with the proform of eosinophil major basic protein (proMBP), which inhibits the proteolytic activity of PAPP-A. Dimeric PAPP-A is the only active form and proteolyse IGFBP-4 and IGFBP-5. Significant amounts of active PAPP-A is reported at gestational ages between seven and thirteen weeks.[br][bold]Methodology:[/bold] We have developed a well characterized two-step sandwich-type enzymatic microplate glow based chemiluminiscence assay to measure PAPP-A levels in the maternal serum. The assay measures PAPP-A in 50 [micro]L of serum sample (diluted 100 folds in sample diluent) against heterotetrameric PAPP-A calibrators (25-5000 ng/mL). The antibody pair used in the assay measures dimeric PAPP-A and PAPP-A/proMBP complex in almost equal concentration and does not cross-react with proMBP, PAPP-A2 and MMP-9 at twice the physiological concentrations.[br][bold]Validation:[/bold] All analytical results are reported in ng/mL. One ng/mL of htPAPP-A = 2.56 [mu]IU/mL. Total imprecision calculated on 3 samples over 10 runs, 4 replicates per run, using CLSI EP5-A guidelines was 3.1% at 249ng/mL, 4.3% at 476ng/mL and 5.7% at 732.5ng/mL. The limit of detection calculated using six serum samples in the range of of 3.95-159.15ng/mL over 10 runs is 7.73ng/mL. The functional sensitivity of the assay at 15% CV was 11.65ng/ml. Dilution studies showed an average recovery of 96-109%. The median PAPP-A value on a first trimester samples (n=50) was 643.68ng/mL. When potential interferents (hemoglobin, triglycerides and bilirubin) were added at two times their physiological concentration, PAPP-A concentrations were within [plusmn]10% of the control.[br][bold]Conclusions:[/bold] A quantitative, robust and fully characterized microplate PAPP-A chemiluminiscence assay has been developed to measure PAPP-A in maternal serum. The approximate median PAPP-A levels found in a first trimester maternal serum can be measured with [lt] 7 % CV using this assay. The performance of the assay is acceptable for investigation of clinical utility in a variety of maternity related disorder.[br]* For Research Use Only. Not for use in diagnostic procedures.[br][br]Nothing to Disclose: AK, BK, ASP, CO 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1778 49 144 SAT-23 PO32-01 Saturday 83 2012


84 ENDO12L_SAT-24 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Pilot Study on Short-Term Effects of Aromatase Inhibitor Therapy on Cognitive Function in Postmenopausal Women with Breast Cancer Shalini Dabbadi, Paula Gendreau, Susan Tannenbaum, Susan Rabinowe, Richard Kaplan, Faryal Mirza University of Connecticut Health Center, Farmington, CT; University of Connecticut Health Center, Farmington, CT; University of Connecticut Health Center, Farmington, CT; Saint Francis Hospital and Medical Center, Hartford, CT BACKGROUND: Mild cognitive impairment (MCI) has been reported in cross-sectional studies of postmenopausal women with breast cancer receiving aromatase inhibitors (AIs). A potential underlying mechanism for this finding may be the reduction in the formation of female sex hormones from precursor androgens with AI therapy. Estrogen receptors have been identified in several areas of the brain important in cognitive performance including prefrontal cortex (short-term memory), the hippocampus (learning and storage) and the amygdala (modulation of memory consolidation).[br]OBJECTIVES: To determine the effect of short term therapy with AIs, letrozole and anastrozole, on cognitive function in postmenopausal women with breast cancer.[br]METHODS: Comprehensive neurocognitive testing was performed in postmenopausal women initiating therapy with anastrozole or letrozole for breast cancer. Subjects were evaluated at baseline and at 12 weeks after initiating therapy using the Rey Auditory Verbal Learning Test, Benton Visual Retention Test, Trail Making Test, the Stroop Color and Word Test and the Controlled Oral Word Association Test (COWAT). The Cognitive Failures Questionnaire and Beck Depression Inventory were also administered to assess perceived cognitive deficits and symptoms of depression. All tests were administered according to standardized published procedures.[br]RESULTS: 28 postmenopausal women with mean age of 59 [plusmn] 6 years completed the study. A significant decline in immediate memory (t = 2.6, df = 27, p [lt]0.02) and delayed memory (t = 2.3, df = 27, p [lt]0.03) was observed while the COWAT scores improved significantly (t = -2.8, df = 27, p [lt]0.01). There were no significant changes on the Stroop or Trail Making Tests (attention, executive functioning, non-verbal memory and processing speed for color and word). There were also no significant differences in self-reported depression and anxiety scores. Greater estrogen decline correlated with lower delayed memory scores on follow-up (r = -0.30), although it did not reach statistical significance.[br]CONCLUSION: Recall memory for a word list declined in the first three months of therapy with AIs in women with breast cancer suggesting the role of estrogen depletion in MCI. Our results suggest the need to further explore the assessment of short and long term cognitive function with adjuvant endocrine treatment of breast cancer as these findings may temporarily or permanently impact the quality of life in this population.[br][br]Sources of Research Support: Connecticut Breast Health Initiative. General Clinical Research Center, University of Connecticut Health Center.[br][br]Nothing to Disclose: SD, PG, ST, SR, RK, FM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1792 49 145 SAT-24 PO32-01 Saturday 84 2012


85 ENDO12L_SAT-25 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) SHBG, but Not Estrogen and Testosterone, Is Beneficially Associated with Adipokines in Postmenopausal Women Roksana Karim, Wendy J Mack, Chun-Ju Chien, Howard N Hodis, Frank Z Stanczyk University of Southern California, Los Angeles, CA; University of Southern California, Los Angeles, CA; University of Southern California, Los Angeles, CA; University of Southern California, Los Angeles, CA [underline]Background:[/underline] Adipose tissue is a major source of estrogens in postmenopausal women. Little is known about the association of estrogens and adipokines in this population. Till date, only one cross-sectional study evaluated the association of adiponectin with sex hormones and sex hormone binding globulin (SHBG) in postmenopausal women. The associations of leptin, resistin, and ghrelin with sex hormones and SHBG are largely unknown in this population.[br][underline]Methods:[/underline] A cross-sectional study in 643 postmenopausal women not taking any hormone therapy. Using stored frozen samples from the baseline visit, serum concentrations of estrone (E1), total estradiol (E2), and total testosterone (T) were measured by validated extraction/chromatographic RIAs; SHBG was measured by direct chemiluminescent immunoassay. Free estradiol (FE2) and free testosterone (FT) levels were calculated. Serum leptin, adiponectin, resistin, and ghrelin concentrations were quantified by RIA. Concentrations of all the analytes were tested for association with time since menopause, BMI, and waist-hip ratio (WHR), using linear regression models. Linear regression models were also used to evaluate the associations of sex hormones and SHBG (dependent variables) with adipokines and ghrelin (independent variables) adjusted for age, race, and BMI.[br][underline]Results:[/underline] The mean (SD) age of the women was 60 (6.9) years, the 68.5% white, and 58% were [gt]10 years from menopause. SHBG, adiponectin, and leptin concentrations but no sex hormones were significantly greater among women [gt]10 years (all p-values [lt]0.03). E1, E2, FE2, and FT were significantly inversely associated with adiponectin and leptin (all p[lt]0.03) adjusted for age, race, and BMI. Only estrogens were significantly inversely associated with ghrelin (all p[lt]0.01). On the contrary, SHBG levels were significantly positively associated with adiponectin and inversely associated with leptin.[br][underline]Conclusions:[/underline] Low adiponectin and ghrelin and high leptin levels are reportedly associated with obesity and increased risk of cardiovascular disease (CVD) whereas high SHBG levels are protective. In that context, our data indicate adverse association of estrogens and testosterone with adipokines and ghrelin. On the contrary, high SHBG levels are beneficially associated with adipokines. Further research is warranted to understand the underlying mechanisms of interaction between adipokines and sex hormones pertaining to CVD.[br][br]Sources of Research Support: Robert E. and May R. Wright Foundation Award, University of Southern California.[br][br]Nothing to Disclose: RK, WJM, C-JC, HNH, FZS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1882 49 146 SAT-25 PO32-01 Saturday 85 2012


86 ENDO12L_SAT-26 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Effects of Conjugated Estrogens and Estradiol on Breast Cancer Growth [italic]In Vitro[/italic] and Antagonism by Bazedoxifene: Rationale for Use of Tissue-Selective Estrogen Complexes (TSECs) Yan Song, Wei Yue, Jiping Wang, Richard J Santen University of Virginia, Charlottesville, VA The Women[apos]s Health Initiative (WHI) trial of menopausal hormone therapy (MHT) demonstrated an increased incidence of breast cancer in women receiving Premarin (conjugated estrogens, CE) plus the progestin, medroxyprogesterone acetate, but strikingly, not with Premarin alone. The use of a TSEC (tissue selective estrogen complex) has been proposed as a novel MHT strategy to eliminate the requirement for a progestin. The first TSEC in clinical development, which pairs the SERM, bazedoxifene (BZA) with conjugated estrogens improves hot flashes, vaginal atrophy, and bone density without stimulation of uterine or benign breast tissue. We posed the question whether this TSEC would exert anti-estrogenic effects on breast and potentially prevent breast cancer. Seven % of women from ages 40-80 harbor occult, undiagnosed breast cancer at autopsy (Santen et al., JCEM 95:S1-66, 2010) and a TSEC might inhibit growth of these tumors. To address this issue, we initially compared estradiol (E[sub]2[/sub]) and CE alone on proliferation and apoptosis in MCF-7 breast cancer cells. CE dose-dependently stimulated growth of MCF-7 cells at a peak concentration (10[sup]-9[/sup] M) ten-fold higher than required for E[sub]2[/sub] (10[sup]-10[/sup] M). We found that both CE and E[sub]2[/sub] alone increased DNA synthesis and reduced apoptosis at similar concentrations and BZA (10[sup]-8[/sup] M) completely blocked these effects. We then used western blot analysis to evaluate several key protein markers linked to proliferation and apoptosis. Both CE and E[sub]2[/sub] significantly stimulated phosphorylation of MAP kinase, Akt, and p70S6K as proliferation markers and up-regulated survivin, Bcl-2, and XIAP as anti-apoptotic factors. These effects of CE and E[sub]2[/sub] could be completely blocked by BZA. Gene expression studies demonstrated that CE and E[sub]2[/sub] were equally potent on expression of cMyc, pS2, and WISP whereas the stimulatory effect of CE on PR and AREG expression was weaker than for E[sub]2[/sub]. BZA effectively blocked each of these effects and alone, demonstrated no estrogen agonistic effects when used under estrogen deprived conditions. In summary, our results indicate that the stimulatory effects of E[sub]2[/sub] or CE on breast tumor cells could be completely abrogated by BZA at concentrations equivalent to those used in women. These studies imply that the CE/BZA TSEC exerts anti-estrogenic effects on breast cancer and might block the growth of occult, undiagnosed breast tumors in postmenopausal women, resulting in a reduction of diagnosed breast tumors.[br][br](1) Santen et al., JCEM 2010; 95:S1.[br][br]Sources of Research Support: Pfizer, Inc.[br][br]Disclosures: RJS: Advisory Group Member, Pfizer, Inc., Novo Nordisk; Consultant, Teva. Nothing to Disclose: YS, WY, JW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2006 49 147 SAT-26 PO32-01 Saturday 86 2012


87 ENDO12L_SAT-27 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Oxytocin Activates NF-[kappa]B and MAPK Pathways in Human Amnion Sung H Kim, Andrew Blanks, Steve Thornton, Mark R Johnson, Phillip R Bennett, Vasso Terzidou Imperial College, London, UK; Warwick Medical School, Coventry, UK Human labour is associated with an increase in prostaglandin (PG) and inflammatory cytokine synthesis within the fetal membranes. Human amnion is known to be one of the major sources of PG production. Incubation of pre-labour amnion cells with oxytocin (OT) results in a marked increase in PGE[sub]2[/sub] synthesis and upregulation of COX-2 and cPLA[sub]2[/sub], key enzymes involved in PGE[sub]2[/sub] synthesis. Our study has identified a novel role for OT in activating NF-kappaB in human amnion and has explored the regulatory mechanisms involved in OT-mediated COX-2 expression in human amnion.[br]Primary amnion cell cultures were established from patients having elective pre-labour caesarean section at term. We found that OT treatment results in nuclear translocation of p65, but not of p50 or RelB NF-kappaB subunits. OT activates p65 and IKK[alpha]/[beta] resulting in degradation of IkappaB[alpha] which could be inhibited using an IKK[beta] inhibitor (TPCA-1).[br]Treatment with OT upregulated mRNA expression of several NF-kappaB regulated genes, including IL-8, CCL2, CCL5 and COX-2 (p[lt]0.05, ANOVA). siRNA mediated knockdown of the p65 NF-kappaB subunit resulted in a significant suppression of OT induced COX-2 upregulation. Pre-treatment with IKK[beta] inhibitor (TPCA-1) also significantly decreased the upregulation of COX-2 by OT (p[lt]0.05, ANOVA).[br]We found that OT activates ERK1/2 and p38 kinase, but not JNK. Inhibition of ERK1/2 and p38 resulted in suppression of OT induced NF-kappaB activation and COX-2 upregulation. Inhibition of G[sub]i[/sub] protein by pertussis toxin (PTX) led to inhibition of OT induced phosphorylation of p65, ERK1/2 and p38, and suppressed OT induced COX-2 and p-cPLA[sub]2[/sub] upregulation.[br]We have demonstrated that OTR couples with G[sub]i[/sub] to activate NF-kappaB in human amnion by a novel mechanism, which requires both IKK and MAPK activation. Our findings suggest a potential further mechanism by which OTR-antagonists and NF-kappaB inhibitors might act in the management of preterm labour.[br][br]Sources of Research Support: Action Medical Research Grant 2010-2013.[br][br]Nothing to Disclose: SHK, AB, ST, MRJ, PRB, VT 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2052 49 148 SAT-27 PO32-01 Saturday 87 2012


88 ENDO12L_SAT-28 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Development of Ovarian Cultures to Rapidly Test Chemotherapeutic Fertotoxicity [italic]In Vitro[/italic] and [italic]In Vivo[/italic] So-Youn Kim, Marilia Cordeiro, Vanida Ann Serna, Jessina Tomas, Lizbeth Gutierrez, Takeshi Kurita, Teresa K Woodruff Northwestern University, Chicago, IL Earlier detection of cancer, coupled with new and improved treatment options, has increased the number of survivors of cancer, especially those who are diagnosed as children. As the number of young cancer survivors increases, so does the concern for their quality of life post-treatment. Unfortunately, off-target effects of radiation and chemotherapies can have unexpected and lasting consequences. For young women, this is particularly worrisome, as the immature oocytes that comprise the ovarian reserve are particularly vulnerable to the effects of radiation and drugs. Our group has developed in vitro ovarian culture and isolated ovarian follicle culture systems in order to advance our understanding of follicle development and the effects of cancer treatments on the ovary, with the goal of finding new ways to restore fertility to young cancer survivors who have been rendered sterile by their cancer treatment. A recent study suggested that the tyrosine kinase inhibitor Gleevec[reg] (imatinib mesylate) protects oocytes from cisplatin-induced cell death when immature animals were treated. Options for preserving fertility in young women prior to cancer treatment are investigational and involve removal of an ovary; even in the face of certain sterility, this can be a difficult decision for young patients and their parents. Experiments aimed at more rapidly testing chemotherapies as well as identifying neo-adjuvant fertoprotective agents can be more rapidly advanced by the deployment of the in vitro follicle culture system and xenograph model described here. To test our in vitro culture system and the xenograph transplant model, we first treated ovaries with Gleevec[reg] in vitro. Here we show that Gleevec[reg] rescues primordial follicles by cisplatin in cultured mouse ovaries by blocking an apoptotic signaling pathway including Bax, cPARP and Caspase 3. Further, we show that the xenograph model recapitulates the in vitro results and extend our analysis to monitor rH2AX foci. Importantly, Gleevec[reg] decreases the number of young follicles containing rH2AX foci inducing DNA damage. These studies show that we have two model systems in which drug testing can occur rapidly, and aid our identification of fertoprotective agents in the future.[br][br]Sources of Research Support: NIH grant(HD041857).[br][br]Nothing to Disclose: S-YK, MC, VAS, JT, LG, TK, TKW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2068 49 149 SAT-28 PO32-01 Saturday 88 2012


89 ENDO12L_SAT-29 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Effects of Bazedoxifene/Conjugated Estrogens on Coagulation Parameters in a 1-Year, Randomized, Placebo- and Active-Controlled, Phase 3 Trial of Postmenopausal Women Sebastian Mirkin, Kelly A Ryan, John R Thompson, Arkadi A Chines Pfizer Inc, Collegeville, PA [bold]Objective:[/bold] In phase 3 clinical trials, bazedoxifene (BZA)/conjugated estrogens (CE) demonstrated efficacy in treating menopausal symptoms and preventing osteoporosis in postmenopausal women with a favorable safety/tolerability profile, including no increase in the risk of coronary heart disease or stroke. The risk of venous thromboembolism for BZA/CE appears to be similar to that for CE and BZA alone. The effects of BZA/CE on coagulation parameters were evaluated in the Selective estrogens, Menopause, And Response to Therapy (SMART)-5 trial.[br][bold]Methods:[/bold] The SMART-5 trial was a 1-year, randomized, double-blind, placebo (PBO)- and active-controlled, phase 3 study in non-hysterectomized, postmenopausal women aged 40 to 65 years (N = 1,843). Coagulation parameters were assessed in a subset of randomized subjects who received BZA 20 mg/CE 0.45 mg (n = 135), BZA 20 mg/CE 0.625 mg (n = 154), BZA 20 mg (n = 73), CE 0.45 mg/medroxyprogesterone acetate (MPA) 1.5 mg (n = 70), or PBO (n = 158).[br][bold]Results:[/bold] Coagulation parameters were evaluated in 590 dosed subjects (mean age [plusmn] standard deviation [SD], 52.9 years [plusmn] 3.4 years; mean years since last menstrual period [plusmn] SD, 2.5 years [plusmn] 1.5 years). At 12 months, BZA 20 mg/CE 0.45 and 0.625 mg and BZA 20 mg showed decreases from baseline in antithrombin III while PBO showed an increase (mean percent change of [ndash]2.3%, [ndash]4.7%, [ndash]3.4%, and 3.7% for BZA 20 mg/CE 0.45 and 0.625 mg, BZA 20 mg, and PBO, respectively; [italic]P[/italic] [lt]0.0001 for BZA/CE and BZA alone vs PBO). The change in antithrombin III for CE 0.45 mg/MPA 1.5 mg (1.8%) was similar to that for PBO. There were no differences among the BZA 20-mg/CE 0.45- and 0.625-mg, BZA 20-mg, CE 0.45-mg/MPA 1.5-mg, and PBO groups in the mean percent changes in Protein C activity (19.3%, 10.8%, [ndash]5.1%, 5.2%, and 1.5%, respectively) or Protein S activity ([ndash]9.3%, [ndash]8.9%, [ndash]5.4%, 5.7%, and [ndash]4.5%, respectively). Compared with PBO, both BZA/CE doses showed decreases in fibrinogen and PAI-1 activity, and an increase in plasminogen ([italic]P[/italic] [lt]0.05 for all). Changes in D-dimer, PAI-1 antigen, prothrombin time, and partial thromboplastin time were generally similar among groups.[br][bold]Conclusion:[/bold] Overall, there was no evidence of clinically meaningful changes in coagulation parameters for BZA 20 mg/CE 0.45 and 0.625 mg compared with PBO at 12 months. Together with the efficacy of BZA/CE on menopausal symptoms and bone preservation, these findings indicate that BZA/CE may be a promising option for postmenopausal women.[br][br]Sources of Research Support: This study was supported by Pfizer Inc.[br][br]Disclosures: SM: Employee, Pfizer, Inc. KAR: Employee, Pfizer, Inc. JRT: Employee, Pfizer, Inc. AAC: Employee, Pfizer, Inc. 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2123 49 150 SAT-29 PO32-01 Saturday 89 2012


90 ENDO12L_SAT-30 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) A Pooled Analysis of the Effects of Bazedoxifene/Conjugated Estrogens on Lipid Parameters from the Selective Estrogens, Menopause, and Response to Therapy Trials Sebastian Mirkin, Kelly A Ryan, Kaijie Pan, Arkadi A Chines, Rogerio A Lobo Pfizer Inc, Collegeville, PA; Columbia University Medical Center, New York, NY [bold]Objective:[/bold] Bazedoxifene (BZA)/conjugated estrogens (CE) is the first tissue selective estrogen complex in clinical development. The Selective estrogens, Menopause, And Response to Therapy (SMART) trials showed that BZA/CE reduced menopausal symptoms and bone loss and was well tolerated in postmenopausal women with a uterus. The effects of BZA/CE on lipid parameters were assessed based on pooled data from the SMART-1, SMART-2, SMART-3, SMART-4, and SMART-5 trials.[br][bold]Methods:[/bold] The SMART trials were randomized, double-blind, placebo (PBO)-controlled, phase 3 studies in non-hysterectomized postmenopausal women. Lipid parameters were assessed at 12 and 24 months for the SMART-1 trial, 3 months for the SMART-2 and SMART-3 trials, and 12 months for the SMART-4 and SMART-5 trials; pooled data were analyzed for women treated with BZA 20 mg/CE 0.45 or 0.625 mg or PBO (N = 4,735).[br][bold]Results:[/bold] BZA 20 mg/CE 0.45 and 0.625 mg were associated with decreases from baseline in total cholesterol (mean changes [95% confidence intervals (CI)] of [ndash]0.3 [[ndash]1.7, 1.1] and [ndash]0.3 [[ndash]1.6, 1.0] mmol/L, respectively, at 12 months; [ndash]0.1 [[ndash]1.6, 1.3] mmol/L for PBO). BZA 20 mg/CE 0.45 and 0.625 mg also reduced levels of low-density lipoprotein cholesterol (LDL-C) from baseline (mean changes [95% CI] of [ndash]0.4 [[ndash]1.6, 0.9] and [ndash]0.4 [[ndash]1.5, 0.7] mmol/L, respectively, at 12 months; [ndash]0.1 [[ndash]1.3, 1.1] mmol/L for PBO). Similar results for total cholesterol and LDL-C were seen at 3 and 24 months. Changes in high-density lipoprotein cholesterol were similar among groups at all time points. The BZA/CE doses showed slight increases in triglycerides at 3, 12, and 24 months. Mean changes (95% CI) in triglycerides for BZA 20 mg/CE 0.45 and 0.625 mg and PBO at 12 months were 0.1 ([ndash]0.9, 1.2), 0.1 ([ndash]0.9, 1.2), and [ndash]0.01 ([ndash]1.0, 1.0) mmol/L, respectively. At 3, 12, and 24 months, the BZA/CE doses showed slight increases in apolipoprotein A (0.13-0.14, 0.16-0.18, and 0.14-0.15 g/L, respectively) compared with minimal changes for PBO, and slight decreases in apolipoprotein B ([ndash]0.05 to [ndash]0.07, [ndash]0.02 to [ndash]0.03, and [ndash]0.02 to [ndash]0.03 g/L, respectively) compared with an increase of 0.03 g/L for PBO at 12 and 24 months.[br][bold]Conclusion:[/bold] Based on this pooled analysis of the SMART studies, BZA 20 mg/CE 0.45 and 0.625 mg were associated with a favorable overall lipid profile in postmenopausal women over 2 years of treatment.[br][br]Sources of Research Support: This study was supported by Pfizer Inc.[br][br]Disclosures: SM: Employee, Pfizer, Inc. KAR: Employee, Pfizer, Inc. KP: Employee, Pfizer, Inc. AAC: Employee, Pfizer, Inc. Nothing to Disclose: RAL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2170 49 151 SAT-30 PO32-01 Saturday 90 2012


91 ENDO12L_SAT-31 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Low Risk of Thrombophilia with Bazedoxifene/Conjugated Estrogens: Summary of Pooled Coagulation Data from the Selective Estrogens, Menopause, and Response to Therapy Trials Sven O Skouby, Rogerio A Lobo, Kelly A Ryan, John R Thompson, Barry S Komm, Arkadi A Chines, Sebastian Mirkin Herlev Hospital, Herlev, Denmark; University of Copenhagen, Copenhagen, Denmark; University of Southern Denmark, Esbjerg, Denmark; Columbia University Medical Center, New York, NY; Pfizer Inc, Collegeville, PA [bold]Objective:[/bold] Bazedoxifene (BZA)/conjugated estrogens (CE) is a menopausal therapy in development and has shown efficacy in reducing menopausal symptoms and bone loss in the Selective estrogens, Menopause, And Response to Therapy (SMART) trials in postmenopausal women. The effects of BZA/CE on coagulation parameters were assessed based on pooled data from the SMART-1, SMART-4, and SMART-5 trials.[br][bold]Methods:[/bold] The SMART-1, SMART-4, and SMART-5 trials were randomized, double-blind, placebo (PBO)- and active-controlled, phase 3 studies in postmenopausal women with a uterus. Mean changes in coagulation parameters were assessed at 12 months in the SMART-4 and SMART-5 trials and at 12 and 24 months in the SMART-1 trial; pooled data were analyzed for women who received BZA 20 mg/CE 0.45 or 0.625 mg or PBO.[br][bold]Results:[/bold] Across the 3 trials, coagulation parameters were assessed in 1,978 randomized and dosed subjects. At 12 months, BZA 20 mg/CE 0.45 and 0.625 mg showed decreases from baseline in antithrombin III compared with an increase for PBO ([ndash]0.05, [ndash]0.06 and 0.02 L/L, respectively; [italic]P[/italic] [lt]0.01 vs baseline for all). At 24 months, BZA 20 mg/CE 0.45 and 0.625 mg and PBO showed similar decreases in antithrombin III ([ndash]0.27, [ndash]0.26, and [ndash]0.21 L/L, respectively; [italic]P[/italic] [lt]0.001 vs baseline for all). Changes in Protein C activity at 12 and 24 months were [ndash]0.01 to [ndash]0.03 and 0.05 L/L, respectively, for the BZA/CE doses compared with [ndash]0.02 and 0.04 L/L, respectively, for PBO ([italic]P[/italic] [lt]0.01 vs baseline for all except BZA 20 mg/CE 0.625 mg and PBO at 12 months). For the 2 BZA/CE doses and PBO, changes in Protein S activity were [ndash]0.02 to [ndash]0.03 and [ndash]0.02 L/L, respectively, at 12 months and 0.06 to 0.08 and 0.15 L/L, respectively, at 24 months ([italic]P[/italic] [lt]0.05 vs baseline for all except PBO at 12 months). There were no apparent changes in fibrinogen, prothrombin time, partial thromboplastin time, PAI-1 activity, plasminogen, and D-dimer for BZA/CE compared with PBO. Pooled incidences of venous thromboembolic and cerebrovascular events were low ([le]0.2%) and similar among groups.[br][bold]Conclusion:[/bold] BZA 20 mg/CE 0.45 and 0.625 mg showed an overall favorable coagulation profile in postmenopausal women, as changes in coagulation parameters were generally comparable to those for PBO over 2 years. Combined with the demonstrated efficacy and safety of BZA/CE in treating menopausal symptoms and preventing osteoporosis, these findings show that BZA/CE may be a promising alternative for non-hysterectomized women.[br][br]Sources of Research Support: This study was supported by Pfizer Inc.[br][br]Disclosures: SOS: Advisory Group Member, Wyeth Pharmaceuticals. KAR: Employee, Pfizer, Inc. JRT: Employee, Pfizer, Inc. BSK: Employee, Pfizer, Inc. AAC: Employee, Pfizer, Inc. SM: Employee, Pfizer, Inc. Nothing to Disclose: RAL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2187 49 152 SAT-31 PO32-01 Saturday 91 2012


92 ENDO12L_SAT-32 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Vasomotor Symptoms and Cardiovascular Risk Markers in Postmenopausal Women Emily D Szmuilowicz, JoAnn E Manson, Ellen W Seely Northwestern University, Chicago, IL; Brigham and Women[apos]s Hospital and Harvard Medical School, Boston, MA; Brigham and Women[apos]s Hospital, Boston, MA [underline]Background[/underline]: We previously found that women who had vasomotor symptoms (VMS) around the time of menopause had lower risk of CVD, stroke, and all-cause mortality and that the predictive value of VMS for clinical CVD events varied by the timing of VMS onset.[br][underline]Methods[/underline]: We examined associations between VMS and both circulating CVD risk biomarkers and blood pressure (BP) in the Women[apos]s Health Initiative Observational Study (WHI-OS). N ranged 649-662 for lipid measures, 2327-2360 for biomarkers, and was [gt]59000 for BP and white blood cell count (WBC) measurements. VMS status was defined as: (1) No VMS: no VMS at menopause onset or at WHI-OS enrollment; (2) Early VMS: VMS at menopause onset, but not at WHI-OS enrollment; (3) Persistent VMS: VMS at both menopause onset and WHI-OS enrollment; (4) Late VMS: VMS at WHI-OS enrollment, but not at menopause onset. The associations between VMS and outcome variables were examined using linear regression. Outcome variables were systolic BP (SBP), diastolic BP (DBP), C-reactive protein (CRP), interleukin-6 (IL-6), vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), E-selectin, white blood cell count (WBC), insulin, glucose, and lipid levels. Covariates included: age, ethnicity, education, smoking, hysterectomy/oophorectomy status, aspirin use, hormone therapy use, and physical activity (partial adjustment); and above plus BMI, hypercholesterolemia, diabetes, and family history of CHD (full adjustment).[br][underline]Results[/underline]: Compared to no VMS, persistent VMS were associated with higher SBP and DBP, and late VMS were associated with higher SBP (but not DBP) in the partially-adjusted model only. Early VMS were not associated with BP in either model. Persistent and late (but not early) VMS were associated with higher WBC in the partially-adjusted model only. In both models, persistent (but not early/late) VMS were associated with higher glucose and insulin, and early (but not persistent/late) VMS were associated with lower VCAM-1. There were no consistent associations between VMS and CRP, IL-6, ICAM-1, E-selectin, or lipids.[br][underline]Conclusions[/underline]: Persistent and late VMS were associated with one or more CVD risk markers (higher BP, WBC, and/or markers of impaired glucose metabolism), while early VMS were associated with lower levels of one marker of endothelial dysfunction. The pathways linking VMS to CVD risk may be mediated by traditional CVD risk factors and biomarkers and may be influenced by the timing of VMS.[br][br]Nothing to Disclose: EDS, JEM, EWS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2236 49 153 SAT-32 PO32-01 Saturday 92 2012


93 ENDO12L_SAT-33 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Cyclic Pattern of 17 [beta]-Estradiol Improves Spatial Memory Performance and Ischemia Outcome in Female Rats Ami Raval, Raquel Borges-Garcia, W Javier Moreno, Isabel Saul, Helen Bramlett Univeristy of Miami, Miami, FL; Univeristy of Miami, Miami, FL; University of Miami, Miami, FL The failure of the Women[apos]s Estrogen for Stroke Trial raised concern regarding the safety of chronic estrogen treatment in women. Recently, we demonstrated that an exogenous bolus of 17[beta]-estradiol to ovariectomized rats protect hippocampus against ischemia via activation of cyclic-AMP response element binding protein (CREB) pathway. In brain phosphorylation of CREB regulates learning and memory. Therefore, in the current study we hypothesize that intermittent estrogen treatment improves cognition and protects the brain from ischemic damage in female rats. Seven days after ovariectomy female rats were injected with 17[beta]-estradiol (5 [micro]g/Kg; i.p.) or oil at intervals of every 48/72h for the period of one month. During last 7 days rats (n=11) were tested using the Morris watermaze to measure cognitive capabilities. The first 4 days measured learning capabilities (learning the position of a platform), the 5[sup]th[/sup] day measured memory (memory of platform location when platform is removed), and the 6[sup]th[/sup] and 7[sup]th[/sup] days measured working memory (learning new platform location). In parallel set of experiments, on 48 hours after last hormone treatment rats were exposed to cerebral ischemia produced by 10 min of bilateral carotid occlusion and systemic hypotension (50 mmHg). Seven days later, rat brains were fixed for histopathological assessment. Results the watermaze study demonstrated that there was a difference in performance between the estradiol and vehicle groups. Data is presented as mean and standard deviation. Experimental group required less time to find the platform (21[plusmn]11s), than the control group (24[plusmn]18s), and remained in the correct quadrant longer during their allotted time (52%vs.12%). Secondly, in the working memory trials, treated rats also required less time to find the platform (21[plusmn]10s) than the control group (25[plusmn]11s). Further results are forthcoming. The results of histopathological analysis demonstrated that the number of live neurons per slice in the CA1 hippocampal region in na[iuml]ve rats was 1100[plusmn]45 (n=4). Ischemia in OvX rats decreased the number of normal neurons by 82% (192[plusmn]10, n= 6, p[lt]0.05). Intermittent estradiol-17[beta] treatment of OvX rats prior to ischemia increased the number of normal neurons 51% (556[plusmn]13, n=8, every 48h) and 41% (446[plusmn]14, n=7, every 72h) compared to the OvX group (p[lt]0.05). Intermittent 17[beta]-estradiol treatment conferred protection in cognitive tasks against ischemia in OvX rats.[br][br]Nothing to Disclose: AR, RB-G, WJM, IS, HB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2270 49 154 SAT-33 PO32-01 Saturday 93 2012


94 ENDO12L_SAT-34 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Effect of Oral Contraceptive Use on Heart Rate Variability during Laboratory-Induced Stress Lacey Wisslead, Lauren Drogos, Leah Rubin, Antonia Savarese, Erin Eatough, Kristen Mordecai, Pauline Maki University of Illinois at Chicago, Chicago, IL; University of Illinois at Chicago, Chicago, IL; University of South Florida, Tampa, FL; Baltimore VA Medical Center, Baltimore, MD Background: Endogenous estrogen plays a critical role in the regulation of autonomic function, including cardiac control as measured by heart rate variability (HRV). Changes in HRV have been observed across the rat estrous cycle. In addition, blockade or removal of endogenous hormones eliminates the corresponding change in HRV (1). Clinical studies have demonstrated that estrogen therapy restores autonomic balance in postmenopausal women, (2) and estrogen, but not progesterone correlates with HRV in healthy young women (3). HRV in young women using oral contraceptives has not been sufficiently studied. The only study to date suggested that OCs do not impact autonomic tone; however, it is unknown if HRV changes between active and inactive pill phases in oral contraceptive users. HRV is also affected by stress (4). In the present study, we examined the effects of progestin-containing OCs on HRV, specifically respiratory sinus arrhythmia (RSA), during conditions of stress and non-stress. We hypothesized that effects of oral contraceptives on RSA will become more pronounced during conditions of stress.[br]Methods: Participants included healthy women aged 18-40 (n = 30; 14 active; 16 inactive pill phase) who were taking the same estrogen-based oral contraceptive for a minimum of six months prior to study entry. Each participant contributed HRV measures obtained under two experimental conditions, one a laboratory-induced stress condition and the other a parallel control condition. HRV data were collected throughout the experimental and control sessions via a UFI Biolog monitor. HRV data were separated into 10 segments per experimental session, and each segment was edited using Cardio Edit.[br]Results: All women showed an increase in stress and anxiety following the TSST (p[apos]s[lt].001).There was a trend for RSA to be lower during the stress compared to the non-stress condition, F(1,28) =3.13, p=0.08, eta2=0.10. Unexpectedly, there were no significant differences between oral contraceptive users in the active versus inactive pill phase on RSA during the stress or non-stress condition, F(1,28) =1.66, p=0.21, eta2=0.06.[br]Conclusions: Contrary to predictions, oral contraceptive use (being in the active pill phase) did not differentially impact HRV during stress. Consistent with previous work in healthy women, our results suggest that there were no significant differences demonstrated on cardiac neural regulation.[br][br](1) T. B. Kuo, C. T. Lai, F. C. Hsu et al., Endocrinology 15 (6), 2613. (2) I. Virtanen, O. Polo, P. Polo-Kantola et al.,. Maturitas 37 (1), 45 (2000). (3) A. S. Leicht, D. A. Hirning, and G. D. Allen, Experimental physiology 88 (3), 441. (2003). (4) A. V. Klinkenberg, U. M. Nater, A. Nierop et al., Acta. obstetricia et gynecologica Scandinavica 88 (1), 77 (2009).[br][br]Nothing to Disclose: LW, LD, LR, AS, EE, KM, PM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2333 49 155 SAT-34 PO32-01 Saturday 94 2012


95 ENDO12L_SAT-35 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Association between Copy Number Variation (CNV) Polymorphism in [italic]CNTNAP4[/italic] Gene and Aging in Women: Effect Modification by Estrogen Receptor Genetic Variant Leonid Iakoubov, Malgorzata Mossakowska, Magdalena Owczarz, Monika Puzianowska-Kuznicka GeneCona, LLC, Vallejo, CA; International Institute of Molecular and Cell Biology, Warsaw, Poland; Medical Center of Postgraduate Education, Warsaw, Poland; Mossakowski Medical Research Centre, Warsaw, Poland [bold]BACKGROUND:[/bold] Accumulation of myelin-related abnormalities is characteristic for some neuropsychiatric disorders, as well as for normal aging. Several genes from neurexin superfamily encode proteins that critically support the integrity of myelin-covered fibers. Polymorphisms in a few of these genes are associated with autism, epilepsy, and schizophrenia. At least some of associations are sex-specific. We investigated whether CNV polymorphisms in neurexin superfamily genes are associated with aging in a similar sex-related fashion, and if female-specific associations could be affected by a functional polymorphism in the estrogen receptor gene [italic]ESR1[/italic].[br][bold]METHODS:[/bold] Five common intronic CNVs of in-del type that belong to 4 genes from neurexin superfamily were genotyped using TaqMan qPCR. To evaluate the association with aging, polymorphism frequency was compared between two groups of community old adults aged 65-75 and 80-90. The conclusions are based on data from two studies, a discovery study in USA Caucasians (n=90), and replication study in Eastern European Caucasians (n=1296). In each of them, samples from age groups, as well as male/female-originating groups were equally represented. Statistical significance was evaluated using Fisher[apos]s 2x2 exact test. Due to restricted power of USA sample-based study, effects of [italic]ESR1[/italic] variants on the[italic]CNTNAP4[/italic] gene association with aging were tested in Europe sample-based study only and need replication.[br][bold]RESULTS:[/bold] The only association with aging that was statistically significant in each of the two studies was for the CNV in [italic]CNTNAP4[/italic] gene in females. Combined, there were 48% carriers of risk polymorphism in females aged 65-75, versus 36% in aged 80-90, OR=1.65; CI(1.21-2.23) Fisher[apos]s P=0.00127. After additional stratification by [italic]ESR1[/italic] CNV genotype, the difference remained statistically significant in carriers of [italic]ESR1[/italic] non-deletion variant (OR =1.76; CI 1.19-2.59; P=0.00466), while virtually absent from carriers of the deletion allele (OR=0.93; CI 0.53-1.61; P=0.88829).[br][bold]CONCLUSION:[/bold] CNV in [bold][italic]CNTNAP4[/italic][/bold] gene is associated with aging in females. The association is not observed in a subpopulation of female carriers of a functional polymorphism in the [bold][italic]ESR1[/italic][/bold] gene that encodes estrogen receptor with higher sensitivity to the ligand. Female stratification based on variations in [bold][italic]CNTNAP4[/italic][/bold] and [bold][italic]ESR1[/italic][/bold] genes might reveal subpopulations with differential aging consequences in response to modifications in sex hormone status.[br][br]Nothing to Disclose: LI, MM, MO, MP-K 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 576 49 156 SAT-35 PO32-01 Saturday 95 2012


96 ENDO12L_SAT-36 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Simultaneous Measurement of Fourteen Steroid Hormones across the Menstrual Cycle in Healthy Women Using Liquid Chromatography-Tandem Mass Spectrometry Candace Keefe, Richard E Reitz, Corrine Welt, Mildred Goldman, Nigel Clarke Massachusetts General Hospital, Boston, MA; Quest Diagnostics Nichols Institute, San Juan Capistrano, CA Context: Steroid immunoassays and radioimmunoassays have limited sensitivity, specificity and accuracy. These limitations can make it difficult to assess hormone levels in children, women and hypogonadal adults. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) can provide improved accuracy and sensitivity. We have now validated a LC-MS/MS that simultaneously measures 14 steroid hormones, with results reported in reproductive age women.[br]Methods: Daily blood samples were obtained from healthy women with regular, 25-35 day menstrual cycles (n=35). Menstrual cycles were standardized to 28 days and hormone levels were centered to ovulation for comparison across the follicular and luteal phases. Pelvic ultrasounds were performed to ensure the emergence of a dominant follicle. Hormone levels were compared in the early, mid and late follicular phase (EFP, MFP, LFP) and luteal phase (ELP, MLP, LLP) and at the midcycle gonadotropin surge (MGS).[br]Results: Progesterone, 17-hydroxyprogesterone, testosterone and androstenedione levels varied across the cycle. In the follicular phase, 17OH progesterone levels increased at the MGS compared to the EFP [177 (67, 349) vs. 52 (20, 93) ng/dL; median (2.5, 97.5 percentile); p[lt]0.001] and preceded the rise in progesterone. In the MLP, 17OH progesterone [261 (139, 431) vs. (59 (23, 102) ng/dL; MLP vs. MFP; p[lt]0.001] and progesterone levels [15 (6.7, 22.2) vs. (0.1 (0.1, 0.3) ng/mL; p[lt]0.001] were higher than in the MFP, and the LP pattern of 17OH progesterone mirrored that of progesterone. Testosterone [42 (24, 66) vs. 26 (16, 47), 30 (20, 54) ng/dL; MGS vs. EFP, LLP; p[lt]0.001] and androstenedione levels [136 (73, 230) vs. 88 (58, 182), 98 (68, 175) ng/dL; p[lt]0.001] were highest at midcycle. Cortisone was lower in the LP than in the FP (p=0.02), but cortisol and DHEA levels did not change across the cycle.[br]Conclusion: This study validates previous findings that testosterone and androstenedione levels peak at the midcycle and 17-hydroxyprogesterone levels mirror the rise and fall of progesterone in the luteal phase. The assay will be a powerful tool for examining multiple adrenal and gonadal steroids simultaneously, which will be useful in the diagnosis of congenital adrenal hyperplasias.[br][br]Disclosures: RER: Chairman, Quest Diagnostics. MG: Employee, Quest Diagnostics. NC: Employee, Quest Diagnostics. Nothing to Disclose: CK, CW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2363 49 157 SAT-36 PO32-01 Saturday 96 2012


97 ENDO12L_SAT-37 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Analysis of Estrogens in Plasma Samples by LC-MS/MS, with a Reduced Sample Preparation Using SelexION Ion Mobility Technology To Remove Interferences Stefanie Krepenhofer, Bruno Casetta, Michael Jarvis, Michal Weinstock AB SCIEX, Concord, Canada (For Research Use Only. Not for Use in Diagnostic Procedures) The two estrogens, Estrone (E1) and 17[beta]-Estradiol (E2) are the primary female sex hormones. The objective of this work was to develop an accurate LC-MS/MS analytical method to detect Estrone (E1) and 17[beta]-Estradiol (E2), using simplified sample preparation to save time and cost. In general, the analysis of these compounds by LC-MS/MS is challenging due to the large number of interferences in addition to high chemical noise observed in the data. Therefore, we have evaluated the use of differential mobility spectrometry (DMS) to improve the selectivity of the detection, which has enabled us to achieve lower Limits of Detection (LOD) for the estrogens in plasma samples, after a simple sample preparation consisting only of protein precipitation.[br]Protein was precipitated from anonymized plasma samples using acetonitrile. After centrifugation the supernatant was directly injected on a 2D chromatographic separation system using a C8 clean-up column (10 [mu]m, 50 x 4.6 mm) and a Phenomenex Luna C8 separation column (5 [mu]m, 100 x 2.1 mm). The HPLC separation used a 10 minute flow gradient of methanol and water containing 0.2 mM NH[sub]4[/sub]F. The mass spectrometric detection of the estrogens was accomplished using an ABSCIEX QTRAP[reg] 5500 system operating in positive ionization mode. For improved selectivity and sensitivity, the instrument was equipped with the SelexION[trade] differential mobility device, which enabled the removal of potential interferences based on differences in their ion mobility characteristics. As a result of the additional selectivity provided by the SelexION device, a significant reduction of the background interferences was achieved. Using this approach it was possible to detect an Estrone (E1) concentration below 1 ppt and an 17[beta]-Estradiol (E2) concentration below 5 ppt in plasma, using the simple sample preparation described.[br][br]Disclosures: SK: Employee, AB Sciex. BC: Coinvestigator, AB Sciex. MJ: Employee, AB Sciex. MW: Employee, AB Sciex. 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2231 49 158 SAT-37 PO32-01 Saturday 97 2012


98 ENDO12L_SAT-38 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Estradiol Measurement Inter-Laboratory Comparison Study and Commutability Assessment Julianne Cook Botelho, Megan Vidal, Matthew Gatcombe, Hans Cooper, Hubert W Vesper Centers for Disease Control and Prevention, Atlanta, GA Accurate laboratory results are a matter of general concern in clinical medicine. This also holds true for results from Estradiol (E2) testing, especially at lower levels found in postmenopausal women. The goal of the CDC Hormone Standardization Program is to improve diagnosis, treatment, prevention and research of diseases and disorders by standardization of hormone measurements. This Inter-Laboratory Study is the first step in this process for E2 measurements.[br]An assessment of E2 measurement has been made by 17 laboratories with 23 different platforms from clinical, commercial, and research laboratories which include mass spectrometry based laboratory developed tests (LDTs) as well as immunoassay manufacturers and end users. Participants measured 40 single donor adult male and female (pre and postmenopausal) samples in duplicate over 3 different days over a range of 2.5-300 pg/mL. The serum material used for this study was produced following a standard protocol developed by the Clinical Laboratory and Standards Institute (CLSI Protocol C37-A) shown to contain minimal matrix effects and to behave similar to regular patient samples. The second part of this study was to assess the commutability of E2 standard reference materials (SRM 971) at 2 levels and of 9 commercial testing materials (CTM) used in external quality assurance (EQA) programs or as potential calibrator materials. Knowledge about the commutability of a material is important to assess its suitability as calibrators or trueness controls. The commutability assessment will evaluate whether SRMs and CTMs behave similarly as patient samples on an analytical system.[br]Current suggested criteria based on biological variability for estradiol recommend an acceptable bias of [plusmn]8.3%, imprecision of [le]11.4%, and a total error limit of [plusmn]27.2%. Bias assessments were made according to CLSI Protocol EP9-A2 and commutability by CLSI Protocol C53-A. These assessments will provide information about the measurement performance of individual assays, the measurement variability across laboratories, the bias of methods as compared to a reference method and an assessment of material suitability for calibrators or trueness controls. Furthermore, this study will help in better defining current needs and gaps in E2 measurements, which will help with standardizing the measurement more efficiently.[br][br]Nothing to Disclose: JCB, MV, MG, HC, HWV 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2237 49 159 SAT-38 PO32-01 Saturday 98 2012


99 ENDO12L_SAT-39 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Testosterone Measurement by Mass Spectrometry [mdash] A Tale of Three Internal Standards Laura Jane Owen, Brian George Keevil UHSM, Manchester, UK Introduction[br]Testosterone measurement by liquid chromatography tandem mass spectrometry (LC-MS/MS) is well accepted as the preferred technique for the analysis of serum testosterone in both males and particularly females. However variation is seen between LC-MS/MS assays and is most likely to be due to method differences between laboratories. One area of inconsistency amongst routine LC-MS/MS assays is the choice of internal standard. We investigated the effects of three internal standards on the results obtained.[br]Methods[br]Stable isotopes of testosterone were prepared in methanol. Testosterone with two deuterium (D2), five deuterium (D5) and three carbon thirteen enrichment (C13) were separately assessed.[br]Sera, calibrators and quality controls (100 [micro]L) were extracted using 0.5 mL of ether following the addition of 10 [micro]L of internal standard. All aliquots were prepared in triplicate, one for each type of internal standard. After mixing the ether was transferred to the wells of a 96-deep well block then evaporated to dryness. Extracts were reconstituted with 100 [micro]L of 50% mobile phases and analysed using a Waters Acquity LC and Quattro Premier tandem mass spectrometer. This method had previously shown to have excellent agreement with a reference method using the D2 internal standard.[br]Results[br]Lower results were obtained when using D5 testosterone when compared to D2 testosterone. The Passing-Bablock regression equation was; testosterone (D5) nmol/L = 0.86 x testosterone (D2) + 0.04.[br]The C13 internal standard also gave lower results but was closer than the D2 target than the D5 internal standard. The Passing-Bablock regression analysis was; testosterone (C13) nmol/L = 0.90 x testosterone (D2) + 0.02.[br]Discussion[br]The choice of internal standard alone can have a significant affect on the results obtained by LC-MS/MS assays for testosterone using this chromatography. The effects of the combination of chromatography and internal standard choice should be investigated during method development.[br][br]Nothing to Disclose: LJO, BGK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 553 49 160 SAT-39 PO32-01 Saturday 99 2012


100 ENDO12L_SAT-40 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Dynamics of Serum Testosterone during the Menstrual Cycle Evaluated by Daily Measurements with an ID-LC-MS/MS Method and a 2[sup]nd[/sup] Generation Automated Immunoassay Hong N Bui, Patrick M Sluss, Stuart Blincko, Marinus A Blankenstein, Annemieke C Heijboer VU University Medical Center, Amsterdam, Netherlands; Massachusetts General Hospital, Boston, MA; Abbott, Wiesbaden, Germany [bold]Context: [/bold]Testosterone levels in normally cycling women are generally assumed to be elevated around the time of ovulation. The clinical relevance of changing testosterone levels during the menstrual cycle, however, is unclear. Poor performance of current direct immunoassays for testosterone at low concentrations confounds this issue.[br][bold]Objective[/bold]: To assess daily testosterone fluctuation during the menstrual cycle by a thoroughly validated isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS) method and to evaluate whether a ARCHITECT[sup][reg][/sup] 2nd Generation Testosterone fully automated direct immunoassay is equally suited for this purpose.[br][bold]Design[/bold][bold]:[/bold] Serum samples obtained daily during the menstrual cycles of 25 healthy women, characterized by biochemical and physical examination.[br][bold]Measurements: [/bold]Testosterone, progesterone, estradiol, LH, FSH, and SHBG were measured.[br][bold]Results:[/bold] Performance of the ID-LC-MS/MS method (1) was concordant with a published reference method (2)(y = 1.007 x [ndash] 1.61 ng/dL; r = 0.9998). Comparison of the ARCHITECT[sup][reg][/sup] 2nd Generation Testosterone results to ID-LC-MS/MS yielded y = 1.095 x + 3.00 ng/dL (r = 0.9031). Overall, testosterone levels were significantly higher mid-cycle, but a peak was not discernable in each individual. Intra-individual variation exceeded the group average in the menstrual cycle; the ratio of extremes (max/min) found in individuals ranged from 1.6-3.7. Apart from a persistent positive bias, the immunoassay measured the same testosterone profiles as the ID-LC-MS/MS method. The reference interval was 8.65-48.7 ng/dL (0.30-1.69 nmol/L) for ID-LC-MS/MS and 14.4-57.6 ng/dL (0.50-2.00 nmol/L) for the 2[sup]nd[/sup] generation Architect.[br][bold]Conclusion: [/bold]Our ID-LC-MS/MS method measured low testosterone levels accurately. The ARCHITECT[sup][reg][/sup] 2nd Generation Testosterone immunoassay had acceptable performance across the entire range measured in women. On average, the elevation of mid-cycle testosterone levels is statistically significant, although not relevant for clinical practice since the day-to-day variation is higher and independent of the menstrual cycle. In this light, we recommend measuring testosterone on at least 2 independent occasions for diagnostic purposes.[br][br](1) Bui HN et al., Ann Clin Biochem 2010; 47: 248-252. (2) Thienpont LM et al., Clin Chem 2008; 54: 1290-1297.[br][br]Disclosures: SB: Employee, Abbott Laboratories. Nothing to Disclose: HNB, PMS, MAB, ACH 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 611 49 161 SAT-40 PO32-01 Saturday 100 2012


101 ENDO12L_SAT-41 POSTER SESSION: Female Reproductive Endocrinology: Pregnancy [amp] Menopause (1:30 PM-3:30 PM) Reference Ranges for Circulating Testosterone in the Follicular and Luteal Phases of the Menstrual Cycle in a Healthy, Community-Based Sample of Women in the Framingham Heart Study Andrea D Coviello, Kerrie P Nelson, Shalender Bhasin Boston University School of Medicine, Boston, MA; Boston University School of Public Health, Boston, MA [bold]Background:[/bold] Normative ranges of circulating total (TT) and free testosterone (FT) in healthy women across the menstrual phase have not been well characterized; in part due to lack of sufficiently sensitive and reliable testosterone assays that allow for accurate measurement of testosterone in the low concentrations found in women.[br][bold]Objective:[/bold] To formulate reference ranges for TT and FT in healthy women with regular cycles from a community based population using liquid chromatography tandem mass spectroscopy (LC-MSMS).[br][bold]Methods:[/bold] TT and SHBG were measured in women from Generation 3 of the Framingham Heart Study (FHS). Women, 19-45 years, were included if they reported regular menstrual cycles between 28-35 days and were not taking hormonal contraceptives. Women were excluded if they were under or overweight (BMI[lt]18 or [gt]25 kg/m[sup]2[/sup]), smoked, had impaired fasting glucose, diabetes, hypertension, hyperlipidemia, cardiovascular disease or cancer. Menstrual cycle length, first day of the last menstrual cycle, and the regularity of menstrual cycles was obtained during the FHS examination. Based on these responses and the date of blood draw, women were classified as being in the follicular (first 10 days), luteal (last 14 days), or ovulatory phase. TT was measured by LC-MSMS (sensitivity 2 ng/dL), SHBG by immunofluorometric assay, and FT was calculated. Hormones were log transformed for analyses. [bold]Results: [/bold]Mean[plusmn]SD age 37[plusmn]6 years, BMI 22.0[plusmn]1.6 kg/m[sup]2[/sup], fasting glucose 87[plusmn]5 mg/dl, total cholesterol 170[plusmn]25 mg/dl, HDLc 64[plusmn]15 mg/dl. Hormones are presented as the median (2.5[sup]th[/sup], 97.5[sup]th[/sup] percentile range). [underline]Follicular Phase (n=67):[/underline] TT 23.7 (8.3, 48.3) ng/dl, FT 1.8 (0.7, 4.4) pg/ml, and SHBG 89.7 (43.0, 184.4) nM. [underline]Ovulatory Phase (n=33):[/underline] TT 34.7 (10.1, 48.3) ng/dl, FT 2.8 (1.1, 4.6) pg/ml, and SHBG 93.3 (28.3, 154.3) nM. [underline]Luteal Phase (n=101):[/underline] TT 28.5 (11.3, 62.5) ng/dl, FT 2.4 (0.9, 7.2) pg/ml, SHBG 96.1 (42.9, 161.3) nM. Total and Free T varied across phase (TT p=0.001, FT p=0.002) but SHBG did not (p=0.7). Total and Free T were significantly lower in the follicular phase compared to the luteal phase (p[lt]0.01).[br][bold]Conclusion: [/bold]We present reference ranges for TT and FT measured by LC-MSMS in healthy premenopausal women with regular menstrual cycles. Both TT and FT were significantly different between the follicular and luteal phases suggesting that timing of measurement with the menstrual cycle is important when assessing premenopausal women[apos]s androgen status.[br][br]Sources of Research Support: RO1 HL094755.[br][br]Nothing to Disclose: ADC, KPN, SB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1684 49 162 SAT-41 PO32-01 Saturday 101 2012


102 ENDO12L_SAT-42 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Dexamethasone Decreases Expression of Genes Involved in Cholesterol Biosynthesis and Steroidogenesis in Stallion Testes Nancy H Ing, David W Forrest, Penny K Riggs, Adrianna L Lee, Shavahn Loux, Charles C Love, Dickson D Varner, Steve P Brinsko, Thomas H Welsh Texas A[amp]M University, College Station, TX; Texas A[amp]M University, College Station, TX Glucocorticoids negatively impact testicular steroidogenesis in males of every mammalian species studied but the molecular mechanisms that are responsible remain unclear. High levels of glucocorticoids may arise from stress or medical syndromes or treatments. In male horses (stallions) and men, subfertility is not uncommon. In three mature stallions, a single dose of dexamethasone (DEX, 0.1 mg/kg i.v.) decreased the peripheral blood concentrations of testosterone by 70% within 12 h, while four control stallions were unaffected. Castration at that time yielded testicular tissues for analyses of global gene expression using microarrays. Of thirteen differentially expressed genes identified, seven were confirmed on northern blots or by quantitative RT-PCR. Expression of one gene (NFkappaB inhibitor alpha or NFKBIA) increased by treatment, consistent with anti-inflammatory effects of DEX. Expression of six genes was depressed by DEX. The glucocorticoid receptor alpha (GR) gene, the protein product of which transduces DEX effects, was down-regulated by DEX as expected. The other five genes down-regulated were involved in cholesterol biosynthesis [squalene epoxidase and 24-dehydrocholesterol reductase] or steroidogenesis [luteinizing hormone receptor, glutathione S-transferase A3 (GSTA3) and P450 aromatase]. The GSTA3 protein is intriguing because it isomerizes the immediate precursors of testosterone and progesterone in non-rodent mammalian species, including man. We cloned the entire GSTA3 mRNA from stallion testes. The cDNA sequence localized the gene and regulatory sequences on horse chromosome 20. PCR primers were designed to selectively amplify GSTA3 and not other family members. The encoded GSTA3 protein contains the five amino acid residues that are critical for steroid isomerase activity. The protein will be expressed and analyzed for activity in the laboratory of Dr. Bengt Mannervik, an expert on the GSTA family of enzymes. Within the testes, Leydig cells express the GR gene most highly, as well as the genes depressed by DEX. Primary cultures of Leydig cells from stallion testes also showed dose-dependent regulation of GSTA3 and NFKBIA mRNA concentrations by DEX. Therefore, we propose a model in which DEX acts directly on Leydig cells to attenuate expression of a population of genes that results in decreased testosterone production. This mechanism may be useful therapeutically for altering steroidogenesis.[br][br]Sources of Research Support: American Quarter Horse Foundation grants to NHI and THW; Link Endowment.[br][br]Nothing to Disclose: NHI, DWF, PKR, ALL, SL, CCL, DDV, SPB, THW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1623 50 163 SAT-42 PO35-01 Saturday 102 2012


103 ENDO12L_SAT-43 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Mechanisms of WNT4 Action in Leydig Cell Function Pascal Bernard, Janelle Ryan, Dagmar Wilhelm, Vincent R Harley Prince Henry[apos]s Institute of Medical Research, Melbourne, Australia; The University of Queensland, Brisbane, Australia Male sexual differentiation requires the production of male-specific hormones from both fetal Sertoli and Leydig cells in the gonads. Fetal Leydig cells produce testosterone essential for the virilisation of the male embryo. Despite their crucial role, little is known about Leydig cell development and function. WNT4, a Wnt ligand, is important for both male and female gonadal development. In males, it regulates proper vascular development and androgen production. WNT4 regulates the expression of genes encoding enzymes in the testosterone biosynthetic pathway in mice and WNT4 heterozygous mutations in humans leads to excess production of androgens. While strong evidence suggests that WNT4 regulates testosterone synthesis at the transcriptional level, very little is known about its mechanism of action. A number of steroidogenic genes involved in the production of testosterone are transcriptionally regulated by nuclear hormone receptor Steroidogenic Factor 1 (SF1). One report indicates that WNT4 may regulates SF1-mediated transcriptional activation of steroidogenic genes, however the precise mechanism is unknown.[br]Using a specific antibody raised against Wnt4, we discovered that Wnt4 is specifically expressed in Leydig cells of the developing testis in mice. Using microarray expression analysis we have identified a new target gene of Wnt4, the prorenin receptor. The prorenin receptor is strongly expressed in Leydig cells and is absent in these cells in Wnt4 knockout mice. Since the prorenin receptor acts as a receptor for Wnt signalling, we hypothesize that Wnt4 upregulates the expression of the prorenin receptor and acts via the prorenin receptor to regulate steroidogenesis in the testis. We are investigating the role of Wnt4 and the prorenin receptor in mice using Wnt4 and prorenin receptor knockout models.[br][br]Nothing to Disclose: PB, JR, DW, VRH 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1399 50 164 SAT-43 PO35-01 Saturday 103 2012


104 ENDO12L_SAT-44 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Hormone-Induced 14-3-3Gamma Adaptor Protein Regulates Steroidogenic Acute Regulatory Protein Activity and Steroid Biosynthesis in MA-10 Leydig Cells Yasaman Aghazadeh, Malena Rone, Josiph Blonder, Timothy Veenstra, Buck Hales, Martine Germaine Culty, Vassilios Papadopoulos McGill University, Montr[eacute]al, Canada; National Cancer Institute at Frederick, Frederick, MD; Southern Illinois University, Carbondale, IL; McGill University, Montr[eacute]al, Canada; McGill University, Montr[eacute]al, Canada Cholesterol is the sole precursor of steroid hormones in the body. The import of cholesterol to the inner mitochondrial membrane, the rate-limiting step in steroid biosynthesis, relies on the formation of a protein complex that assembles at the outer mitochondrial membrane called the transduceosome(1). The transduceosome contains several mitochondrial and cytosolic components, including the steroidogenic acute regulatory protein (STAR) (1). Human chorionic gonadotropin (hCG) induces [italic]d[/italic][italic]e novo[/italic] synthesis of STAR, a process shown to parallel maximal steroid production (2). In the hCG-dependent steroidogenic MA-10 mouse Leydig cell line, the 14-3-3Gamma protein was identified in native mitochondrial complexes by mass spectrometry and immunoblotting, and its levels increased in response to hCG treatment. The 14-3-3 proteins bind and regulate the activity of many proteins, acting via target protein activation, modification, and localization (3). In MA-10 cells, cAMP induces 14-3-3Gamma expression parallel to STAR expression. Silencing of 14-3-3Gamma expression potentiates hormone-induced steroidogenesis. Binding motifs of 14-3-3Gamma were identified in components of the transduceosome, including STAR. Immunoprecipitation studies demonstrate a hormone-dependent interaction between 14-3-3Gamma and STAR that coincides with reduced 14-3-3Gamma homodimerization. The binding site of 14-3-3Gamma on STAR was identified to be S194 in the STAR-related sterol-binding lipid transfer (START) domain, the site phosphorylated in response to hCG. Taken together, these results demonstrate that 14-3-3Gamma negatively regulates steroidogenesis by binding to S194 of STAR, thus keeping STAR in an unfolded state, unable to induce maximal steroidogenesis (4). Over time 14-3-3Gamma homodimerizes and dissociates from STAR allowing this protein to induce maximal mitoxhondrial steroid formation.[br][br](1)Papadopoulos V et al., Mol Cell Endocrinol 2007; 59-64. (2)Stocco DM et al.,Endocr Rev 1996; 221-44. (3)Aitken A, Plant Mol Biol 2002; 993-1010. (4)Arakane F et al.,J Biol Chem 1997; 32656-62.[br][br]Sources of Research Support: CHR MOP Grant 102647 awarded to VP. RI-MUHC fellowship awarded to YA.[br][br]Nothing to Disclose: YA, MR, JB, TV, BH, MGC, VP 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 567 50 165 SAT-44 PO35-01 Saturday 104 2012


105 ENDO12L_SAT-45 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Developmental Changes of microRNA Profiles of Rat Leydig Cells: From Stem to Adult Ren-Shan Ge, Yufei Zhang, Kaiming Yuan, Zhijian Su, Ying Su, Xiaoheng Li, Qingquan Lian Wenzhou Medical College, Wenzhou, China; Mudanjiang Medical University, Mudanjiang, China; Jinan University, Guangzhou, China We have identified a developmental sequence that leads to the establishment of the adult Leydig cell (ALC). The first identified cell in the sequence is the progenitor Leydig cells. This fibroblast-like, luteinizing hormone receptor positive (LHR[sup]pos[/sup]) and 3[beta]-hydroxysteroid dehydrogenase positive (3[beta]HSD[sup]pos[/sup]) cell is shown to be from stem Leydig cell and then gives rise to the immature and adult Leydig cell. The hallmark of stem Leydig cell is its capability of self-renewal and of specifying into Leydig cell lineage. A fundamental unanswered question is how stem Leydig cells are specified, maintained and instructed to differentiate? The intrinsic cues including microRNAs (miRNAs) in stem Leydig cells possibly orchestrate the self-renewal and specification. In the present study, we systemically performed the miRNA arrays (653 common miRNAs) for stem, progenitor, immature and adult Leydig cells. We found that 3 miRNAs (miR-31, -221 [amp]-222) were 10 fold downregulation from stem to progenitor Leydig cells, with miR-31 downregulation by 600 fold. Using PicTar and TargetScanS, we find that Leydig cell specific transcription factor steroidogenic factor 1 3[apos] untranslated region (NR5A1-3[apos]UTR) has miR-31 binding site. miR-124 and 325 were significantly downregulated, mi-532-5p was upregulated from progenitor to immature Leydig cells. In conclusion, many miRNAs regulate the transition of stem Leydig cells in the Leydig cell lineage.[br][br]Sources of Research Support: The study is partially funded by NSFC 30871434.[br][br]Nothing to Disclose: R-SG, YZ, KY, ZS, YS, XL, QL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1417 50 166 SAT-45 PO35-01 Saturday 105 2012


106 ENDO12L_SAT-46 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Silver Nanoparticles Affect Leydig Cell Function [italic]In Vivo[/italic] Thomas Garcia, Guilherme Costa, Luiz Renato Franca, Marie-Claude Hofmann University of Illinois, Urbana, IL; Federal University of Minas Gerais, Belo Horizonte, Brazil Spermatogonial stem cells (SSCs) provide the foundation for spermatogenesis and are vital for the perpetuation of species. Previous studies from our lab examined the in vitro effects of silver nanoparticles (AgNPs) on an SSC cell line. Our data showed that AgNPs interfered with SSC proliferation in a dose and size-dependent manner and that AgNPs disrupted components of the GDNF signaling pathway, a pathway critical for self-renewal of SSCs in vivo. If AgNPs were to induce a similar effect in vivo, spermatogenesis and fertility would be severely impaired. To test this, CD1 male mice were given injections of vehicle and 1 mg/kg spherical AgNPs of 10nm diameter via the tail vein every 3 days, five times, beginning at postnatal day 45. This in vivo dose is roughly equivalent to the lowest in vitro dose with effects (10 [mu]g/ml) from our previous studies, and [sim]2,000 times greater than that found in the blood in a previous AgNP inhalation study. Although serum levels of LH and FSH were unaffected at 15, 60, and 120 days, serum and intratesticular levels of testosterone were significantly increased at 15 days. Additionally, significant changes in seminiferous epithelium morphology and Leydig cell nucleus and individual size (morphology) were observed at 15 and 60 days. Testis gene expression analysis of cholesterol biosynthesis, cholesterol transport, testosterone and estradiol biosynthesis, and GH-related genes revealed 3-[beta]-hydroxysteroid dehydrogenase mRNA significantly upregulated in treated animals at 15 days. In conclusion, we found significant histopathological alterations in Leydig cells, and significant changes in testicular testosterone levels and a critical biosynthesis gene. Thus, our results demonstrate that AgNPs have the potential to significantly impact testis health through altered steroid production.[br][br]Sources of Research Support: NIH T32 ES007326 and NIH HD054607.[br][br]Nothing to Disclose: TG, GC, LRF, M-CH 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2328 50 167 SAT-46 PO35-01 Saturday 106 2012


107 ENDO12L_SAT-47 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Oxidative Stress Induced by the Xenoestrogen Bisphenol A in Rat Leydig Cells Is Influenced by Diet Manjunatha K Nanjappa, Manuj Ahuja, Muralikrishnan Dhanasekaran, Frank F Bartol, Robert L Judd, Benson T Akingbemi Auburn University, Auburn, AL; Auburn University, Auburn, AL Bisphenol A (BPA) is a high volume production chemical used in the manufacture of polycarbonate plastics and epoxy resins. Greater than 90% of the population in the United States has measurable BPA levels in their blood. Concern exists that exposure to BPA may predispose individuals to adverse health outcomes. Here, objectives were to determine if exposure to environmental chemical BPA and/or consumption of high energy diets induces oxidative stress in Leydig cells. Long Evans male rats (n=14) were exposed to BPA by maternal gavage at 0, 2.5 and 25 [micro]g/kg body weight from gestational day 12 to postnatal day (PND) 21 and were fed a BPA-free diet from weaning to PND 70. Subsequently, animals from each treatment group were maintained for a period of 28 days (PND 71 to 98) on a normal or high fat diet (NFD or HFD, Harlan-Teklad, Indianapolis, IN) with 16% or 42% of energy content from fat. Animals were sacrificed within 24 h of day 98 postpartum when serum and Leydig cells were obtained to assess oxidative stress and antioxidant enzyme activity. Serum levels of thiobarbituric acid reactive substance, a marker for oxidative stress, were greater in male rats exposed to BPA and maintained on HFD than in NFD control animals (P[lt]0.05). However, serum glutathione (GSH) levels increased only in animals exposed to the 25 [micro]g/kg dose of BPA and fed HFD compared to control (P[lt]0.05). Further, exposure to BPA at 25 [micro]g/kg and maintenance on HFD induced oxidative stress in Leydig cells as indicated by increased generation of reactive oxygen species (ROS) compared to the NFD control group (P[lt]0.05). Interestingly, antioxidant enzymes were differentially affected by the interaction of BPA and HFD. For example, superoxide dismutase activity was decreased in Leydig cells from all HFD animals (P[lt]0.05), whereas catalase activity was up-regulated by exposure to BPA regardless of diet (P[lt]0.05). The results were replicated in vitro because incubation of Leydig cells with BPA (10 nM, 18 h) increased ROS production compared to control (P[lt]0.05). In addition, culture of BPA-free Leydig cells with hydrogen peroxide (0, 25, 50 [micro]M; 18 h), a prototype ROS generator, decreased testosterone (T) production (P[lt]0.05). Collectively, results indicate that BPA exposure and/or maintenance on HFD induces oxidative stress. Therefore, BPA disruption of Leydig cell T secretion may be due, in part, to increased ROS levels.[br][br]Sources of Research Support: This study was supported in part by NIH ES-015886.[br][br]Nothing to Disclose: MKN, MA, MD, FFB, RLJ, BTA 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1974 50 168 SAT-47 PO35-01 Saturday 107 2012


108 ENDO12L_SAT-48 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Activin Signaling Regulates Sertoli Cell Differentiation and Function Peter K Nicholls, Peter G Stanton, Justin Chen, Paul Gregorevic, Craig A Harrison Prince Henry[apos]s Institute of Medical Research, Clayton, Australia; Baker IDI Heart and Diabetes Institute, Melbourne, Australia Throughout development and in early postnatal life, proliferating Sertoli cells of the testis direct the formation of the seminiferous epithelium. At puberty, Sertoli cells enter a [apos]terminally[apos] differentiated state, characterised by the establishment of a functional blood testis barrier (BTB), the ability to sustain spermatogenesis, and withdrawal from mitosis. Activin A, a member of the transforming growth factor-[beta] (TGF-[beta]) superfamily, is a key regulator of Sertoli cell proliferation during embryogenesis, and a decrease in activin signalling corresponds with terminal differentiation. In this study, we sought to understand the function of activin upon differentiated Sertoli cells using an in vivo model of elevated systemic activin A. Chronic activin signalling reduces testis mass and induces a cumulative testicular dysgenesis phenotype in adult mice, consistent with a failure of Sertoli cells to support spermatogenesis. Activin was found to potently disrupt BTB function in adult mice, and tight junction formation in differentiated 20 dpp rat Sertoli cells. Furthermore, manipulation of activin signalling re-initiates a program of cellular proliferation in rat Sertoli cells. Proliferative cells were characterised by reduced expression of mature markers (including clusterin [0.32-fold, p[lt]0.01] and claudin-11 [0.23-fold, p[lt]0.01]), and re-expression of juvenile markers (including cytokeratin-18 [18-fold, p[lt]0.01] and ID3 [3.5-fold, p[lt]0.01]) following activin exposure. Our results confirm that Sertoli cells maintain the capacity to proliferate beyond puberty and show that activin promotes a functional de-differentiation of rodent Sertoli cells. This study indicates that activin signalling must be strictly controlled in the adult in order to maintain Sertoli cell function in spermatogenesis.[br][br]Sources of Research Support: National Health and Medical Research Council of Australia Project Grants (#1006488 and #1009144).[br][br]Nothing to Disclose: PKN, PGS, JC, PG, CAH 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1454 50 169 SAT-48 PO35-01 Saturday 108 2012


109 ENDO12L_SAT-49 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Estrogen Receptors and Sertoli Cell Function Thais FG Lucas, Catarina S Porto, Maria de Fatima M Lazari Escola Paulista de Medicina - UNIFESP, S[atilde]o Paulo, Brazil Aim: Sertoli cells play a key role in the control of germ cell development, and are important targets for androgen and estrogen actions. In rat Sertoli cells, 17beta-estradiol (E2) activates translocation of estrogen receptors (ESRs) to the plasma membrane, and activates ERK1/2 phosphorylation (Biol Reprod 78:101, 2008). Expression of Cyclin D1 is increased by E2 and the ESR1-selective agonist PPT, but not by the ESR2-selective agonist DPN, indicating the role of ESR1 in Sertoli cell proliferation (Biol Reprod, 2012). In mammals, progression through the cell cycle is regulated by the association of Cyclins and Cyclin-dependent kinases (CDK), and the activity of these complexes is inhibited by p27Kip1 (reviewed in Genes Dev 18:2699, 2004). The role of E2 in the regulation of p27Kip1 has not been explored. Moreover, gene expression involved in differentiation, hormone synthesis and junction proteins in Sertoli cells may be regulated by GATA-1 transcription factor (reviewed in Mol Endocrinol 22:781, 2008). GATA-1 expression in Sertoli cells is not regulated by androgen (Endocrinology 146:2674, 2005), but the role of E2 has not been explored. We now report the regulatory role of E2, PPT and DPN on the expression of p27kip1 and GATA-1 in rat Sertoli cells. Methods and Results: Primary culture of Sertoli cells was obtained from 15-day old Wistar rats. Cells were incubated in the absence (control) and presence of E2 (0.1 nM), PPT (10 nM), or DPN (10 nM) for 24 hours at 35[deg]C. Cells were untreated or pretreated with the MEK inhibitor U0126 (20 [mu]M), the PI3K inhibitor wortmannin (1 [mu]M), the disruptor of the CREB:CBP complex KG-501 (25 [micro]M), or NFkB1 nucleus translocation inhibitor SN50 (1 [micro]M). Afterwards, cells were stimulated with E2, PPT or DPN, and the expression of p27kip1 and GATA-1 was determined by Western Blot. E2 or DPN increased the expression of p27kip1 and GATA-1 (2.3 and 1.5-fold, respectively), whereas PPT had no effect. Wortmannin or KG-501 blocked the effects of E2 or DPN on the expression of p27kip1 and GATA-1, while U0126 and SN50 had no effect. Conclusion: 17beta-estradiol modulates Sertoli cell proliferation through ESR1-mediated increase of cyclin D1 and ESR2-AKT-CREB-mediated increase of p27kip1. Possible effects of E2-ESR2 on gene transcription may be mediated by increased expression of GATA-1. The present study reinforces the important role of estrogen for normal testis development and homeostasis.[br][br]Sources of Research Support: FAPESP and CNPq.[br][br]Nothing to Disclose: TFGL, CSP, MdFML 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 927 50 170 SAT-49 PO35-01 Saturday 109 2012


110 ENDO12L_SAT-50 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Activation of PPAR[alpha] and PPAR[beta]/[delta] Regulates Sertoli Cell Metabolism Mariana Regueira, Maria Fernanda Riera, Silvana Lucia Artagaveytia, Maria Noel Galardo, Eliana Herminia Pellizzari, Selva Beatriz Cigorraga, Silvina Beatriz Meroni Hospital de Ni[ntilde]os R Guti[eacute]rrez, Buenos Aires, Argentina Sertoli cells (SC) provide the structural and nutritional support for germ cell development. SC actively metabolizes glucose and converts it to lactate, which is the major energy source for germ cells. On the other hand, it has been postulated that SC uses fatty acids (FA) as a source of energy. The transcription factors PPAR[alpha] and PPAR[beta]/[delta] [mdash]members of the PPARs nuclear receptor family[mdash] participate in the expression of genes involved in FA metabolism in many cells types. The aim of this work was to study the participation of PPAR[alpha] and PPAR[beta]/[delta] activation in the regulation of genes involved in FA metabolism and in lactate production in SC. Cultures of SC obtained from 20-day-old rats were incubated for different periods of time without (control) or with variable doses of WY14643 (WY) or GW0742 (GW) [mdash]pharmacological activators of PPAR[alpha] and PPAR[beta]/[delta] respectively. Carnitine-palmitoyltransferase 1 (CPT1) mRNA levels were detected by Northern Blot and long- and medium-chain 3-hydroxyacyl-CoA dehydrogenases (LCAD, MCAD) and the fatty acid transporter CD36 (FAT/CD36) mRNA levels were analyzed by RQPCR. Results expressed as fold-increase in mRNA levels combining three independent experiments (mean[plusmn]SD), obtained in 48h incubations with 10[micro]M WY and 5[micro]M GW are shown; [bold]CPT1[/bold] WY: 2.3[plusmn]0.6*; GW: 3.3[plusmn]0.2*; [bold]LCAD[/bold] WY: 1.9[plusmn]0.4*; GW: 2.8[plusmn]0.3*; [bold]MCAD[/bold] WY: 1.9[plusmn]0.3*; GW: 1.8[plusmn]0.4* and [bold]FAT/CD36[/bold] WY: 2.5[plusmn]0.6*, GW: 2.7[plusmn]0.5* (*p[lt]0.05 vs. control). While PPAR[alpha] activation had no effect on lactate production, PPAR[beta]/[delta] activation increased it (Control: 7.9[plusmn]0.5; GW: 11.4[plusmn]0.6[sup]# [/sup][mu]g/[mu]gDNA,[sup] #[/sup]p[lt]0.001 vs. control). Phosphorylation of the Pyruvate Dehydrogenase Complex (PDC) [mdash]enzymatic complex that regulates pyruvate to acetylCoA flow[mdash] was also evaluated by Western Blot. While an increase in phospho-PDC by PPAR[beta]/[delta] activation was observed, PPAR[alpha] activation had no effect. It is suggested that the increased phosphorylation of PDC by PPAR[beta]/[delta] activation, which results in inhibition of enzymatic activity, may be responsible for an increase in pyruvate availability that is converted to lactate. Altogether these results suggest that in SC, PPAR[alpha] and PPAR[beta]/[delta] participate in the regulation of FA metabolism. PPAR[beta]/[delta] also regulates lactate production and phosphorylation of PDC, which suggests a participation of this transcription factor in the provision of lactate for germ cell development.[br][br]Sources of Research Support: PIP2008N[ordm]806; PICT2007N[ordm]1004.[br][br]Nothing to Disclose: MR, MFR, SLA, MNG, EHP, SBC, SBM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 556 50 171 SAT-50 PO35-01 Saturday 110 2012


111 ENDO12L_SAT-51 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Evidence of Defective Release of Testosterone from XXY Mouse Testis Iqbal Munir, Jeffrey Kim, Lue YanHe, Leung Andrew, Christina Wang, Swerdloff Ronald Harbor-UCLA Medical Center, UCLA School of Medicine, Torrance, CA [bold]Objective: [/bold]In an experimental XXY mouse model of Klinefelter Syndrome, we have previously demonstrated that low peripheral serum testosterone (T) and elevated gonadotropins are associated with hyperplasia and hypertrophy of Leydig cells. Isolated Leydig cells from XXY testis produce significantly higher amounts of T [italic]in vitro[/italic] than XY controls. The aim of the study was to further characterize T production in XXY testis and to find direct evidence of a defect in the release of T to the circulation in XXY mice.[br][bold]Methods: [/bold]XXY mice were generated in our lab as published before. Testes of 7 adult XXY mice and their 7 XY littermates were used in the experiments. Testicular venous system was ligated and blood collected from the venous effluent. Testes were dissected out, decapsulated and incubated in the serum free medium in the presence or absence of 10 ng/ml of lutenizing hormone (LH) for 3 h. T was measured in the incubation medium, testicular homogenate and testicular venous effluent using LC-MS/MS. Expression of androgen binding protein (ABP) was evaluated in freshly isolated testicular homogenate by Western blot.[br][bold]Results: [/bold]Incubation of decapsulated whole testis in culture medium showed higher testicular content of T in XXY mice (25 [plusmn] 4.3 ng/testis, mean [plusmn]SEM,) in comparison to XY (17[plusmn] 2.6 ng/testis). Addition of LH to the culture medium further increased T content in XXY animals (40 [plusmn] 8.3 ng/testis) and XY testis (35 [plusmn] 7.3 ng/testis). There was no difference in the amount of T released in the medium from XY vs. XXY testis. T concentration in the testicular venous effluent, was 3-fold lower in the venous effluent from XXY testis (6.4 [plusmn] 0.85 ng/ml) compared to XY testis (18.3 [plusmn] 3.4 ng/ml, p=[lt]0.05). Western blot analysis demonstrated two subunits of ABP approximately 48 and 38 kDa. The 48 kDa subunit expression was higher in XXY testis whereas 38 kDa subunit had higher expression in XY testis.[br][bold]Conclusions: [/bold]Our data provide further evidence that synthesis of T is not impaired in the XXY testis. The amount of T is not lower in the testis of XXY mice but there is a defect in the release of T from testis to the venous circulation as shown by the lower concentration of T in testicular venous effluent. The similar concentrations of T in the culture media of decapsulated XY and XXY testes reflect leakage from the interstitial space. Differences in the ABP subunits in XY vs XXY testis may play a role in sequestration of T in mouse model XXY testes.[br][br]Nothing to Disclose: IM, JK, LY, LA, CW, SR 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1543 50 172 SAT-51 PO35-01 Saturday 111 2012


112 ENDO12L_SAT-52 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Expression and Role of FoxO3a in the Testis Young-Suk Choi, Eun Jig Lee Yonsei University College of Medicine, Institute of Endocrine Research, Seoul, Republic of Korea; Yonsei University College of Medicine, Seoul, Republic of Korea; Northwestern University Feinberg School of Medicine, Chicago, IL Infertility was positively correlated with age in female FoxO3a KO mice. Degeneration in early proliferation of granulose cell was found. This study focuses mainly on the roles of FoxO3a in testis. The expression of FoxO3a starts from the embryonic day 13.5 and this lasts until the post pubertal week 3 (PPW3) in testicle. After PPW3, most of the expression of FoxO3a occurs in the nucleus of Leydig cell, but after PPW5, FoxO3a occurs both in the nucleus and cytoplasm. When GnRH signaling is defected in 5 weeks old male C57/BL6 mouse, FoxO3a was increased in the nucleus. When LH is treated on R2C cell, there was increase in phosphorylation of FoxO3a by AKT that is indicating LH regulates FoxO3a localization.[br]When active form of FoxO3a-TM adenovirus was introduced to R2C cell, a Leydig tumor cell, the concentration of testosterone hormone and StAR protein was reduced. Also when FoxO3a and StAR promoter-Luc vector were co-transfected into FT293 cell for reporter assay, FoxO3a inhibited StAR promoter activity. In conclusion, FoxO3a is a modulator of LH signal and plays an important role in testosterone production in the testis.[br][br](1)Murayama C., et al., Mol Cell Endocrinol 2012; 350(1): p. 1-9. (2)IHong Z.Y., et al,Cancer Lett 2012; 314(1): p. 34-40. (3)Fukuda S., et al.,J Ovarian Res, 2009. 2(1): p. 17.[br][br]Nothing to Disclose: Y-SC, EJL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 785 50 173 SAT-52 PO35-01 Saturday 112 2012


113 ENDO12L_SAT-53 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Gonadotroph Responsiveness to PACAP Signaling through the Short and HOP1 Isoforms of the PAC1 Receptor Rong Q Yang, Kimberly E Leake, Stephen J Winters, Joseph P Moore University of Louisville School of Medicine, Louisville, KY; University of Louisville School of Medicine, Louisville, KY There is increasing evidence that PACAP plays a role in the development and regulation of gonadotroph function. PACAP signals through VPAC1 and VPAC2, two G-protein-coupled receptors that are also activated by VIP, and through the PACAP specific receptor, PAC[sub]1[/sub]R. There are multiple splice variants of PAC[sub]1[/sub]-R (1). The Short and the Hop1 isoforms are expressed in gonadotrophs and act through the G[alpha]s pathway, while the Hop1 isoform also stimulates the G[alpha]q pathway to increase intracellular calcium (2). We sought to determine which PAC[sub]1[/sub]-R subtype is most influential in PACAP activation of the PACAP promoter and the follistatin promoter, a key PACAP target that mediates its effect to reduce FSH[beta] and GnRH-R expression. L[beta]T2 gonadotroph cells were transfected with 50 ng of either Short or HOP1 receptor expression vector and with a PACAP or follistatin promoter - luciferase construct. Transfected cells were treated for 6 h with increasing doses of PACAP, and luciferase activity was quantified. Over-expression of either Short or Hop1 increased follistatin promoter activation by PACAP equally. Mutation of the AP1 or CRE sites in the follistatin promoter reduced PACAP activation by the Short receptor only. Overexpression of either receptor increased PACAP promoter activation but the Hop1 receptor was more effective. Mutation of the proximal CRE of the PACAP promoter abolished stimulation. To identify genes activated by PACAP, L[beta]T2 cells were transfected with control or HOP1 vector and cultured with or without 10 nM PACAP for 6 h. In both groups, PACAP stimulated genes associated with PKA and p53 pathways and TRK receptor signaling equally and decreased DAR2 signaling molecules. Signaling pathway molecules stimulated more robustly in HOP1 transfected cells included PI3K/AKT, ERK/AKT, NGS and BMP pathways. Signaling pathway molecules stimulated only in HOP1 transfected cells included IP3 metabolism, calcium signaling, PKCb, JAK/STAT and NO pathways and receptor signaling molecules associated with GnRH, PPARa/RxRa, VEGF, TGFb, EGF, IGF-1, AR, ER, GR, and Wnt/b-catenin. Some of the genes significantly altered by PACAP exposure and relevant to gonadotroph function include increases in INHA, cMyc, junD, SMAD4, LIMHX1, LEP and ACVR2B and decreases in INHBB, CREBBP, ACVR1, and EGR1. Signaling through the HOP1 receptor likely involves cross-talk with multiple signaling pathways and may modulate gonadotroph responses to multiple endocrine and paracrine factors.[br][br](1) Vaudry D et al., Pharmacol. Rev. 2009; 61:283-357. (2) Mustafa T et al., J. Biol. Chem. 2007; 282:8079.[br][br]Sources of Research Support: NIH R01-HD050571 awarded to JPM. NIH Summer Endocrine Research Training Program Grant T35 DK072923.[br][br]Nothing to Disclose: RQY, KEL, SJW, JPM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2173 50 174 SAT-53 PO35-01 Saturday 113 2012


114 ENDO12L_SAT-54 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Comparative Expression of the Androgen Receptor in Immature and Mature Testes from Monkey and Human Espernaza Berensztein, Tony Plant, Roberto Ponzio, Diego Chirico, Marco A Rivarola, Alicia Belgorosky Hospital de Pediatria Garrahan, Buenos Aires, Argentina; Pittsburgh University, Pittsburgh, PA; Facultad de Medicina, Universidad de Buenos Aires, Buenos Aires, Argentina The androgen receptor (AR) is highly expressed in Sertoli cells (SC) of the adult human testis (HT) but not during infancy when expression of this receptor is found primarily in peritubular (PC) and interstitial cells (IC) (Berensztein et al. Pediatr Res 2006). In human infancy, serum gonadotropins and testosterone levels reach pubertal values but spermatogenesis is not initiated suggesting that FSH stimulation at this stage of development is not able to induce AR expression in SC. To analyze the role of gonadotropins in the regulation of the expression of testicular AR we have analyzed 1) the ontogenesis of AR expression by immunohistochemistry both in HT and in a higher primate model, the Macaca mulatta testis (MT),and 2) the effect of [italic]in vivo[/italic] LH and FSH stimulation on AR expression in SC in juvenile MT. RESULTS. 1) In both HT and MT, a similar cellular distribution of AR was found in all age groups: AR was positive in Leydig cells (LC), IC, PC and SC, and negative in germ cells (GC). In normal HT, the % of SC expressing AR (%AR) in Neonates ([lt] 1 month (m) old, n=6) was 2.25%, and in Infants (1-7 m, n=10) 2.96%, values significantly lower than in Prepubertal (5.9%,1-11 years, n=8) (p[lt]0.05) and in Pubertal (92.3.1%,14-15 y, n=4) subjects; in the adult subjects, it was 100%. Moreover, a significant positive linear regression of % AR as a function of age was found (r=0.951, p[lt]0.001). In MT, % AR in Neonates (9-18 d old, n=2) was 0.5; in Infants (4-5 m, n=2) 0, values significantly lower than in Juvenile (51%, 13-36 m, n=5) and Pubertal animals (86.1%, 42-48m, n=2); in adults (60 m, n=4) %AR was 96.7%. 2) Gonadotropin stimulation for 11 days of juvenile MT induced AR expression in SC. Specifically, %AR during treatment with FSH (79.3 [plusmn]13.6), LH (69.2 [plusmn]13) and FSH+LH (85.5[plusmn] 10.7) was significantly greater, p[lt]0.05 (Bonferroni test) with respect to that during vehicle (35.5[plusmn]17.6 %). Taken together, these results suggest 1) that upregulation of AR expression in SC during the transition from the neonatal to juvenile phase of development in primates is relatively gonadotropin independent, and 2) that LH and FSH appear to be equally important in driving the gonadotropin dependent upregulation of AR expression at puberty. Finally, validating the nonhuman primate model of Macaca mulatta for the human situation opens a range of possibilities to enrich our current knowledge of the postnatal development of HT.[br][br]Sources of Research Support: NIH grant,U54 HD 08160 (supported the generation of the monkey tissue).[br][br]Nothing to Disclose: EB, TP, RP, DC, MAR, AB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 185 50 175 SAT-54 PO35-01 Saturday 114 2012


115 ENDO12L_SAT-55 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Involvement of AMPK in Testicular Function Pauline Tartarin, Edith Guibert, Aminata Toure, Marc Foretz, Joelle Dupont, Benoit Viollet, Pascal Froment INRA, Nouzilly , France; Inserm, U1016, Institut Cochin; Cnrs UMR8104; Univ Paris Descartes, Sorbonne Paris Cit[eacute], Paris, France The 5 [apos]AMP-activated protein kinase or AMPK is a key protein kinase in the regulation of cell metabolism. It is also an important indicator of energy reserves. AMPK is the target of a drug, metformin, frequently used to treat diabetes and also certain female infertility associated with insulin resistance (syndrome polycystic ovaries). However few studies investigated the role of metformin and its target, AMPK, on male fertility. Thus, we analysed the involvement of this kinase by using 1/a male mouse line inactivated for the catalytic alpha 1 subunit of AMPK, 2/wild type males newborn mice from females treated with metformin during the first half of gestation.[br]Inactivation of AMPK led to a decrease in fertility associated with hyperandrogenism of testicular origin. Strangely, no modification of the testis structure was observed: neither in testis size or cell composition nor in apoptosis or in proliferation. However, sperm alpha1 AMPK-/- showed a lack of mobility in association with head structural anomalies, compared with male controls. Mitochondria in alpha1 AMPK-/- spermatozoa present also alterations that could be a cause of the lower mobility. In addition, hyperandrogenia was linked to hyperactivity in Leydig cells. Gonadotropins secretion were not modified in alpha1 AMPK-/- mice.[br]On the other hand, administration of metformin (an activator of AMPK) to pregnant mice showed a reduction in the testicular volume, diameter of the seminiferous tubules and the number of Sertoli cells. During administration of metformin to mothers, the intratesticular testosterone concentration in male fetuses decreased by about 30% compared to male fetuses from control mothers.[br]These two approaches suggest a role of AMPK in the testicular control (during testis development, sperm and testoterone production) similarly to its action in ovarian functions (oocyte quality and steroid production).[br][br]Sources of Research Support: This work was supported by the Agence National pour la Recherche (Fertinergy project).[br][br]Nothing to Disclose: PT, EG, AT, MF, JD, BV, PF 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1909 50 176 SAT-55 PO35-01 Saturday 115 2012


116 ENDO12L_SAT-56 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Distinct Expression Patterns Predict Differential Roles of miRNA-Binding Proteins, Lin28 and Lin28b, in the Mouse Testis Francisco Gaytan, Susana Sangiao-Alvarellos, Maria Manfredi-Lozano, David Garcia-Galiano, Antonio Romero-Ruiz, Silvia Leon, Francisco Ruiz-Pino, Fernando Cordido, Leonor Pinilla, Manuel Tena-Sempere University of Cordoba, Cordoba, Spain; University of A Coru[ntilde]a, A Coru[ntilde]a, Spain Lin28 (also termed Lin28a) and Lin28b are related RNA-binding proteins, involved in the control of microRNA maturation, especially of the let-7 family, with putative roles in the regulation of stem cell differentiation and early (embryo) development. However, their function in adult tissues and during postnatal maturation remains ill defined. Despite the general assumption that Lin28 and Lin28b share similar targets and overlap in terms of function, conclusive demonstration of such a redundancy in different tissues and cell types is still missing. Recently, expression of Lin28 has been reported in the rodent testis, and human genomic studies have disclosed the association between variations in or around the LIN28B locus and the age of menarche; observations that suggest putative roles of Lin28 proteins in mammalian reproduction, whose mechanisms are yet to be defined. We document herein the pattern of RNA expression and protein distribution of Lin28 and Lin28b in the mouse testis during postnatal development and in adulthood. Expression of Lin28 and Lin28b mRNAs was detectable in mouse testis across postnatal maturation, from neonatal to adult periods. Of note, Lin28 peptides displayed totally different patterns of cellular distribution: Lin28 protein was prominently located in undifferentiated and type-A1 spermatogonia within the seminiferous tubules, whereas Lin28b was confined to interstitial Leydig cells and spermatids. These differential patterns of distribution were conserved at earlier stages of postnatal maturation. Moreover, relative mRNA levels of Lin28 and Lin28b disparately varied between neonatal and pubertal periods, with peak levels of Lin28 in the infantile testis and sustained elevation of Lin28b mRNA in young adult male gonads; an age when relative levels of let-7a and let-7b miRNAs were significantly suppressed. Altogether, our data are the first to document the divergent patterns of cellular distribution and mRNA expression of Lin28 and Lin28b in the mouse testis along postnatal maturation; a phenomenon which is suggestive of distinct functional roles of these two related, but apparently not overlapping, miRNA-binding proteins in the male gonad.[br][br]Sources of Research Support: BFU2011-25021 (Ministerio de Economia y Competitividad, Spain).[br][br]Nothing to Disclose: FG, SS-A, MM-L, DG-G, AR-R, SL, FR-P, FC, LP, MT-S 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1427 50 177 SAT-56 PO35-01 Saturday 116 2012


117 ENDO12L_SAT-57 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) A Specific Region in the 5[prime] Flanking Region of Gonadotropin-Regulated Testicular RNA Helicase (GRTH/DDX25) Gene Directs Its Cell-Specific Expression in Testicular Germ Cells Raghuveer Kavarthapu, Chon-Hwa Tsai-Morris, Masato Fukushima, Joaquin Villar, James Pickel, Maria L Dufau National Institutes of Health, Bethesda, MD; National Institutes of Health, Bethesda, MD GRTH is a multifunctional protein post-transcriptional regulator of genes that is essential for round spermatid elongation and completion of spermatogenesis. This helicase also prevents the Leydig cells (LC) from overstimulation of gonadotropin-induced androgen pathway by promoting degradation of StAR protein. In transgenic mice (Tg) carrying deletions of the GRTH 5[apos] flanking regions upstream of the promoter domain with GFP as the reporter gene, we previously demonstrated that the 205 bp promoter of the gene contains the necessary elements for LC specific expression. No expression in germ cells (GC) was found when Tg mice carrying the 5[apos] sequence extended to 3.6 kb, and 1.4 kb 5' to the ATG codon contains a functional half-site androgen response element. To further define regulatory regions of GRTH required for expression in GC during development, Tg mice were generated carrying 5[apos] flanking sequence 6.4 kb (6.4 kbTg) and/or various deletions. Within the 3.6-6.4 kb region, similar clusters of other known sequences that direct GC specific expression were identified. Immunohistochemistry showed that 6.4 kbTg directed GFP expression in both GC and LC. Deletion of the sequence between 1.4 kb to 3.6 kb (6.4-3.6 kb/1.4 kb Tg) direct GFP expression in GC only. In these lines, GFP expression was found to be stage specific in spermatocytes, round spermatids and elongating spermatids. Maximal expression of GFP in spermatocytes was observed in the pachtyene spermatocytes entering metaphase of meiosis. No expression was found in Sertoli cells. Western blots confirmed the GPF expression in all of these Tg lines. Our studies have generated and characterized transgenic lines that can be utilized to define basal and hormonally regulated transcription of the GRTH gene in a cell specific manner, and to further advance our knowledge on the regulation of GRTH action in male reproduction.[br][br]Sources of Research Support: The IRP, NICHD.[br][br]Nothing to Disclose: RK, C-HT-M, MF, JV, JP, MLD 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1937 50 178 SAT-57 PO35-01 Saturday 117 2012


118 ENDO12L_SAT-58 POSTER SESSION: Regulation of Testicular Function (1:30 PM-3:30 PM) Expression and Gene Regulation of ARR19-[beta] in Mouse Testis Hana Kang, Keesook Lee Chonnam National University, Gwangju, Republic of Korea ARR19s, also known as Cmtm2 and cklfsf2, are MARVEL transmembrane domain containing proteins and characterized as two isoforms, ARR19-[alpha] and ARR19-[beta], with [sim]50% identity of amino acids in mice. ARR19-[alpha], which is highly expressed in the testis, was recently shown to be a factor regulating the function of testicular Leydig cells. In this study, we investigated the expression of ARR19-[beta] and its gene regulation in the testis. Besides highly expressed in the testis, ARR19-[beta] was also weakly expressed in other adult tissues such as epididymis, lung and kidney in mice. Further studies revealed that ARR19-[beta] expression was developmentally regulated in the testis. Immunohistochemical analysis of mouse testis showed that ARR19-[beta] protein was detected only in interstitial cells in young testis, whereas it was highly expressed in both haploid germ cells and interstitial Leydig cells in adult testis. ARR19-[beta] protein was also detected in epididymal sperms. In order to study the gene regulation of ARR19-[beta] in the testis, we analyzed 2 kb upstream sequence of the ARR19-[beta] promoter, which revealed the presence of three putative GATA-1 binding motifs. GATA-1 is the key regulator of ARR19-[alpha] expression in mouse Leydig cells. As well as GATA-1, CREB also markedly increased the activity of the ARR19-[beta] promoter. These findings demonstrate that ARR19-[beta] is the target gene of both GATA-1 and CREB. Interestingly, GATA-1 is repressed by cAMP, whereas CREB is activated by cAMP, in testicular Leydig cells. Therefore, ARR19-[beta] expression may be developmentally regulated by GATA-1 in young mice without LH/cAMP signaling and by CREB in adult mice with LH/cAMP signaling. Together, these findings suggest that ARR19-[beta] may be involved in the development and function of germ cells and Leydig cells in the testis.[br][br]Nothing to Disclose: HK, KL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 411 50 179 SAT-58 PO35-01 Saturday 118 2012


119 ENDO12L_SAT-59 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Estrogen Enhances Recovery of Fertility in Irradiated Rat Testis Wei Zhou, Connie C Weng, Marvin L Meistrich University of Texas MD Anderson Cancer Center, Houston, TX We previously showed in rats, in which irradiation had completely blocked spermatogonial differentiation, that testosterone (T) suppression with GnRH-antagonist and antiandrogen stimulated spermatogenic recovery and addition of estradiol (E2) accelerated this recovery. We now report for the first time, that addition of E2 enhances recovery of fertility in irradiated (6Gy) LBNF1 rats. 10 wk after irradiation, LBNF1 rats were treated with GnRH-antagonist and antiandrogen (Group XAF), or GnRH-antagonist and antiandrogen with addition of E2 (Group XAFE) for 6 wks. The rats were then allowed to recover for 6 wks without any hormonal suppression/treatment. For fertility recovery analysis, rats were mated with Sprague-Dawley female rats for 4 wks. Although condensed spermatids began to appear in testis by 22 wks after irradiation, in cauda epididymis the sperm were only observed at 26 wks after irradiation. At both time points, the sperm head counts were higher in XAFE group than in XAF group. By 26 wks after irradiation, 60% of the rats in XAFE group (n=5) recovered their fertility and for each fertile male, 1.7 litters of rats were born. The average litter size is 10.4, comparable to that of control normal rats (litter size=11.5). None of the rats in XAF group produced any pups. Testis histology analysis showed that by 26 wks after irradiation, although about 50% of the seminiferous tubules in rats of XAF group still exhibited spermatogenesis differentiation, around 1/3 of these differentiated tubules were abnormal, with missing layer(s) of differentiated germ cells or abnormal structure or apoptosis; while in XAFE groups, over 70% of the tubules exhibited spermatogenesis differentiation; in the 3 rats that had recovered fertility, over 90% of the tubules showed spermatogenesis recovery, and less than 10% of these tubules were abnormal. In XAF group, only 20% of the seminiferous tubules contained condensed spermatids by 26 wks after irradiation; while in XAFE group, about 60% of the tubules contained condensed spermatids. These data suggested that E2 not only enhances fertility recovery in irradiated rat, but also prolongs the normal spermatogenesis cycles after spermatogenesis induction by testosterone suppression. Further study and elucidation of the mechanism will help understand the role of estrogen in male spermatogenesis and may provide novel therapy methods for patients of azoospermia induced by cancer therapies.[br][br]Nothing to Disclose: WZ, CCW, MLM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 590 51 180 SAT-59 PO35-02 Saturday 119 2012


120 ENDO12L_SAT-60 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Contrasting FSH Signaling Actions during Sertoli Cell Proliferation and Differentiation Periods: Potential Regulation Via Differential Expression of G Protein Subunits Coupled to the FSH Receptor Ana P Jacobus, William H Walker Magee Womens Research Institute-School of Medicine-University of Pittsburgh, Pittsburgh, PA Sertoli cells receive, integrate, and transmit signals required for spermatogenesis. In the immature testes, Sertoli cells proliferate. During puberty, Sertoli cells stop proliferating and then differentiate so that they are able to support germ cell development. Sertoli cells proliferation and maturation are regulated by several hormones and growth factors; however, only follicle-stimulating hormone (FSH) stimulates both the proliferation of Sertoli cells immediately after birth and the differentiation of the cells during puberty. The extent of Sertoli cell proliferation and the timing of differentiation determines the final number of Sertoli cells and therefore, the number of germ cells that can be supported. To better understand the processes that control Sertoli cell development, we are investigating the mechanisms by which FSH signaling can be switched from promoting proliferation to differentiation. We hypothesize that the expression or activities of G protein subunits associated with the FSH receptor are altered during the proliferative and differentiation phases of the Sertoli cell development. Our preliminary data indicates that in proliferative Sertoli cells cultured from 5 day-old rats, FSH activates PI3K, AKT, c-Raf and Erk Kinase (MEK). These data suggest that FSH may act via G[beta][gamma] associated with G[alpha]i. In differentiated Sertoli cells isolated from 20 day-old rats, cAMP levels are elevated through FSH activation of G[alpha]s, leading to an increase of PKA activity and consequently c-Raf/MEK and AKT activities are decreased. These data suggest that FSH preferentially acts via G[alpha]s in differentiated Sertoli cells from 20 day-old rats. We will assay the levels and activities of the G protein subunits coupled to FSH receptor in 5 and 20-day-old Sertoli cells. We expect to find that the levels of G[alpha]i, G[alpha]s and their associated G[beta][gamma] subunits are altered during proliferation and differentiation such that FSH signaling results in opposing regulation of MAPK and AKT pathways during the two stages of development.[br][br]Sources of Research Support: CAPES post doctoral fellowship n0 0634/11-5.[br][br]Nothing to Disclose: APJ, WHW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1787 51 181 SAT-60 PO35-02 Saturday 120 2012


121 ENDO12L_SAT-61 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Simultaneous Insulin and Leptin Signaling in POMC Neurons Promotes Fertility and Metabolic Homeostasis in Male Rodents Latrice D Faulkner, Abigail R Dowling, Jennifer W Hill The University of Toledo, Toledo, OH; The University of Toledo, Toledo, OH Hypothalamic pro-opiomelanocortin (POMC) neurons in the arcuate nucleus are critical regulators of energy balance and glucose homeostasis. The POMC precursor is enzymatically processed to produce both the anorectic peptide [alpha]-melanocyte-stimulating hormone ([alpha]-MSH) and the endogenous opioid [beta]-endorphin. [beta]-endorphin exerts a continuous restrictive influence on GnRH secretion and the reproductive axis in both primates and rodents. Indeed, [beta]-endorphin immunoreactive terminals synapse on the GnRH neuron soma in these same species. Therefore, POMC neurons may function as critical nodes where metabolic and reproductive signals communicate.[br]Leptin and insulin convey information regarding energy stores to the CNS. POMC neurons contain both insulin and leptin receptors that regulate their activity. These regulatory signals also mediate the production of the pro-hormone convertases responsible for the post-translational processing of the POMC precursor. Here we demonstrate that male rodents lacking both insulin and leptin receptors in POMC neurons (LepR/IRPOMC mice) exhibit reduced fertility. Elevated testosterone levels, testis weights, and LH levels reflect this defect. We also examine whether the arcuate nucleus concentration of peptides produced from POMC processing and levels of pro-hormone convertases are affected. Our results suggest that the absence of leptin and insulin signaling in POMC neurons increases basal LH release and disrupts reproductive function via reduction in the inhibitory opioid tone on GnRH neurons. Thus, perception of leptin and insulin by POMC neurons is not only required for normal body weight and glucose homeostasis, but also to permit normal fertility. Additional studies are required to determine whether this action is mediated via direct or indirect neuronal connections.[br][br]Sources of Research Support: NIH Grant R00-HD-056491.[br][br]Nothing to Disclose: LDF, ARD, JWH 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2007 51 182 SAT-61 PO35-02 Saturday 121 2012


122 ENDO12L_SAT-62 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Osteocalcin Mediates Insulin Regulation of Male Reproductive Functions Franck Oury, Mathieu Ferron, Huizhen Wang, Rajendra Kumar, Gerard Karsenty Columbia University, New York, NY; University of Kansas Medical Center, Kansas City, KS Genetics and biochemical evidence have established that bone, via the osteoblast-derived hormone osteocalcin, favors testosterone biosynthesis in the male mouse following its binding to GPRC6A on Leydig cells of the testis. The unraveling of this unexpected way of regulating male fertility raises the question as to know whether osteocalcin acts downstream of LH or instead defines a second pathway necessary for male fertility. Osteocalcin injection failed to correct any of the fertility defects in the pituitary Luteinizing hormone (LH)-deficient mice while in contrast Human chorionic gonadotropin (hCG) injection could correct the fertility defect in Osteocalcin-/- mice. Together these data indicate that osteocalcin does not act downstream of LH to regulate male fertility. Taking advantage of what is already known about osteocalcin regulation, we then showed through the analysis of cell-specific gene deletion and cell ablation mouse models that: (i) Insulin signaling in osteoblasts favors male fertility by modulating osteocalcin activity, (ii) bone resorption is the cellular mechanism used by insulin signaling in osteoblasts to enhance osteocalcin bioactivity. This study provides evidence that there is a direct link between energy metabolism and male fertility, and reveals that osteocalcin is necessary for this crosstalk to occur.[br][br]Nothing to Disclose: FO, MF, HW, RK, GK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2342 51 183 SAT-62 PO35-02 Saturday 122 2012


123 ENDO12L_SAT-63 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Cyclooxygenase 2 (COX2)/15d-PGJ2 System in Hamster Sertoli Cells: Its Modulation by FSH/Testosterone and Its Involvement on Glucose Uptake Maria Eugenia Matzkin, Eliana Herminia Pellizzari, Ricardo Saul Calandra, Selva Beatriz Cigorraga, Monica Beatriz Frungieri Institute of Biology and Experimental Medicine (IBYME-CONICET), Buenos Aires, Argentina; University of Buenos Aires, Buenos Aires, Argentina; Center for Endocrinological Investigations (CEDIE-CONICET), Buenos Aires, Argentina We have previously described a stimulatory effect of testosterone (T) on cyclooxygenase 2 (COX2) expression, prostaglandin (PG) synthesis as well as the involvement of PGs in the modulation of T production in hamster Leydig cells (1, 2). In the present study, we investigated the existence of a COX2/PGs system in hamster Sertoli cells (SC), its regulation by hormones (FSH and T) and the effect of one PG, 15d-PGJ2, on SC glucose uptake.[br]The Syrian hamster is a seasonal breeder. The exposure of adult hamsters to a short photoperiod (6h light/day) for 16 weeks triggers a drastic decrease in plasma levels of LH, FSH and T, as well as a severe testicular regression. SC were purified from testes of regressed adult hamsters and cultured under basal conditions for 48h. Subsequent incubation of SC in the presence of FSH (100 ng/ml), T (1 [mu]M) or the plasma membrane-impermeable T-BSA (1 [mu]M) for 1h significantly induced COX2 expression and MAPK1/2 phosphorylation (determined by Western blot) and 15d-PGJ2 production (quantified by immunoassay). The stimulatory effect of FSH and T-BSA on COX2 expression was abolished by the selective MEK1/2 inhibitor, U0126. In addition, T-BSA-induced COX2 expression and MAPK1/2 phosphorylation were reverted by the antiandrogen bicalutamide.[br]The presence of PPAR[gamma], the nuclear receptor for PGJ2 derivatives, was detected in hamster SC by RT-PCR and immunocytochemistry. 15d-PGJ2 (1 [mu]M) decreased the uptake of [sup]3[/sup]H-2-deoxyglucose (2DOG), a non-metabolizable glucose analogue, into SC. This effect was abolished by the PPAR[gamma] antagonist, BADGE. On the other hand, FSH, T and T-BSA increased 2DOG uptake, an effect further stimulated in the presence of meloxicam, a selective COX2 inhibitor. Thus, FSH and T regulate glucose uptake in hamster SC through, at least, two mechanisms, a direct positive modulation and an indirect negative effect exerted via 15d-PGJ2/PPAR[gamma].[br]In summary, these results demonstrate the presence of a COX2/PGs system in hamster SC and its up-regulation by FSH and T. Furthermore, results obtained in the presence of the plasma membrane-impermeable T-BSA suggest that T exerts a stimulatory effect on COX2/PGs through a non-classical mechanism that involves the presence of androgen receptors and MAPK1/2 activation. Overall, the COX2/15d-PGJ2 system might act as a local modulator of FSH and T actions on hamster SC function.[br][br](1) Matzkin ME et al., Reproduction 2009; 138:163. (2) Frungieri MB et al., Endocrinology 2006; 147:4476.[br][br]Sources of Research Support: Consejo Nacional de Investigaciones Cinetificas y Tecnicas (Conicet); Agencia Nacional de Promocion Cientifica y Tecnologica (ANPCyT); Facultad de Medicina, Universidad de Buenos Aires (UBACYT); Fundacion Roemmers.[br][br]Nothing to Disclose: MEM, EHP, RSC, SBC, MBF 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 42 51 184 SAT-63 PO35-02 Saturday 123 2012


124 ENDO12L_SAT-64 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Impact of Aging and Obesity on Hepatic Sex Hormone-Binding Globulin Gene Expression in Women and Men Jyothi Gogineni, Marjan Karegar, Dushan T Ghooray, Charles R Scoggins, Stephen J Winters University of Louisville, Louisville, KY; University of Louisville, Louisville, KY Sex hormone binding globulin (SHBG) is a homodimeric glycoprotein produced by hepatocytes that transports testosterone and other steroids in human plasma, reduces their clearance, and regulates their entry into target cells. Multiple factors influence the plasma level of SHBG including genotype, sex, age, weight, diet, exercise, and a variety of hormones including thyroxine, growth hormone, cortisol, testosterone and estradiol. SHBG levels are low in obesity, and low levels beginning in childhood predict the development of the Metabolic Syndrome (MetS) and Type 2 diabetes (T2DM). However, the mechanisms that regulate SHBG are only partly understood. Experiments in HepG2 hepatocarcinoma cells and mice expressing an SHBG transgene reveal that the transcription factor HNF4[alpha] activates the SHBG promoter, COUP-TF antagonizes this effect, the peroxisome proliferator-activated receptor PPAR[gamma] represses transcription, and TNF[alpha] reduces SHBG expression partly through HNF4[alpha]. So far, however, no studies have examined SHBG gene expression in human liver. In this study, we obtained normal liver tissue from 25 patients (11F, 14M) age 46-82 yrs with colorectal cancer and ECOG performance status 0 who were undergoing partial hepatic resection as treatment for their disease. SHBG and HNF4[alpha] mRNA levels were measured by PCR, serum SHBG levels by ELISA, and clinical data were obtained. The BMI ranged from 16-34 in women and 16-37 kg/m[sup]2[/sup] in men. In agreement with previous results, serum SHBG levels were higher in women than men and diverged with increasing age, were markedly increased in women treated with estradiol or tamoxifen, and decreased with increasing BMI. SHBG mRNA levels were positively correlated to the plasma level of SHBG (r=0.37), were higher in subjects with lower BMI (r=-0.3), but were similar in men and women, and were unaffected by age. The expression level of HNF4[alpha] was highly positively correlated (r=0.81; p[lt]0.01) with the level of SHBG mRNA, and was also higher in thin patients than in those who were obese, but neither sex nor age predicted HNF4[alpha]. Moreover, while plasma SHBG was lower in those patients with MetS (n=9) or T2DM (n=8), SHBG mRNA and HNF4[alpha] levels were similar. Our results imply that the plasma level of SHBG is regulated by transcriptional and post-translational mechanisms, HNF4[alpha] is an important determinant of SHBG expression in human liver, and multiple factors contribute to the change in SHBG that occurs with aging or obesity.[br][br]Nothing to Disclose: JG, MK, DTG, CRS, SJW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 843 51 185 SAT-64 PO35-02 Saturday 124 2012


125 ENDO12L_SAT-65 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) GLUT3 and CASPR5 [mdash] Novel Genetic Factors in Male Infertility Alexander W Pastuszak, Carolina J Jorgez, Larry I Lipshultz, Dolores J Lamb Baylor College of Medicine, Houston, TX [bold]Introduction and Objective:[/bold] While several known genetic alterations cause male infertility, it is likely that many other currently unrecognized genetic etiologies exist. Here, we present genomic data implicating the glucose transporter GLUT3, as well as contactin-associated protein CASPR5, in male fertility.[br][bold]Methods:[/bold] Blood-derived genomic DNA from 22 men with non-obstructive azoospermia (NOA) and normal Y-chromosome microdeletion and karyotype analyses, as well as 4 fertile male controls, was used for array comparative genomic hybridization (aCGH) to assess for copy number variations (CNVs). Copy number gains and losses were identified and candidate fertility genes selected based on 1) the frequency of CNVs in a gene, 2) the magnitude of the gain/loss, and 3) available data on gene expression. CNVs were validated using Taqman qPCR, and DNA sequencing performed for genes of interest. Immunohistochemical staining of testis sections utilized commercially available antibodies and standard protocols.[br][bold]Results:[/bold] Copy number gains were identified using aCGH in GLUT3 in 2/22 NOA patients, and in CASPR5 in 1/22 patients and in no controls. These CNVs were confirmed in all patients using Taqman qPCR copy number assays (CNAs). Subsequent CNAs of DNA from 43 infertile men yielded 5 more men with GLUT3 gains, and one additional man with a loss in CASPR5. The frequency of GLUT3 CNVs in the general population (including fertile and infertile individuals) is approximately 5%, whereas our observed gain frequency is 16%. CNVs in CASPR5 are found in 0.002% of the general population and in 4% of our infertile male population. Sequencing of GLUT3 yielded SNPs in exons 2 (2465A[gt]C, 36K[gt]T), 6 (6505A[gt]G, 258L[gt]L), and 10 (14680C[gt]T, 436T[gt]T), none of which were predicted to be damaging. Staining for GLUT3 in testis of a male with 3 copies of GLUT3 and hypospermatogenesis demonstrated cytoplasmic Leydig cell and spermatocyte staining.[br][bold]Conclusions:[/bold] Copy number gains and losses in GLUT3 and CASPR5, respectively, were identified in several infertile males. Future work in the form of CASPR5 sequencing and subcellular localization and functional assays analyzing the impact of expression of both genes on fertility-related cell signaling pathways, as well as the study of animal models, will further elucidate the roles of these genes in male fertility.[br][br]Sources of Research Support: AUA Foundation Russell Scott, Jr., MD, Resident Research Award (AWP); NIH grants K12 DK0083014 (CJJ, DJL), the Multidisciplinary K12 Urologic Research (KURe) Career Development Program (CJJ, DJL), and by P01HD36289 from the Eunice Kennedy Shriver National Institute for Child Health and Human Development, NIH (DJL).[br][br]Nothing to Disclose: AWP, CJJ, LIL, DJL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1411 51 186 SAT-65 PO35-02 Saturday 125 2012


126 ENDO12L_SAT-66 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Circulating Endothelial Cells as Marker of Vascular Damage in Male Hypogonadism Domenico Milardi, Giuseppe Grande, Antonella Giampietro, Linda Tartaglione, Sara Palumbo, Francesca Vendittelli, Riccardo Marana, Alfredo Pontecorvi, Cecilia Zuppi, Ettore Capoluongo, Laura De Marinis Universit[agrave] Cattolica del S Cuore, Rome, Italy; Universit[agrave] Cattolica del S Cuore, Rome, Italy; Universit[agrave] Cattolica del S Cuore, Rome, Italy Testosterone (T) deficiency has become a frequently diagnosed condition in our current society with epidemic obesity and is associated with cardiovascular risk (1). Recent studies has established the importance of altered vascular endothelium function to cardivascular disease. The damage to the endothelium might also cause endothelial cell detachment, resulting in increased numbers of circultating endothelial cells (CECs) within the bloodstream (2).[br]To investigate if hypogonadism could modify CEC count in pherypheral bloodstream, we investigated CEC count by the CellSearch System in peripheral blood of twenty hypogonadic patients (25 to 55 years), studied after neurosurgical operation to remove nonsecreting pituitary adenoma or craniopharyngioma. The control group comprised ten age- and gender-matched healthy subjects. Antropomethric parameters, body mass index and waist circunference (WC) were evalutated. Serum total cholesterol, HDL cholesterol, triglycerides, glycosylated haemoglobin levels, T, estradiol, dihydrotestosterone, SHBG, LH and FSH were determined. CEC count was investigated by the CellSearch System, a semiauthomatic method to accurately and reliably enumerate CECs, sorted based on a CD146+, CD105+, DAPI+, CD45- phenotype (3).[br]CEC count/ml was increased in hypogonadic patients vs control group (mean[plusmn] SE 16.53[plusmn]7.55 vs 1.03[plusmn]0.14 cells/ml; p[lt]0.05). Correlation analysis in hypogonadic patients showed inverse correlation between T levels and CEC count/ml, positive correlation between waist circumference and CEC count/ml and inverse correlation between T levels and waist circumference. Regression analysis yielded a significant result for the model, with the highest adjusted R[sup]2[/sup] for T and WC, the most consistently significant variables.[br]Hypogonadism has been shown to be an indipendent determinant of endothelial disfunction, thus contributing to vascular pathology (4). The increase in CEC count in the group of hypogonadic patients represent a direct index of endothelial damage and could be a possible link between T deficiency and cardiovascular disease. WC in hypogonadal man may represent an adjuntive risk factor for endothelial damage and cardiovascular disease. Male hypogonadism induces endothelial disfunction. Visceral obesity may be implicated in the endothelial-disfunction induced by T deficiency.[br][br](1) Ullah MI et al., Horm Metab Res 2011; 43(3):153. (2) Blann AD et al., Thromb Haemost 2005; 93(2):228. (3) Strijbos MH et al., Thromb Haemost 2009; 102(2):347. (4) Foresta C et al., Clin Endocrinol 2008; 68(2):284.[br][br]Nothing to Disclose: DM, GG, AG, LT, SP, FV, RM, AP, CZ, EC, LDM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1924 51 187 SAT-66 PO35-02 Saturday 126 2012


127 ENDO12L_SAT-67 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Regenerated Luminal Epithelial Cells Are Derived from Pre-Existing Luminal Epithelial Cells in Adult Mouse Ventral Prostate Laura E Pascal, June Liu, Sudhir Isharwal, Daniel Metzger, Raquel Ramos Garcia, Jan Pilch, Susan Kasper, Karin Williams, Per Basse, Joel Nelson, Pierre Chambon, Zhou Wang University of Pittsburgh School of Medicine, Pittsburgh, PA; Universit[eacute] de Strasbourg, Coll[egrave]ge de France, Strasbourg, France; University of Cincinnati, Cincinnati, OH; University of Pittsburgh School of Medicine, Pittsburgh, PA Introduction: Determining the source of regenerated luminal epithelial cells in the adult prostate during androgen deprivation and replacement will provide insights into the origin of prostate cancer cells and their fate during androgen deprivation therapy. Prostate stem cells in the epithelial layer have been suggested to give rise to luminal epithelium. However, the extent of stem cell participation to prostate regrowth is not clear.[br]Methods: A recently developed genetic lineage tracing strategy was used to follow the fate of mouse prostatic luminal epithelial cells during androgen withdrawal and androgen replacement. To control for the specificity of prostatic luminal epithelial cell labeling and the time of labeling, a previously reported PSA[minus]CreERT2 transgenic mouse strain expressing tamoxifen (TAM)[minus]inducible CreERT2 recombinase driven by a 6[minus]kb human PSA promoter/enhancer was used. PSA[minus]CreERT2 mice were crossed with mT/mG mice to generate bigenic PSA[minus]CreERT2/R26[minus]EGFP mice. The luminal epithelial cells in intact prostate were genetically marked by transient administration of TAM, and the fate of marked luminal cells was determined during prostate regression and regrowth in the ventral prostate and compared to the fate of luminal epithelial cells previously determined in the lateral prostate.[br]Results: The PSA[minus]CreERT2[minus]based genetic lineage marking approach labeled fully[minus]differentiated luminal cells in the intact prostate, but not in the castrated prostate. In intact, non[minus]castrated PSA[minus]CreERT2/R26R[minus]GFP mice, [sim]70% of the luminal epithelial cells in the lateral prostate were labeled by GFP in the lateral prostate; and the percent labeling remained constant between intact and androgen manipulated mice. The efficiency of GFP genetic labeling by TAM was less in the ventral prostate than in the lateral prostate. Approximately 50% of the luminal epithelial cells were labeled by GFP in the ventral prostate, and the percent GFP labeling remained constant between intact and androgen manipulated mice. [br]Conclusions: Pre[minus]existing luminal epithelial cells were shown previously to be a source of regenerated luminal epithelial cells in the adult murine lateral prostate. This self-duplication of luminal epithelial cells was confirmed in the ventral prostate. Prostatic luminal epithelial cells could survive androgen deprivation and were capable of proliferating upon androgen replacement. This mechanism does not appear to be lobe specific.[br][br]Sources of Research Support: National Institutes of Health Grants 5 R37 DK51193, R01 CA 108675, 1 P50 CA90386, and P30 CA047904. LEP was a trainee of NIH T32 DK007774 training program and JL was a recipient of the Mellam Family Foundation Fellowship.[br][br]Nothing to Disclose: LEP, JL, SI, DM, RRG, JP, SK, KW, PB, JN, PC, ZW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 468 51 188 SAT-67 PO35-02 Saturday 127 2012


128 ENDO12L_SAT-68 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Steroidogenic Enzyme Expression in the Human Fetal Prostate Keely McNamara, Elana Rosman, Hiro Sasano, Yasuhiro Nakamura, Shen Yao, Ellen Shapiro, Hongying Huang, Alexander Kirschenbaum, Alice C Levine Tohoku University School of Medicine, Sendai, Japan; Mount Sinai School of Medicine, New York, NY; New York University School of Medicine, New York, NY; Memorial Sloan Kettering Cancer Center, New York, NY [bold]Background: [/bold]Androgens are necessary for normal prostate development. Recent reports demonstrate expression of steroidogenic enzymes in a subset of castration-resistant prostate cancer (PCa) cells and these findings have led to the development of new treatments for advanced disease. We previously demonstrated cell-type specific expression of type II 5-alpha-reductase (SRD5A2) in epithelial and mesenchymal cells of the human fetal prostate. We hypothesize that a subset of epithelial cells in the developing human prostate express steroidogenic enzymes involved in de novo androgen synthesis and that this expression in primary PCa may be a marker for more aggressive disease. [bold]Methods[/bold]: Tissues were obtained from 12 male fetuses (8-22 weeks gestation) at the time of pregnancy interruption and with the approval of the institutional review boards (Mount Sinai and New York University). All samples had been fixed in 10% formalin, embedded in paraffin and later deparaffinized and immuhistochemically stained for c17 and 17[beta]HSD5. Briefly, a citrate based (ph6) antigen retrieval step was used to retrieve the antigens. A methanol/H2O2 block was employed to quench endogenous peroxidase expression. Visualization of the antibody was carried out using the Histofine Kit (Nichirei, Japan) which is based on the principle of streptavidin-biotin amplification. Specificity of C17 staining was verified by antibody preabsorption. [bold]Results: [/bold]There was marked immunoreactivity of c17 in both ductal and acinar epithelial cells as in some stromal cells of 8, 9, 11, 12, 13, 16, 19, 21 and 22 weeks gestation human fetal prostates. There was also some weaker expression of 17BHSD5 in both stromal and epithelial cells of the same specimens. [bold]Conclusion:[/bold] Human fetal prostate epithelial cells express c17 as early as 8 weeks gestation in a subset of cells in the proximal ducts. These c17 positive fetal prostate epithelial cells may represent a progenitor population. PCa cells that similarly express steroidogenic enzymes may, therefore, represent a more primitive, potentially more aggressive subpopulation.[br][br]Nothing to Disclose: KM, ER, HS, YN, SY, ES, HH, AK, ACL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1598 51 189 SAT-68 PO35-02 Saturday 128 2012


129 ENDO12L_SAT-69 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Enhancement of the Antitumor Action of Calcitriol in Ectopic and Orthotopic Xenografts of Human Prostate Cancer Cells Harboring CYP24A1 shRNA in Mice Shaye Kamal Lewis, Julie N Stewart, Josephine B Addai, Hima V Vangapandu, Annisa L Lewis, Kristin Dittmar, Robia G Pautler, Dolores J Lamb Baylor College of Medicine, Houston, TX; Weil Cornell Medical College and The Methodist Hospital, Houston, TX; Weil Cornell Medical College and The Methodist Hospital, Houston, TX; Baylor College of Medicine, Houston, TX In many prostate cancer patients, androgen deprivation therapy results in a temporary reduction of tumor growth followed by androgen-independent tumor growth. Currently, there are no effective treatments for androgen-independent prostate cancer. Calcitriol inhibits the growth of cancer cells from multiple tissues including the prostate. An CYP24A1 is an important enzyme involved in the metabolism of calcitriol into a biologically inactive form. CYP24A1 is over-expressed during advanced stages of prostate cancer leading to calcitriol resistance. We tested the hypothesis that administration of low doses of calcitriol to DU145 prostate cancer cells stably expressing shRNA CYP24A1 will enhance inhibition of prostate cancer growth in xenograft mouse models. Tumors were produced by either intra-prostatic or subcutaneous injection of SCID or nude mice. Calcitriol or sesame oil (vehicle) was administered by oral gavage. MR imaging (Bruker Biospin Pharmascan 7.0T MRI) was done to assess intra-prostatic tumor growth. After 7 weeks of treatment, s.c. DU145 tumors stably expressing shRNA CYP24A1 receiving calcitriol showed a marked reduction in tumor volume compared to all experimental groups (150% difference in tumor volume compared with shRNA controls treated with calcitriol, p[lt]0.0001). Similarly, intra-prostatic tumor volume and proliferation decreased between 60 and 90 days of treatment in shRNA CYP24A1 xenograft-bearing mice treated with calcitriol. Metastasis occurred after orthotopic (38% of orthotopic engraftments, n=34; p[lt]0.0001, Fisher[apos]s exact test) but not s.c. injections. Local tumor invasion was predominantly observed beneath the urothelium of the bladder. Distant tumor dissemination was mostly located in the lymph nodes and diaphragm. Of importance, significantly fewer CYP24A1 shRNA xenograft-bearing animals treated with calcitriol (22%, n= 9; p= 0.0456) displayed metastatic tumors than did the other experimental groups. In both ectopic and orthotopic xenografts experiments, calcitriol treatments were not associated with any significant toxicity as evidenced by the absence of calcium phosphate deposits in the kidneys of all experimental groups. Thus, selective targeting CYP24A1 may facilitate the use of low doses of calcitriol to inhibit growth of prostate cancer cells while avoiding toxicity.[br][br]Sources of Research Support: This work was supported by W81XWH-07-1-0022 PC061154 Idea Development Award from the Department of Defense, Prostate Cancer Research Program Initiative.[br][br]Nothing to Disclose: SKL, JNS, JBA, HVV, ALL, KD, RGP, DJL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2235 51 190 SAT-69 PO35-02 Saturday 129 2012


130 ENDO12L_SAT-70 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Dicer1 Ablation in the Mouse Epididymis Causes Dedifferentiation of the Epithelium and Imbalance in Sex Steroid Signaling Ida Bjorkgren, Anton Krutskikh, Ilpo Huhtaniemi, Matti Poutanen, Petra Sipila University of Turku, Turku, Finland; University of Turku, Turku, Finland; Imperial College London, London, UK; University of Turku, Turku, Finland The epididymis, required for sperm maturation and storage, is a complex organ divided into several segments. Final differentiation of the epididymal epithelium takes place after birth and gives rise to a segment specific gene expression pattern. Recent studies on human and rat epididymides have shown that microRNAs (miRNAs) are differentially expressed at juvenile and adult stages, indicating a role for RNA interference (RNAi) in the postnatal development of the epididymis. To study the role of RNAi in epididymal development and function, we have generated a conditional knock-out (cKO) mouse line of Dicer1, the RNase III enzyme needed for processing pre-miRNAs to functional miRNAs. The lack of Dicer1 in the most proximal segments of the epididymis caused dedifferentiation of the epithelium with increased cell proliferation and a reduction in segment specific gene expression. The [italic] Dicer1 [/italic] cKO epididymis also presented with an imbalance in the expression of sex steroid receptors. Similar to androgen receptor [italic] (Ar) [/italic] cKO mice or mice with excess estrogen signaling, the expression of [italic] Ar [/italic] was downregulated while the expression of Estrogen receptor 1 increased in the most proximal segments of the [italic] Dicer1 [/italic] cKO epididymis. This suggests a role for the RNAi pathway as an important regulator of balanced estrogen-androgen action in the mouse epididymis.[br][br]Nothing to Disclose: IB, AK, IH, MP, PS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1192 51 191 SAT-70 PO35-02 Saturday 130 2012


131 ENDO12L_SAT-71 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Variants in Follicle-Stimulating Hormone Receptor Gene in Infertile Brazilian Men and the Correlation to FSH Serum Levels and Sperm Count Carla Peluso, Milton Ghirelli-Filho, Marcello Machado Gava, Denise Maria Christofolini, Caio Parente Barbosa, Bianca Bianco Faculdade de Medicina do ABC, Santo Andre, Brazil; Faculdade de Medicina do ABC, Santo Andre, Brazil In male, follicle-stimulating hormone (FSH) is essential for normal spermatogenesis and fundamental for Sertoli cell function and the induction and maintenance of qualitatively and quantitatively normal spermatogenesis by a specific receptor (FSHR). The aim of the study was to analyze the distribution of the follicle-stimulating hormone (FSH) receptor ([italic]FSHR[/italic]) Ala307Thr and Asn680Ser polymorphisms in infertile Brazilian men and evaluate the possible role of these polymorphisms on serum levels of FSH and in sperm count. A case-control study was performed comprising 138 infertile men with non-obstructive azoospermia (n=53) or severe oligozoospermia (n=85), and 217 fertile men as controls. Genotyping of [italic]FSHR[/italic] polymorphisms was performed by real time PCR. The results were analyzed statistically and a [italic]p[/italic]-value [lt]0.05 was considered significant. According to the sperm count, relatively similar [italic]FSHR[/italic] polymorphisms genotype and allele frequencies were found among the groups, and combined genotypes of two polymorphisms did not identify a haplotype associated with sperm count. Considering FSH serum level according to genotypes of the Ala307Thr and Asn680Ser polymorphisms individually, statistical analysis showed no difference among the groups. When the combined genotypes of the [italic]FSHR[/italic] polymorphisms were compared to FSH serum levels, no difference was also found among the groups. In conclusion, the findings demonstrate that, in Brazilian population studied, genetic variations, Asn680Ser and Thr307Ala, of the [italic]FSHR[/italic] gene are not correlated with serum FSH levels or sperm count in male infertility.[br][br]Sources of Research Support: This work was supported by grants from FAPESP (Funda[ccedil][atilde]o de Amparo a Pesquisa do Estado de S[atilde]o Paulo) #2011/08681-1.[br][br]Nothing to Disclose: CP, MG-F, MMG, DMC, CPB, BB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 8 51 192 SAT-71 PO35-02 Saturday 131 2012


132 ENDO12L_SAT-72 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Can Intratesticular Injection of Hypertonic Saline Replace Surgical Castration? Eun Young Jeon, Byung Kuk Kwak, Yong-Pil Cheon, Sung-Ho Lee Sangmyung University, Seoul, Republic of Korea; Sungshin Women[apos]s University, Seoul, Republic of Korea Recent evidence revealed that the intratesticular injection of hypertonic saline(20%) resulted in nadir testosterone level and sterile state in rats. To confirm the efficacy of this method in reproductive endocrine studies further, we investigated the anatomical changes and several target gene expression levels in the reproductive tissues. Single dose of hypertonic saline(750 [micro]l/testis) were administered into the testes of adult rats(5 months-old). Bilateral orchiectomy was performed at the same day of injection. Following 1, 2, 4 weeks and 3 months post-injection, the tissues were collected. The body weights were not changed significantly in both orchiectomy group and saline-injection group compared to those in intact group. The wet weights of reproductive tissues(testis, epididymis, seminal vesicle and prostate) and adrenal were significantly decreased in orchiectomy group and saline injected group from 4 weeks post-treatment when compared to those in intact group. The testes exerted slight atrophy and the tunica albuginea seemed to be intact in saline injected group. Histologically, however, larger parts of testicular tissue underwent necrosis and were barely recognizable after hematoxylin-eosin staining. Sloughing of immature germ cells from the basement membrane along with shedding cells in the intraluminal space was notable in most seminiferous tubules from the saline injected testis. In RT-PCR studies, the mRNA levels of hypothalamic GnRH and kisspeptin were significantly elevated in both orchiectomy group and saline injected group. Likewise, the expression levels of the subunits for pituitary gonadotropin(i.e. common alpha subunit, LH-beta and FSH-beta subunit) also were significantly increased. The present study confirmed that the direct injection of hypertonic saline into testis can induce a castration-like, testosterone-depriving effects on the accessory sex organs. Both chemically castrated and surgically castrated animals were almost identical not only in their histological features and but also in their expression patterns of hypothalamus-pituitary reproductive endocrine factors. Our findings suggest that this simple and inexpensive method would be a good replacement for the conventional orchiectomy and the hormone-based chemical castration(e.g. GnRH antagonists treatment).[br][br]Nothing to Disclose: EYJ, BKK, Y-PC, S-HL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1502 51 193 SAT-72 PO35-02 Saturday 132 2012


133 ENDO12L_SAT-73 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Determination of Peripheral Gonadal Hormones after the Treatment of Atypical Antipsychotics, Olanzapine and Risperidone in Wistar Male Rats Tulin Yanik, AK Mehmet, Denis Sezlev, Canan Kursungoz, Bulent Kurt, Ozan Akay, Suleyman Akarsu METU, Ankara, Turkey; GATA, Ankara, Turkey; GATA, Ankara, Turkey Sexual dysfunction is seen in approximately 30-80% of schizophrenic patients, which significantly reduces the quality of life. The antipsychotic treatment is one of the important causes of the sexual dysfunction in these patients. Two of the generally prescribed atypical antipsychotics, olanzapine and risperidone have less extrapyramidal side effects; although, they cause sexual and reproductive dysfunctions in males. The antipsychotics affect the levels of hypothalamus-pituitary-gonad (HPG) hormones that are mainly gonadotropin releasing hormone (GnRH), prolactin, follicle stimulating hormone (FSH), leutinizing hormone (LH) and also total testosterone which may lead to delay in spermatogenesis, decrease in sperm motility and quality, and cause morphological changes in testes. Atypical antipsychotics stimulate both dopaminergic and serotonergic systems and trigger release of LH, FSH, and testosterone. Therefore, we have investigated the peripheral levels of testosterone, LH, FSH and histological properties of the testes in the wistar-male rats (n=15) that were treated with olanzapine (4mg/kg/day) and risperidone (2mg/kg/day) orally for 4-wks. Plasma FSH, LH and testosterone levels were analyzed by ELISA. The plasma FSH levels were decreased by [sim]30% in the olanzapine treated group (1.47 mlU/ml) vs control (2.09 mlU/ml) and increased by [sim]35% in the risperidone group (3.20 mlU/ml [plusmn] 1.35 SEM) vs control. The plasma LH levels did not significantly change in both olanzapine and risperidone groups (0.55mlU/ml) vs control (0.53 mlU/ml). The plasma testosterone levels were also similar in all groups that were 0.43 mlU/ml in olanzapine and 0.69 mlU/ml risperidone groups vs control (0.61 mlU/ml). Although the animals[apos] testes were enlarged during the drug treatment, we did not detect any abnormalities in the diameter of seminifer tubules and their inflammation at the intertubular distance, Leydig cell morphology, as well. Finally, this slight change of FSH, a peptide evidenced to stimulate spermatogenesis, may render a beginning of change in the HPG axis during the atypical antipsychotic treatment.[br][br]Nothing to Disclose: TY, AKM, DS, CK, BK, OA, SA 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2337 51 194 SAT-73 PO35-02 Saturday 133 2012


134 ENDO12L_SAT-74 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Analysis of Herpesvirus Infection in Human Infertility Mo Chen, Masashi Higuchi, Li-Yi Cai, Shun-ichiro Izumi, Takako Kato, Yukio Kato Meiji University, Kawasaki, Japan; Meiji University, Kawasaki, Japan; Meiji University, Kawasaki, Japan; Maternal and Child Health Hospital, Wuxi, China; Tokai University, Isehara, Japan We developed a transgenic (TG) rat using chimera gene composed of porcine FSHb promoter fused to the reporter gene, Human Herpesvirus1 Thymidine Kinase (HHV1-TK). TG rats show ectopic expression of HHV1-TK in the testes and exhibit male infertility. To clarify mechanism underlying the disruption of spermatogenesis in those TG rats is valuable to understand normal spermatogenesis. Recently we demonstrated an accumulation of HHV1-TK in the round spermatid of TG rat testis. In addition, we identified the ectopic transcription start site in the downstream of the translation start site of the HHV1-TK gene, resulting the production of truncated HHV1-TK. Observation that ectopic expression of HHV1-TK is due to an endogenous promoter/transcription site suggests us that HHV1 infected in human expresses its TK gene by ectopic promoter in testis resulting in accumulation of truncated HHV1-TK and this triggers male infertility as shown in HHV-TK TG rat. Hence, in this study, we have investigated a relationship between infection of herpes virus and human male infertility.[br]Semina were donated by Chinese male infertile patients (153 men, aged 21-49 years) with informed consent, followed by DNA preparation for PCR. Semen volume, sperm number and sperm mortality were examined. Nested-PCR was performed for four types of helpes virus. Amplified DNA was extracted after agarose gel electrophoresis and analyzed the nucleotide sequence[br]The virus DNAs of HHV1, HHV3, HHV5 and HHV6 were confirmed by PCR, electrophoresis and DNA sequencing and the virus infection was identified in 59 patients (39%). Number of patients infected were 39 (26%) for HHV1, 33 (22%) for HHV5, 6 (4%) for HHV4 and 3 (2%) for HHV6, respectively. Moreover, double-infection was found in 22 out of 59 specimens (37%), most of which were double-infection of HHV1 and HHV5s. However, relationship between virus infection and sperm abnormality was not identified.[br]This study demonstrated a high-frequency of viral infection in human male infertile patients. However, we did not observe significant difference in abnormality in semen volume, sperm number and sperm mortality between virus-infected and -uninfected patients. The results may indicate that viral infection in human has no relation to male infertility but we did not examine the expression of viral gene, especially viral TK gene. Accordingly, it is indispensable to examine whether viral TK gene is expressed in the testis of virus-infected infertile patient.[br][br]Nothing to Disclose: MC, MH, L-YC, S-iI, TK, YK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1465 51 195 SAT-74 PO35-02 Saturday 134 2012


135 ENDO12L_SAT-75 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) The Putative Humanin Receptor Subunits Are Present in the Testis Prasanth Surampudi, Yue Jia, YanHe Lue, Jeffrey Kim, Christina Wang, Ronald Swerdloff Labiomed at Harbor UCLA, Torrance, CA [bold]Background:[/bold] Humanin (HN), an 24 amino acid mitochondrial peptide, is found in several tissues including the brain and testis. Our previous studies demonstrated that administration of HN peptide protects testicular germ cell from apoptosis induced by hormonal deprivation in vivo [amp] hyperthermia in vitro. It has been demonstrated in neuronal cells that the cytoprotective action of HN is mediated in part through a membrane bound trimeric receptor, comprised of subunits WSX-1, GP-130, and CNTFR[alpha], that utilizes STAT3 as a signaling pathway. Thus, we hypothesize that the cytoprotective action of HN may be mediated through a similar receptor in the testis.[br][bold]Objective:[/bold] We attempted to identify and localize the presence of the HN receptor subunits WSX1, GP130, and CNTFR[alpha] in mouse testes.[br]Methods: Adult (12-week-old) mice (C57BL/6J, control and GnRH antagonist treated for two weeks) were used in the study. Immediately after sacrifice, one testis from each animal was dissected and snap-frozen in liquid nitrogen for protein extraction and Western blot (WB). The contralateral testis was perfused with Bouin[apos]s solution. Paraffin embedded testicular sections were subjected to immunohistochemistry (IHC) and immunofluorescence to determine the localization of the putative HN receptor subunits.[br][bold]Results:[/bold] Western blot showed that WSX1 was detected in the testes as at [sim]70kDa, which was absent in spleen samples from WSX1 knockout mice. GP130 was detected as a 130 kDa protein in the testes. CNTFR[alpha] was not detected in testes by WB using the available antibodies studied previously in neuronal cells. IHC revealed the presence of strong immunoreactivity of WSX1, GP130 [amp] CNTFR-[alpha] in Leydig cells. We found that WSX1 [amp] GP130 were localized in the cytoplasm of spermatocytes, but not CNTFR[alpha]. There were strong immunoexpression of both WSX1 [amp] GP130 in spontaneous apoptotic germ cells in control mice, and in apoptotic germ cells induced by testicular hormonal deprivation in GnRH-antagonist treated mice.[br][bold]Conclusion[/bold]: 1) HN receptor subunits WSX1, GP130 and CNTF[alpha] are present in mouse testes; 2) These three subunits of HN receptor are localized in Leydig cells; 3) Both WSX1 and GP130 are present in the cytoplasm of spermatocytes and apoptotic germ cells. Functional studies are ongoing in our laboratory to determine the role of each receptor subunit. This will help us understand the mechanism of the cytoprotective actions of HN in the testes and its modulation for clinical applications.[br][br]Disclosures: RS: Investigator, Aeterna-Zentaris. Nothing to Disclose: PS, YJ, YL, JK, CW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2274 51 196 SAT-75 PO35-02 Saturday 135 2012


136 ENDO12L_SAT-76 POSTER SESSION: Male Reproduction [amp] Fertility (1:30 PM-3:30 PM) Humanin Is Detected in the Testis after Intraperitoneal (ip) Administration Yue Jia, Junxiang Wan, Yanhe Lue, Aikoui Ohanyan, Kuk Wha Lee, Ronald S Swerdloff, Pinchas Cohen, Christina CL Wang Los Angeles Biomedical Research Institute and Harbor-UCLA Medical Center, Torrance, CA; Mattel Children[apos]s Hospital, David Geffen School of Medicine at UCLA, Los Angeles, CA Objective: Humanin (HN) has been identified as an endogenous cytoprotective peptide. HN protects neuronal, pancreatic beta, and cardiac cells from apoptosis after stress. In our previous study, intra-testicular injection of HN protected male germ cells from apoptosis after hormonal deprivation. The present study determines the distribution of HN in the testis after ip injection of HN.[br]Methods: In experiment 1, adult male mice (C57BL/6, 14 weeks old) received a single ip dose of HN peptide (40 mg/Kg body weight) and sacrificed at 15, 30, 60, 120, and 240 minutes after injection. In experiment 2, male mice were sacrificed at 240 minutes after different ip doses (4, 10, and 40mg/Kg/BW) HN peptide. Control mice only received placebo (saline) injection. One testis of each animal was collected and kept at -80C for Western blot and a specific ELISA assay used to detect the level of exogenous HN. The other testis was fixed with Bouin[apos]s fixative for immunohistochemistry to localize the HN peptide.[br]Results: In experiment 1, two peaks were detected by ELISA after 40 mg/Kg/BW HN ip injection, one at 15 minutes (6.9 ng/mg)(vascular phase) and the second at 240 minutes (8.0 ng/mg) (tissue phase). Western blot analyses confirmed that exogenous HN was detected at 15 and 240 minutes. In experiment 2, HN level in testis was 1.6 ng/mg in the control mice, rose after HN 4 and 10 mg/Kg/BW, reaching 8.6 ng/mg after 40 mg/Kg/BW HN ip injection. Exogenous HN expression was only detected after HN 40mg/Kg injection by immuno-blot. Using immunohistochemistry, specific HN staining was found in Leydig and early germ cells in mouse testis at 240 minutes after 40mg/Kg/BW HN injection.[br]Summary: Four hours after ip injection of 40 mg/Kg/BW HN peptide, exogenous HN peptide was detected in testis and localized mainly in Leydig cells and early germ cells. These data help define target sites for exogenous HN in the testes and may have clinical application since preliminary data from our laboratory shows that ip injection of HN at 40mg/Kg/BW prevents cyclophosphamide induced acute pro-apoptotic effect on male germ cell in mice.[br][br]Disclosures: RSS: Investigator, Aeterna-Zentaris. Nothing to Disclose: YJ, JW, YL, AO, KWL, PC, CCLW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1251 51 197 SAT-76 PO35-02 Saturday 136 2012


137 ENDO12L_SAT-85 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Retinaldehyde Dehydrogenase 1 Is a Novel Regulator of Adaptive Thermogenesis in White Adipose Tissue Florian W Kiefer, Cecile Vernochet, Patrick O[apos]Brian, Steffen Spoerl, Jonathan D Brown, C Ronald Kahn, Jorge Plutzky Brigham and Women[apos]s Hospital, Harvard Medical School, Boston, MA; Harvard Medical School, Boston, MA Two functionally distinct types of fat are present in mammals: white adipose tissue (WAT), the primary site of triglyceride storage, and brown adipose tissue (BAT), which promotes energy dissipation through adaptive thermogenesis. Factors that determine white versus brown fat differentiation and function remain poorly understood. Recent data link vitamin A and its retinoid metabolites to the regulation of adipogenesis and energy balance. Retinoid metabolism is tightly controlled by an enzymatic network in which retinaldehyde dehydrogenases (Aldhs) are the rate-limiting enzymes converting retinaldehyde (Rald) to retinoic acid. Previously, we demonstrated that lack of the Aldh isoform 1a1 protected mice from diet-induced obesity by inducing a hypermetabolic state. However, the mechanisms for this phenotype remained unclear.[br]Here we show that Aldh1a1 is predominately expressed in white but not brown fat. Genetic Aldh1a1 deficiency resulted in increased expression of classic BAT markers in WAT of standard chow-fed mice. Moreover, Aldh1a1-deficient mice manifested increased mitochondrial enzyme activity and oxygen consumption in WAT and were resistant to cold exposure as compared to controls. Using antisense approaches, WAT-selective Aldh1a1 knockdown limited weight gain and improved glucose homeostasis in high fat-fed obese mice by induction of a similar thermogenic program. In investigating underlying mechanisms, we found that Rald, whose endogenous concentrations are elevated in Aldh1a1 deficiency, is a transcriptional regulator of uncoupling protein 1 (UCP1), the key mediator of adaptive thermogenesis. Rald stimulation in white adipocytes increased UCP1 expression 100-fold in a retinoic acid receptor (RAR) but not retinoic X receptor (RXR)-dependent manner. Rald selectively bound and activated RAR but not RXR, induced UCP1 promoter activity, and recruited the co-activator PGC-1[alpha] to the UCP1 promoter.[br]These data establish Aldh1a1 and Rald as novel determinants of adipocyte plasticity and adaptive thermogenesis in white adipose tissue with potential therapeutic consequences for obesity and type 2 diabetes.[br][br]Sources of Research Support: HL048743, AR054604-03S1, NIH/NIDDK DK056626, DK048873, DK048873-14S2, and the Austrian Science Fund (FWF): J3107-B19.[br][br]Nothing to Disclose: FWK, CV, PO, SS, JDB, CRK, JP 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 225 52 198 SAT-85 PO27-02 Saturday 137 2012


138 ENDO12L_SAT-86 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Adipose Tissue[ndash]Specific Deletion of the Mitochondrial Transcription Factor A (TFAM) Causes Lipodystrophy and Hepatosteatosis Cecile Vernochet, Ron C Kahn Joslin Diabetes Center, Boston, MA Mitochondrial dysfunction in adipose tissue has been reported in obesity, type 2 diabetes and metabolic syndrome, but whether this dysfunction participates in the development or is a result of these disorders remains an open question. Mitochondrial transcription factor A (Tfam) plays a key role in mitochondrial function by controlling both mtDNA stability and transcription. To investigate the physiological consequences of mitochondrial impairment in adipose tissue, we generated a mouse deficient for Tfam specifically in adipocytes (Ad-TFKO) using Adiponectin-Cre and Tfam floxed allele. Ad-TFKO mice are leaner than control mice on normal chow diet (CD) by 16% and 28% less after 6 weeks upon HFD. In the CD, Ad-TFKO mice brown adipose tissue was almost inexistent and completely overtaken by white adipose tissue, whereas subcutaneous white fat was reduced by 61% and perigonadal fat by 52%. Ad-TFKO mice also had 14% decreased in basal energy expenditure with no significant change in spontaneous activity. Interestingly, fed glucose levels in Ad-TFKO mice were elevated by 35% compared to controls but fasting glucoses were similar. Ad-TFKO also exhibited a similar glucose clearance during an insulin tolerance test but remained hyperglycemic compared to control throughout. Ad-TFKO has a 2 fold increase in liver mass and histology revealed marked triglycerides accumulation. Thus TFAM KO in fat impaired white and brown adipose tissue expansion and provoked lipid localization to the liver leading to hepatosteatosis and systemic insulin resistance. These results thus suggest that mitochondria dysfunction in adipose tissue could indeed participate and accelerate development of metabolic syndrome.[br][br]Nothing to Disclose: CV, RCK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1550 52 199 SAT-86 PO27-02 Saturday 138 2012


139 ENDO12L_SAT-87 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Unacylated Ghrelin Suppresses High-Fat Diet[ndash]Induced Phenotypic Switch in Adipose Tissue Macrophages, and Enhances Differentiation of Bone Marrow-Derived Endothelial Progenitor Cells Patric JD Delhanty, Lucy Baldeon-Rojas, Martin Huisman, Iris van den Berge, Thierry Abribat, Axel PN Themmen, Pieter JM Leenen, AJ van der Lely Erasmus Medical Center, Rotterdam, Netherlands; Erasmus Medical Center, Rotterdam, Netherlands; Metabolic Diseases, Lyon, France The incidence of obesity and type 2 diabetes (T2D) is of epidemic proportions, although pre-diabetes, characterized by insulin-resistance (IR) occurs in an even larger population. The development of T2D, and associated morbidities, would likely be prevented by reversal of insulin-resistance. We now have evidence that unacylated ghrelin (UAG) has this beneficial effect, although the mechanisms are unclear. Obesity-induced conversion of anti-inflammatory macrophages (MP) to an inflammatory phenotype in fat is an important effector of IR, and is thought to be driven by recruitment of inflammatory monocytes from bone marrow (BM). To investigate if UAG modulates these processes, we infused mice for 4 wks with either saline or UAG. Half these mice were given a normal diet (ND), and half a high fat diet (HFD) during the last 2 wks of infusion to induce IR and increased fat mass. Epididymal fat was removed at wk 4, and the stromal-vascular cell fraction (SVF) isolated and analyzed by FACS. The MP population in the SVF was defined using F4/80 as a marker. Inflammatory (M1) and anti-inflammatory (M2) adipose tissue MPs (ATM) were defined by CD11c or CD301 immunoreactivity, respectively. Compared with the ND, HFD caused a significant 5-fold increase in ATM, accompanied by significant induction (from 6[plusmn]1 to 13[plusmn]0.8%, p[lt]0.001) of M1, and suppression (97.4[plusmn]0.3 to 96.2[plusmn]0.4%, p[lt]0.01) of M2 cells in the ATM population. However, UAG treatment significantly reversed these effects (M1, 6.1[plusmn]1%; M2, 98.8[plusmn]0.2% of ATM). To investigate if this was related to modulation of BM function, we assessed monocyte populations in BM of [plusmn]HFD/[plusmn]UAG treated mice, but surprisingly found no effect of either HFD or UAG. To explore this further, we stimulated BM cells in vitro to develop into three lineages: endothelial progenitor cells (EPC), MPs and dendritic cells (DC). No effect of HFD was observed on recovery of these cell types, and UAG did not affect the differentiation of MPs or DC. However, UAG stimulated the differentiation of EPC 126[plusmn]9% (p[lt]0.05) compared to controls. These data suggest short-term HFD causes an M2 to M1 transition in ATM that is corrected by UAG treatment. Thus, UAG appears to have a direct effect on fat-resident MP phenotype, but not on BM composition. UAG agonists may prevent obesity-induced IR in part by blocking its pro-inflammatory effects in fat. UAG also promoted the potential of BM to generate EPC, with potentially cardiovascular protective properties.[br][br]Nothing to Disclose: PJDD, LB-R, MH, IvdB, TA, APNT, PJML, AJvdL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 921 52 200 SAT-87 PO27-02 Saturday 139 2012


140 ENDO12L_SAT-88 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Estrogen and Retinoid Interactions in the Regulation of Adipose Triglyceride Lipase Barbara Reichert, Rumana Yasmeen, Fangping Yang, Petra Kotzbeck, Margret Poeschl, Rudolf Zechner, Ouliana Ziouzenkova Ohio State University, Columbus, OH; Ohio State University, Columbus, OH; University of Graz, Graz, Austria Visceral obesity is a complex disorder involving dysregulation of fat storage and lipolysis. Identifying mechanisms and regulation of the rate-limiting enzyme of lipolysis in adipose tissue, adipose triglyceride lipase (ATGL), is key in understanding and treating this disorder. Our previous studies implicated vitamin A metabolites, retinaldehyde and retinoic acid (RA), as key regulators of visceral fat accumulation in females. Here, we investigate the effects of estrogen and vitamin A metabolites in regulating ATGL.[br]We utilized ovariectomized mice as a model of postmenopause to examine the roles of vitamin A and estrogen in vivo. We ovariectomized (OVX) or sham-operated wild-type (WT; n=9 OVX, n=19 sham) and Aldh1a1-/- (n=10 OVX, n=10 sham) mice, which lack the principal enzyme for RA production from retinaldehyde in adipose tissue. Tissues were collected three months after surgery from mice receiving a regular chow diet. Ovariectomized WT mice acquired 240% more visceral fat than WT sham. Ovariectomized mouse visceral fat had increased Aldh1a3 (244%) and RA-sensitive Cyp26A1 expression compared to sham, suggesting increased RA production in estrogen-deprived mice. Ovariectomized Aldh1a1-/- mice with disrupted RA production resisted visceral fat accumulation and had improved glucose tolerance. Correspondently, ATGL protein in visceral fat was 190% higher in Aldh1a1-/- compared to WT ovariectomized mice.[br]To investigate the influences of estrogen and vitamin A seen in vivo, we utilized mature 3T3-L1 adipocytes and human SGBS adipocytes in vitro. We stimulated these adipocytes with retinaldehyde or RA in the presence and absence of estrogen. ATGL protein levels were measured by Western blot and lipase activity by non-esterified fatty acid (NEFA)-release assay. 3T3-L1 adipocytes stimulated with retinaldehyde had increased ATGL protein. Retinaldehyde or estrogen increased NEFA release in SGBS adipocytes; however combination of these metabolites did not further increase NEFA release, suggesting a common pathway for retinaldehyde and estrogen action.[br]In conclusion, our data suggest that estrogen suppresses retinaldehyde catabolism by Aldh1, and retinaldehyde, in turn, induces ATGL and protects ovariectomized Aldh1a1-/- female mice from visceral obesity. By elucidating Aldh1[apos]s role in sexually dimorphic fat accumulation, we can potentially target visceral obesity in women.[br][br]Sources of Research Support: Alpha Omega Alpha Carolyn L. Kuckein Student Research Fellowship.[br][br]Nothing to Disclose: BR, RY, FY, PK, MP, RZ, OZ 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1546 52 201 SAT-88 PO27-02 Saturday 140 2012


141 ENDO12L_SAT-89 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) FGF21 Suppresses Lipolysis Via Akt-Dependent Reduction of cAMP Levels in Mouse and Human Adipose Tissues Xuan Ge, Aimin Xu, Cheng Chen, Karen SL Lam The University of Hong Kong, Hong Kong, China; The University of Hong Kong, Hong Kong, China Fibroblast Growth Factor 21 (FGF21) is a liver-secreted hormone that plays a regulatory role in glucose and lipid metabolism in animals. During fasting, free fatty acids (FFA) released from the adipose tissue stimulate hepatic FGF21 production [italic]via[/italic] the activation of PPAR[alpha]. However, the role of FGF21 in lipolysis and the underlying mechanism remain poorly characterized. This study aimed to address these questions using in vivo and ex vivo approaches as well as the FGF21 knockout (KO) mice. A single bolus injection of FGF21 acutely reduced basal and isoproterenol-stimulated serum FFA and glycerol levels in C57 mice and FGF21 KO mice. A greater suppression of lipolysis was seen in the FGF21 KO mice which also had higher baseline serum FFA and glycerol levels. At the molecular level, such an inhibitory effect of FGF21 on lipolysis was accompanied by the decreased phosphorylation of hormone sensitive lipase (HSL) at ser660. Explant studies showed that FGF-21 induced Akt phosphorylation in both mouse and human white adipose tissue (WAT). This was accompanied by an elevation in phosphodiesterase 3B (PDE3B) activity, a reduction in foskolin (FSK)[ndash]stimulated cAMP level, and decreased basal and FSK-stimulated lipolysis, all of which could be attenuated by an Akt inhibitor. Taken together, these findings suggest that FGF21 plays a physiological role in regulating basal lipolysis activity, and suppresses basal and stimulated lipolysis through a reduction in cAMP levels, consequent to Akt-dependent activation of PDE3B, in both mouse and human WAT.[br][br]Sources of Research Support: Acknowledgement: This study was supported by a Collaborative Research Grant (HKU3/CRF/09) from the Hong Kong Research Grant Council.[br][br]Nothing to Disclose: XG, AX, CC, KSLL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 102 52 202 SAT-89 PO27-02 Saturday 141 2012


142 ENDO12L_SAT-90 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) SRA/SRAP Regulates Adipocyte Differentiation and Insulin Sensitivity through Insulin Receptor Signaling Shannon Liu, Ruichuan Xu, Isabelle Gerin, William P Cawthorn, Ormond A MacDougald, Alan R Saltiel, Ronald J Koenig, Bin Xu University of Michigan Medical Center, Ann Arbor, MI; University of Michigan Medical Center, Ann Arbor, MI; Universit[eacute] Catholique de Louvain, Brussels, Belgium; University of Michigan, Ann Arbor, MI; University of Michigan Medical Center, Ann Arbor, MI The Sra1 gene expresses both a non-coding RNA denoted Steroid Receptor RNA Activator (SRA), and an encoded protein (SRAP). Our previous work showed that SRA enhances adipogenesis and insulin sensitivity. To assess the mechanism, we differentiated the ST2 mesenchymal precursor cell line into adipocytes using either a standard hormone cocktail of methylisobutylxanthine, dexamethasone and insulin, or all possible 1 and 2 hormone combinations. These studies showed that SRA overexpression promotes ST2 cell differentiation to adipocytes if the hormone cocktail includes insulin, suggesting that the SRA effect involves insulin signaling. In addition, SRA overexpression inhibits phosphorylation of p38 mitogen activated protein kinase (MAPK) and c-Jun NH2-terminal kinase (JNK) in the early mitotic clonal expansion of ST2 cells. Conversely, knockdown of endogenous SRA/SRAP increases phosphorylation of JNK. Next, we used 3T3-L1 cells to study the effects of SRA/SRAP on glucose uptake. Overexpression of SRAP in 3T3-L1 adipocytes increased insulin-stimulated glucose uptake. Knockdown of endogenous SRA/SRAP in 3T3-L1 mature adipocytes increased basal glucose uptake [sim]2 fold, whereas it inhibited glucose uptake in response to high doses of insulin. IGF-I receptor and GLUT4 proteins were induced by SRA/SRAP knockdown, and may contribute to the associated increased basal glucose uptake. Importantly, knockdown of SRA/SRAP in mature 3T3-L1 adipocytes leads to reduce protein levels of both insulin receptor [beta] (IR[beta]) and its precursor, without changing insulin receptor mRNA levels. This effect is partially eliminated by the proteasome inhibitor MG132, suggesting that SRA/SRAP negatively regulates proteasomal degradation of IR[beta] and its precursor. SRA/SRA knockdown also results in decreased autophosphorylation of IR[beta] and tyrosine phosphorylation of insulin receptor substrate -1 (IRS-1), but these changes are only partially accounted for by decreased IR[beta] protein. We found that SRA/SRAP directly binds to the insulin receptor, suggesting the effects on IR[beta] and its precursor may result from physical interaction. In summary, the present study reveals that SRA/SRAP regulates adipocyte differentiation and insulin sensitivity through maintaining the protein content of the insulin receptor and modulating insulin receptor signaling.[br][br]Nothing to Disclose: SL, RX, IG, WPC, OAM, ARS, RJK, BX 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1570 52 203 SAT-90 PO27-02 Saturday 142 2012


143 ENDO12L_SAT-91 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) The Clock Gene Bmal1 Regulates Adipogenesis Via Wnt Signaling Pathway Bingyan Guo, Somik Chatterjee, Lifei Li, Ji M Kim, Jeongkyung Lee, Vijay K Yechoor, Laurie J Minze, Willa A Hsueh, Ke Ma The Methodist Hospital Research Institute, Houston, TX; Baylor College of Medicine, Houston, TX Circadian clock is present in adipose tissue, but whether it regulates adipocyte development is unknown and the molecular link between circadian dysregulation and obesity is poorly understood. Here we report that disruption of the clock gene, brain and muscle Arnt-like 1(Bmal1), in mice led to increased adipogenesis, adipocyte hypertrophy and obesity, which was evident as early as day 7 of postnatal development without significant alterations in overall energy homeostasis, compared to wild-type controls. This is due to its cell-autonomous effect as Bmal1 deficiency from embryonic fibroblasts, as well as stable shRNA knockdown (KD) in 3T3-L1 preadipocyte and C3H10T1/2 mesenchymal stem cells promoted adipogenic differentiation. We demonstrate that attenuation of Bmal1 function resulted in down-regulation of many genes in Wnt signaling pathway, a known inhibitory mechanism of adipogenesis. Analysis of promoters of these genes ([beta]-catenin, Dishevelled2, TCF3) revealed Bmal1 occupancy, indicating direct circadian regulation by Bmal1, and serum shock was able to elicit their intrinsic circadian oscillation patterns. As a result, Bmal1 KD suppressed Wnt signaling activity while its overexpression augmented this activity. In addition, Wnt3a treatment partially restored the blunted Wnt activity, and in turn, suppressed the increased adipogenesis in KD cells. Taken together, our study demonstrates a novel function of the core clock gene Bmal1 as a negative regulator of adipocyte development and this function is mediated through direct transcriptional regulation of canonical Wnt signaling pathway. Our study suggests that this molecular pathway could be the mechanistic link between circadian disruption and development of obesity.[br][br]Nothing to Disclose: BG, SC, LL, JMK, JL, VKY, LJM, WAH, KM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1950 52 204 SAT-91 PO27-02 Saturday 143 2012


144 ENDO12L_SAT-92 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Testosterone Inhibits Adipogenesis through Modulation of the BMP4 Signaling Pathway Gregorio Chazenbalk, Prapti Singh, David Abott, Dumesic Daniel David Geffen School of Medicine at UCLA, Los Angeles, CA; University of Wisconsin, Madison, WI OBJECTIVE: Adipogenesis is a paracrine-mediated process whereby adipose stem cells (ASCs) become preadipocytes through commitment and then differentiate to adipocytes. Androgen acts through its receptor (AR) to inhibit human subcutaneous (SC) abdominal preadipocyte differentiation. We have previously demonstrated that testosterone (T) inhibits adipogenesis during conversion of ASCs to preadipocytes. The present study utilizes SC abdominal adipose from nonobese women to determine if the inhibitory effect of T in adipogenesis is exclusively mediated through AR events and if occurs during ASC commitment to preadipocytes.[br]DESIGN: Prospective study[br]MATERIAL AND METHODS: Lipoaspirate was obtained from SC abdominal adipose of nonobese patients. ASCs were isolated and then incubated in adipogenic medium for 3 days without and with testosterone (T, 50 nM), 5[alpha]-Dihydrotestosterone (DHT, 5 nM), androgen receptor (AR) antagonist flutamide (F, 100 nM), and Bone Morphogenetic Protein 4 (BMP4, 5 nM), a major regulator of ASC commitment to preadipocytes. Expression of adipogenic gene markers PPARg, CEBP-[alpha] and CEBP-[beta] was determined by qRT-PCR and expressed as fold changes (treated vs. untreated samples).[br]RESULTS: T decreased gene expression of PPARg (0.5 fold, p[lt]0.001), CEBP-[alpha] (0.6 fold, p[lt]0.001) and CEBP-[beta] (0.7 fold, p[lt]0.001). Similarly, DHT decreased gene expression of PPARg (0.4 fold, p[lt]0.002), CEBP-[alpha] (0.5 fold, p[lt]0.003) and CEBP-[beta] (0.7 fold, p[lt]0.001). F partially reversed T inhibition of PPAR[gamma] (0.3 fold, p[lt]0.02 T vs T+F, p[lt] 0.006 F vs F+T), CEBP-[alpha] (0.3 fold, p[lt]0.007 T vs T+F, p[lt] 0.003 F vs T +F) but completely reversed the inhibitory effect of T in CEBP-[beta] (0.3 fold, p[lt] 0.008 T vs T+F, n.s. F vs T+F). BMP4 stimulated PPAR[gamma] (1.96 fold, p[lt]0.010) and CEBP-[alpha] (1.88 fold, p[lt]0.001) and these BMP4 actions were inhibited by T. BMP4 did not alter CEBP-[beta] gene expression.[br]SUMMARY: Androgen inhibits early stages of adipogenesis through the BMP4 signaling pathway and this action appears mediated in part through aromatization of androgen to estrogen.[br]CONCLUSIONS: Androgen excess may diminish SC abdominal adipogenesis and reduce the capacity of this adipose store to safely store fat.[br][br]Nothing to Disclose: GC, PS, DA, DD 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2292 52 205 SAT-92 PO27-02 Saturday 144 2012


145 ENDO12L_SAT-93 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Do Differences in Metabolic Function between Adipose Tissue Depots Play a Role in Obesity Development? Korinna Huber, Mirijam Bedurftig, Jennifer Miles-Chan, Nichola Thompson, Bernhard H Breier University of Veterinary Medicine, Hannover, Germany; University of Fribourg, Fribourg, Switzerland; The University of Adelaide, Adelaide, Australia; Massey University, Albany Campus, Auckland, New Zealand Successful prevention and therapeutic strategies to combat the growing [ldquo]obesity crisis[rdquo] are hampered by our lack of understanding of different pathways to obesity. A number of metabolically distinct pathways to obesity with different health outcomes are recognised (1,2,3). Here we investigate the role of adipose tissue and underlying mechanisms of one such pathway through interrogation of responses to metabolically active hormones [italic] ex vivo [/italic] and the expression of molecular markers of insulin signalling and lipolytic responses in both subcutaneous (ScAT) and retroperitoneal adipose tissue (RpAT).[br]Pregnant Wistar rats were either fed chow [italic] ad libitum [/italic] or undernourished throughout pregnancy, generating control (AD) or prenatally undernourished (UN) offspring (1). At 9 months of age, ScAT and RpAT were collected to investigate insulin-stimulated glucose uptake, catecholamine-induced glycerol release and insulin signalling proteins.[br]The mean adipocyte size was significantly higher (p[lt]0.001) in both types of adipose tissues from UN in comparison with AD offspring. Insulin increased glucose uptake [italic] ex vivo [/italic] in ScAT from both AD and UN offspring (p[lt]0.001); the response to insulin was stronger in AD offspring (p[lt]0.01). In RpAT, insulin stimulated glucose uptake only in AD offspring (p[lt]0.001) but not in UN offspring. While protein expression of insulin signalling markers, InsR, PI3K, PKC[zeta] and mTOR, were similar in ScAT of the two groups, significantly lower levels (p[lt]0.01) were observed in RpAT of UN in comparison with AD offspring. Lipolytic responses to [beta]3-adrenergic stimulation [italic] ex vivo [/italic] differed between adipose tissue depots and were influenced by prenatal nutrition. Lipolytic responses in ScAT were present (p[lt]0.01) in UN offspring, but were significantly lower (p[lt]0.01) compared to AD offspring. The lower lipolytic response in ScAT from UN offspring was completely inhibited by insulin, while in AD offspring, insulin only partially reduced (p[lt]0.01) the stimulation of glycerol release. In contrast, [beta]3-adrenergic stimulation of lipolysis was similar in RpAT from AD and UN offspring and insulin-dependent inhibition of glycerol release was absent.[br]These findings emphasize the role of different adipose tissue depots in the pathophysiology of obesity. A shift in insulin sensitivity and lipolytic responses in different adipose tissue depots may explain specific patterns of fat storage and energy availability that contribute to distinct health consequences.[br][br](1) Thompson NM, Norman AM, Donkin SS, Shankar RR, Vickers MH, Miles JL, Breier BH (2007). Prenatal and postnatal pathways to obesity: different underlying mechanisms, different metabolic outcomes. Endocrinology 148(5): 2345-2354. (2) Huber K, Miles JL, Norman AM, Thompson NM, Davison M, Breier BH (2009). Prenatally-induced changes in muscle structure and metabolic function facilitate exercise-induced obesity prevention. Endocrinology. 150 (9): 4135-4144. (3) Miles JL, Huber K, Thompson NM, Davison M and Breier BH (2009). Moderate daily exercise activates metabolic flexibility to prevent prenatally-induced obesity. Endocrinology. 150 (1):179-186.[br][br]Nothing to Disclose: KH, MB, JM-C, NT, BHB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 841 52 206 SAT-93 PO27-02 Saturday 145 2012


146 ENDO12L_SAT-94 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Nutrition-Independent Morphological Changes in Adipose Tissue during Critical Illness Mirna Bastos Marques, Lies Langouche, Annelies Aertgeerts, Sarah Vander Perre, Sarah Derde, Greet Van den Berghe KU Leuven, Leuven, Belgium We previously reported that adipose tissue biopsies from critically ill patients display an increased number of small adipocytes and a profound infiltration by M[sub]2 [/sub]type macrophages (1,2). Infiltration of M[sub]1 [/sub]type macrophages in adipose tissue has been described in conditions of excessive food intake. It currently remains unknown whether the morphological changes in adipose tissue during critical illness are related to the administration of parenteral nutrition. This study was designed to compare the impact of early parenteral nutrition (PN) with fasting on the morphological alterations within adipose tissue evoked by critical illness.[br]Male mice of 24-weeks were anesthetized and instrumented with central venous catheters. Sepsis was produced by cecal ligation and puncture. Mice were fluid resuscitated and treated with antibiotics. After 24-hour fluid resuscitation, mice were randomly allocated to fasting (n=11) or feeding (PN) (n=11). For comparison, healthy mice (n= 11) were allowed [italic]ad libitum[/italic] chow. Total fat content of the body was quantified with DEXA on days 0 and 5. On day 5, mice were sacrificed and adipose tissue was collected. Adipocyte size was determined in HE-stained sections through computer imaging analysis. Macrophages were identified by ICC (F4/80 Ab) and scored for intensity. Macrophage markers TNF[alpha] (M[sub]1[/sub]) and arginase (M[sub]2[/sub])[sub] [/sub]were quantified with real time PCR.[br]In subcutaneous and visceral adipose tissue from critically ill animals, adipocytes werer significantly smaller than in healthy controls. This change in adipocyte size with illness was unaffected by nutrition. Total fat content was lower in ill than in healthy animals, but again irrespective of nutrition. More macrophages were present in adipose tissue from ill than from healthy mice and, again, this was irrespective of nutrition. The increased number of macrophages in adipose tissue were predominantly of the M[sub]2[/sub] type, as arginase levels were 32-fold increased in all adipose depots, and TNF[alpha] was only slightly elevated.[br]In conclusion, the appearance of small adipocytes infiltrated with more M[sub]2 [/sub]type macrophages during critical illness was independent of PN administration. More small adipocytes during critical illness may reflect an adaptive response to illness, taken their increased capacity to take up glucose and fatty acids. In addition, infiltration of adipose tissue by M[sub]2[/sub] macrophages may be adaptive taken their antiinflammatory and insulin-sensitizing feature. This remains to be investigated.[br][br](1) Langouche L et al, Am J Respir Crit Care Med 2010, 182(4): 507-516. (2) Langouche et al, Crit Care. 2011;15(5):R245.[br][br]Nothing to Disclose: MBM, LL, AA, SVP, SD, GVdB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1431 52 207 SAT-94 PO27-02 Saturday 146 2012


147 ENDO12L_SAT-95 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Increased Mitochondrial Activity in BMP7-Treated Brown Adipocytes, Due to Increased CPT1- and CD36-Mediated Fatty Acid Uptake Kristy L Townsend, Ding An, TianLian Huang, Matthew D Lynes, Hongbin Zhang, Laurie J Goodyear, Yu-Hua Tseng Joslin Diabetes Center, Harvard Medical School, Boston, MA; Boston University, Boston, MA Brown adipose tissue dissipates chemical energy in the form of heat and has recently been shown to actively uptake triglyceride in a regulated manner. Factors that directly control fatty acid uptake and oxidation in brown adipocytes have not yet been fully elucidated. Bone morphogenetic protein 7 (BMP7) is a growth factor capable of inducing brown fat mitochondrial biogenesis during differentiation from adipose progenitors. Administration of BMP7 to mice also results in increased energy expenditure. To determine if BMP7 is able to directly affect the mitochondrial activity of mature brown adipocytes, independent of the differentiation process, we delivered BMP7 to mature brown adipocytes and measured mitochondrial activity. We found that BMP7 increased mitochondrial activity, including fatty acid oxidation and citrate synthase activity. This was accompanied by an increase in fatty acid uptake, and increased protein expression of CPT1 and CD36, which import fatty acids into the mitochondria and the cell, respectively. Importantly, inhibition of either CPT1 or CD36 resulted in a blunting of the mitochondrial activity of BMP7-treated cells. In conclusion, BMP7 increases mitochondrial activity in mature brown adipocytes via increased fatty acid uptake and oxidation, a process which requires the fatty acid transporters CPT1 and CD36.[br][br]Sources of Research Support: K.L.T. was supported by NIH T32-DK007260[ndash]33 and NIH F32DK091996, This work was supported in part by NIH grants DK077097 (Y.-H.T.), and Joslin Diabetes Center[apos]s Diabetes Research Center (P30 DK036836 from the NIDDK), a research grant from the Eli Lilly Research Foundation and by funding from Harvard Stem Cell Institute (to Y.-H.T.).[br][br]Nothing to Disclose: KLT, DA, TH, MDL, HZ, LJG, Y-HT 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 577 52 208 SAT-95 PO27-02 Saturday 147 2012


148 ENDO12L_SAT-96 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Total and High-Molecular-Weight Adiponectin Levels Throughout Life in bGH and GHR-/- Mice Ellen R Lubbers, Aubree K Ziegler, Edward O List, John J Kopchick, Darlene E Berryman Ohio University, Athens, OH; Ohio University, Athens, OH; Ohio University, Athens, OH Bovine growth hormone (bGH) transgenic mice are giant and lean, while growth hormone receptor knockout (GHR-/-) mice are dwarf and obese. The bGH mice have significantly reduced lifespan and reduced insulin sensitivity. In contrast, GHR-/- mice have increased longevity and improved insulin sensitivity. Adiponectin is a powerful insulin sensitizer. It has been suggested that the high molecular weight (HMW) form has the predominate insulin sensitizing bioactivity. The levels of total and HMW adiponectin have been presented in these mice at 6 months of age, with total adiponectin being decreased in bGH mice, but increased in GHR-/-mice compared to wild type (WT) controls. HMW adiponectin and the HMW/total adiponectin ratio was also shown to be increased in GHR-/- mice, but not changed in bGH(1). As adiposity and insulin sensitivity change with age in these mice and a single 6-month time point represents different relative ages in relation to the total lifespan of the mice, an assessment of total and HMW adiponectin throughout life using several time points could help elucidate the role of adiponectin in these unique mouse lines. Serum total and HMW adiponectin were measured side-by-side using an ALPCO ELISA kit. In the bGH mice, total adiponectin is reduced at 2, 6 and 9 months of age, but is not significantly changed at 14 months of age; HMW adiponectin is significantly reduced only at 2 months of age. In bGH mice, the HMW/Total adiponectin ratio showed no significant difference from WT at any age. Circulating total and HMW adiponectin in GHR-/- mice are increased throughout life, at 6, 12 and 24 months of age. Interestingly, the HMW/total adiponectin ratio is increased in GHR-/- mice only at 6-months of age. While observed serum adiponectin levels remain relatively consistent, genotype differences become less pronounced with advancing age. Serum adiponectin will also be compared to leptin and insulin levels in the same samples. Overall, these changes in adiponectin during lifespan highlight the importance of carefully selecting time points for sample collection, as differences of a few months can alter observed genotype differences.[br][br](1)E Lubbers, BMiles, EO List1, JJ Kopchick, DE Berryman. Total and high molecular weight adiponectin levels in mice with altered GH signaling. Poster presented at: Experimental Biology; 2010 April 24-28; Anaheim, CA.[br][br]Sources of Research Support: This work was supported in part by the State of Ohio[apos]s Eminent Scholar Program that includes a gift from Milton and Lawrence Goll, by the AMVETS, by the Diabetes Research Initiative at Ohio University, by NIH grants DK083729, AG031736, and by a Student Enhancement Award.[br][br]Nothing to Disclose: ERL, AKZ, EOL, JJK, DEB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1773 52 209 SAT-96 PO27-02 Saturday 148 2012


149 ENDO12L_SAT-97 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Quantification of Collagen and Effect on Adipocyte Size in bGH Mice Katie M Troike, Lara A Householder, Edward O List, John J Kopchick, Darlene E Berryman Ohio University, Athens, OH; Heritage College of Osteopathic Medicine, Ohio University, Athens, OH; Ohio University, Athens, OH Fibrosis refers to the formation of excess connective or scar tissue in an organ and is usually attributed to changes in the amount and composition of extracellular matrix (ECM) proteins. Growth hormone (GH), a protein secreted from the anterior pituitary, has repeatedly been shown to be positively correlated to ECM deposition in many tissues. The primary ECM protein in adipose tissue is collagen. Increases in collagen have been shown to be directly correlated with a decrease in adipocyte size. Generally, a reduction in adipocyte size would be considered a positive change. However, it has been suggested that it may have negative implications in carbohydrate and lipid metabolism as well as inflammation. To date, no study has assessed the role of GH in ECM deposition in adipose tissue. Bovine growth hormone transgenic mice (bGH) are giant and lean, yet have shortened life spans. In this study, the ECM deposition in adipose tissue was assessed in tissue collected from bGH versus littermate control mice at 10, 26, and 62 weeks of age, in both males and females. Adipose tissue samples from subcutaneous, perigonadal, retroperitoneal, and mesenteric depots were fixed and embedded in paraffin blocks and stained with pico-sirius red. When assessing overall collagen content, the bGH mice show greater ECM deposition when compared to the WT controls at most ages and in both sexes. Furthermore, adipocyte size was reduced in the bGH mice, with the subcutaneous depot showing the most dramatic reduction in size. When comparing different depots, collagen staining appeared most prevalent in the subcutaneous depot as compared to intraabdominal depots in both male and female mice. These results demonstrate a novel role of GH in adipose tissue metabolism and suggest that ECM deposition likely contributes to the depot specific differences in adipose tissue previously reported in mice with modified GH action.[br][br]Sources of Research Support: This work was supported in part by the State of Ohio[apos]s Eminent Scholar Program that includes a gift from Milton and Lawrence Goll, by the AMVETS, by the Diabetes Research Initiative at Ohio University, and by NIH grants DK083729, AG031736.[br][br]Nothing to Disclose: KMT, LAH, EOL, JJK, DEB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2043 52 210 SAT-97 PO27-02 Saturday 149 2012


150 ENDO12L_SAT-98 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Depot Specific White Adipose Tissue Immune Cell Populations in Male C57Bl/6J Mice Stephanie Harshman, Fabian Benencia, Riia Junnila, Edward O List, John J Kopchick, Darlene E Berryman Ohio University, Athens, OH; Heritage College of Osteopathic Medicine, Ohio University, Athens, OH; Ohio University, Athens, OH White adipose tissue (WAT) is a multifaceted organ composed of many cell types within the stromal-vascular fraction (SVF). An assortment of leukocytes is present in SVF including adipose tissue macrophages (ATMs), dendritic cells, and lymphocytes. These immune cells have been shown to have a role in fat tissue inflammation, glucose homeostasis, and insulin resistance through cytokine secretion and cell recruitment. Cell populations change in obese states and appear to alter the endocrine products secreted from adipose tissue and contribute to the inflammation commonly associated with obesity. Many of these general classes of immune cells can be further subdivided based on physiological function. Thus, it is important to not only evaluate the general classes of immune cells present in the tissue under a given condition but also to determine the subclasses in order to better appreciate their role in adipose tissue physiology. Adding to the complexity of adipose tissue are depot specific differences in cellular complexity and endocrine output. To date, little is known regarding the depot differences in immune cell populations or the influence of aging. Therefore, the purpose of this study was to characterize the depot specific differences of immune cell populations in relation to age. Adipose tissue from four distinct depots (inguinal, retroperitoneal, epididymal, and mesenteric) was collected from male C57Bl/6J mice at 5 months of age (young adult) and 21 months of age (aged adult). Dissected adipose tissue was treated immediately with collagenase and the SVF collected. The SVF samples were incubated with a series of fluorphore antibodies and analyzed by FACSAria II flow cytometer with FlowJo software to observe immune cell characteristics. The results demonstrate a unique inflammatory profile in the 21-month subcutaneous and mesenteric depots with significant macrophage and dendritic cell populations compared to retroperitoneal and epididymal depots. In addition, CD45+ cells were identified in subcutaneous, epididymal, and mesenteric depots indicating the presence of B-lymphocytes. The changes in immune cell composition at two age points provided a better understanding of the diverse functionality of immune cell populations, their depot-specific expression, and their potential role in adipose tissue inflammation.[br][br]Sources of Research Support: This work was supported in part by the State of Ohio[apos]s Eminent Scholar Program that includes a gift from Milton and Lawrence Goll, by the AMVETS, by the Diabetes Research Initiative at Ohio University, by a Student Enhancement Award from Ohio University, by an Endocrine Society Summer Fellowship, and by NIH grants DK083729, AG031736.[br][br]Nothing to Disclose: SH, FB, RJ, EOL, JJK, DEB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2099 52 211 SAT-98 PO27-02 Saturday 150 2012


151 ENDO12L_SAT-99 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Effects of Myostatin and Its Propeptide on Transdifferentiation of C2C12 Myoblast to Adipocytes Heng Wang, Ann Yang, Jinzeng Yang University of Hawaii at Manoa, Honolulu, HI; Huazhong Agricultural University, Wuhan, China Myostatin, a key member of TGF-[beta] superfamily in skeletal muscle, inhibits myoblast proliferation and differentiation. There are conflicting reports about the role of myosatin in adipoctyes. Myostatin inhibits preadipocyte differentiation, but promotes adipogeneisis in mesenchymal cells. We previously demonstrated inhibitory effects of myostatin propeptide on myostatin function in both transgenic mice (1-5) and in vivo recombinant protein administration studies (6). Transgenic over-expression of myostatin propeptide results in 76-152% increase in main individual muscles at 12 months of age. Administration of the propeptide to neonatal mice at the age of 11 and 18 days increase 13-25% muscle mass at the age of 57 days. The formations of skeletal myocytes, adipocytes and chondrocytes initially derive from the same multipotent mesodermal precursors during embryonic development. The plasticity of myoblasts to adipogenic and osteogenic lineage determinations is well established in vitro. To investigate the effects of paracrine effects of myostatin on adiposity in skeletal muscle, we initiate studying the effects of myostatin on transdifferentiation of C2C12 myoblast to adipocytes. Myostatin mRNA is expressed in a very low level in proliferating C2C12 cells. After successful transdifferentiation of C2C12 myoblasts into adipocyte-like cells by rosiglitazone and PPAR-[gamma] cDNA tranfection, we demonstrated that expressions of myostatin mRNA were induced by the adipogenic medium during myoblast adipogenesis. The relative level of myostatin mRNA by qPCR showed 20-fold increase compared with that in proliferating cells. The treatment with myostatin (1[mu]g/ml) throughout the transdifferentiation period enhanced adipogenesis as evidenced by the lipid accumulation in cell staining and increased expressions of adipogenic markers of PPAR-[gamma] and C/EBP[alpha]. However, little or no effect of myostatin propeptide on the transdifferentiation was observed. The supplementation of the propeptide at a higher level (5[mu]g/ml) showed inhibitory effects on adipogenesis. These results provide initial cell culture evidences that myostatin enhance adipogenesis during myoblast transdifferentiation, which suggest a different role of myostatin in adipogenesis in aged animals and humans when skeletal muscle is naturally regress to atrophy and accumulation of adipose tissue is prevailing.[br][br]1. Yang J, Ratovitski T, Brady JP, Solomon MB, Wells KD, Wall RJ. 2001. Mol Reprod Dev. 60:351-61. 2. Zhao B, Li EJ, Wall RJ, Yang J. 2009. BMC Genomics 10:305. 3. Yang and Zhao. 2006. Mol. Reprod. Dev. 73:462-9. 4. Zhao B, Wall RJ, Yang J. 2005. Biochem Biophys Res Commun. 337:248-55. 5. Suzuki ST, Zhao B, Yang J. 2008. Biochem Biophys Res Commun. 369:767-73. 6. Li Z, Zhao B, Kim YS, Hu CY, Yang J. 2010. Mol. Reprod. Dev. 77:76-82.[br][br]Sources of Research Support: USDA Hatch/Multistate-National Animal Genome Research Program HAW00244-R; Wuhan [quot]3551 Talent Scheme[quot].[br][br]Nothing to Disclose: HW, AY, JY 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 516 52 212 SAT-99 PO27-02 Saturday 151 2012


152 ENDO12L_SAT-100 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Anti-Inflammatory Effects of Full-Length and Globular Protein Fragments of the New Adipokine CTRP-3 Andreas Schmid, Andrea Kopp, Christa Buechler, Andreas Schaeffler University Hospital of Regensburg, Regensburg, Germany [bold]Introduction[/bold][br]C1q/TNF-related protein-3 (CTRP-3) is a newly discovered adipokine with anti-inflammatory impact on monocytes by down-regulation of pro-inflammatory cytokines, e.g. interleukine 6 (IL-6) and tumor necrosis factor alpha (TNF-alpha), in both adipocytes and monocytes. So far it is unclear which kind of interactions and molecular mechanisms are underlying the observed effects. Our aim is to characterize the differential effects of the globular and the collagenous domain of CTRP-3 and to compare these with the effects exerted by the full length protein.[br][bold]Methods[/bold][br]DNA of recombinant full-length CTRP-3, its globular domain, and its collagen domain were transfected into and expressed in H5 insect cells. The secreted proteins were extracted from the cell culture supernatant and purified by affinity chromatography.[br]Costimulation experiments were performed in 3T3-L1 and monocyte-like THP-1 cells with pro-inflammatory stimuli lipopolysaccharide (LPS) and free fatty acids (FFA) plus CTRP-3 or its globular domain, respectively. Gene expression and secretion of pro-inflammatory cytokines were measured by real time RT-PCR and ELISA techniques. Protein levels of intracellular components of pro-inflammatory signaling were analysed by Western blot.[br][bold]Results[/bold][br]Both full-length CTRP-3 and its globular domain were shown to decrease the amount of secreted IL-6, MCP-1 and resistin after costimulation with LPS and FFA in differentiated 3T3-L1 cells. RT-PCR results confirmed down-regulation of these pro-inflammatory factors on mRNA level. Decrease of secreted IL-6 in THP-1 cells upon costimulation with LPS and CTRP-3 globular domain could be shown as well.[br][bold]Conclusion and outlook[/bold][br]The results suggest that the observed anti-inflammatory impact of CTRP-3 is mediated by its globular domain, whereas the collagen like domain is still to be tested on these effects. As a next step we plan to investigate the anti-inflammatory effects of CTRP-3 in an animal model of LPS-induced SIRS.[br][br]Nothing to Disclose: AS, AK, CB, AS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 20 52 213 SAT-100 PO27-02 Saturday 152 2012


153 ENDO12L_SAT-101 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Mechanisms Underlying Adipocyte Dysfunction Caused by HIV-1 Viral Protein R (VPR) Dinakar Iyer, Neeti Agarwal, Toni Oplt, Ashok Balasubramanyam Baylor College of Medicine, Houston, TX The HIV-1 accessory protein viral protein R (Vpr) disrupts adipogenesis and adipocyte function in vitro. This is due in part due to co-repression of PPAR-gamma (1). Vpr also has other effects in many cell types, e.g., induction of cell cycle arrest at the G2/M phase (by sequestering Cdc25c and keeping the cyclin D/Cdk1 complex hyperphosphorylated); and promotion of apoptosis (by activating JNK, decreasing the ratio of Bcl2 to Bax, and binding to the mitochondrial ANT). We used Vpr mutants to investigate additional mechanisms of adipocyte dysfunction due to Vpr.[br]We used a tetracycline inducible lentivirus system in 3T3-L1 adipogenic cell lines to measure the effects of Vpr and the Vpr R80A mutant (which abolishes Vpr[apos]s ability to cause both G2/M cell cycle arrest and mitochondrial dysfunction, without affecting PPAR-gamma corepression). Synchronized 3T3-L1 preadipocytes were infected with known titers of control vector, Vpr or Vpr R80A constructs, induced for protein expression by doxycyline and allowed to undergo differentiation. Cells were harvested at different intervals (1 day to 10 days after induction), and expression levels of genes involved in adipogenesis were quantified by SyberGreen qPCR assay. Compared to vector-infected cells, Vpr expression induced a profound increase in mRNA expression of cyclins B and D at all time points, indicating altered cell cycle regulation, whereas expression of these cyclins in VprR80A expressing cells was not different from vector-infected controls. Vpr expression caused a global decrease (p[lt]0.01) in expression of genes critical for adipogenesis (PPAR[gamma], C/EBP[alpha], Foxo1) at all time points compared to uninfected or vector-infected cells, with a marked diminution of cells positive for Oil Red O after 10 days. By contrast, Vpr R80A expression permitted preadipocytes to progress through the phases of differentiation with temporally and quantitatively similar adipogenic gene expression as vector-infected cells, with a significantly greater fraction of Oil Red O-positive cells compared to Vpr treatment after 10 days.[br]Thus the Vpr R80A mutant substantially reduces Vpr[apos]s ability to disrupt adipocyte differentiation, indicating that cell cycle arrest, mitochondrial dysfunction, or both, play a prominent role in Vpr[apos]s antiadipogenic effect. Cellular measurements of mitochondrial respiration, glycolysis and fat oxidation are under way to dissect the mechanism.[br][br](1) Shrivastav S et al., 2007 22:234-247.[br][br]Nothing to Disclose: DI, NA, TO, AB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2031 52 214 SAT-101 PO27-02 Saturday 153 2012


154 ENDO12L_SAT-102 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Role of NAD[sup]+[/sup] Signaling through Poly(ADP-Ribose) Polymerase 1 in Adipogenesis Keun Woo Ryu, Xin Luo, William Lee Kraus University of Texas Southwestern Medical Center, Dallas, TX; University of Texas Southwestern Medical Center, Dallas, TX; University of Texas Southwestern Medical Center, Dallas, TX Adipose tissue plays a central role in the development of obesity and insulin resistance, which may involve exceeding the capacity of adipose tissue to expand and subsequent failure to recruit new adipocytes. Adipogenesis is tightly regulated by the sequential regulation of a set of key adipogenic transcription factors. Although the major transcription cascades and protein factors that regulate this process have been identified, our understanding of the precise molecular mechanisms of adipogenesis, particularly during the early stages of differentiation, is incomplete. However, recent studies have shown that nicotinamide adenine dinucleotide (NAD[sup]+[/sup])-dependent enzymes, including poly(ADP-ribose) polymerases (PARPs; e.g., PARP-1) play important roles in adipogenesis.[br]To elucidate the transcriptional regulatory functions of NAD[sup]+[/sup] signaling in adipogenesis, we are using a well-established model of adipogenesis: the murine 3T3L1 preadipocyte cell line. Using a variety of cell-based and molecular assays, we have found that (1) the poly(ADP- ribosyl)ation activity of PARP-1 fluctuates during adipogenesis; (2) RNAi-mediated depletion of PARP-1 alters adipogenesis-related gene expression and modulates the 3T3L1 cell differentiation program; (3) depletion of the nuclear NAD[sup]+[/sup] pools by RNAi-mediated knockdown of nicotinamide mononucleotide adenyltransferase 1 (NMNAT-1), a nuclear NAD[sup]+[/sup] synthase, reduces PARP-1 enzymatic activity enhances the differentiation of 3T3L1 cells; and (4) PARP-1 binds to the promoter regions of adipogenic target genes in a pattern that partially overlaps with the peroxisome proliferator activated receptor gamma (PPAR[gamma]), a key transcriptional regulator of adipogenesis. We are now using a variety of biochemical, molecular, cellular, and genomic assays to elucidate the exact role and regulatory mechanisms of NAD[sup]+[/sup] signaling in adipogenesis.[br]Collectively, these studies will help to elucidate the adipogenic regulatory network, as well as shed light on the mechanisms by which the nuclear NAD[sup]+[/sup] signal can regulate transcription to affect cell proliferation and differentiation. These studies have the potential to reveal new aspects of the pathogenesis of obesity and the potential therapeutic benefits of inhibitors for the NAD[sup]+[/sup]-dependent enzymes.[br][br]Sources of Research Support: This work is supported by a predoctoral fellowship from the DOD/BCRP to X.L. and a grant from the NIH/NIDDK to W.L.K.[br][br]Nothing to Disclose: KWR, XL, WLK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2090 52 215 SAT-102 PO27-02 Saturday 154 2012


155 ENDO12L_SAT-103 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Pyroglutamylated RF-Amide Peptide (QRFP) Gene Is Regulated by Lipopolysaccharide and Interferons in a Cellular Model of Metabolic Endotoxemia between Macrophages and Adipocytes Christian Jossart, Riccarda Granata, Marc G Servant, Sylvie Marleau, Huy Ong Universite de Montr[eacute]al, Montr[eacute]al, Canada; University of Turin, Turin, Italy Background: The orexigenic QRFP has been shown to be involved in feeding regulation. QRFP increases adipogenesis and inhibits lipolysis in 3T3-L1 adipocytes in a paracrine/autocrine manner. The expression of QRFP was found to be induced during differentiation of 3T3-L1 cells. However, in diet-induced obesity (DIO), a mouse model in which plasma concentrations of endotoxin have been shown to be slightly elevated, QRFP expression was decreased compared to lean animals. This metabolic endotoxemia (ME) seems to play a crucial role in the development of obesity. Objectives: 1) To document the mechanisms of regulation of QRFP gene. 2) To assess the effect of ME on QRFP expression in adipocyte and macrophage cell lines. Methods: The expression of QRFP in 3T3-L1 adipocytes and RAW264.7 macrophages was monitored by quantitative PCR following treatments with low doses of lipopolysaccharide (LPS) (1 ng/mL) corresponding to plasma concentration found in ME. TNF-[alpha] (10 ng/mL), palmitate (500 [micro]M) and interferon-beta (IFN-[beta], 100 U/mL) and gamma (IFN-[gamma], 10 ng/mL) have also been tested. Moreover, LPS-treated macrophages conditioned medium has been used on adipocytes. Signaling pathways have been investigated with shRNA knockdowns and pharmacological inhibitors. Results: We report the expression of QRFP in mouse peritoneal macrophages and in RAW and J774 macrophage cell lines. TNF-[alpha] and palmitate did not have any effect on QRFP expression. Low doses of LPS downregulated QRFP by 50% in macrophages but not in adipocytes. A direct, but transient, effect of LPS on QRFP expression in macrophages could be reversed with IKK-2 inhibitor IV or the knockdown of TRIF but not the knockdown of MyD88. Interferons are induced by LPS in macrophages. IFN-[beta] could reduce QRFP expression in both macrophages and adipocytes in similar magnitude than LPS but in a more sustained manner. IFN-[gamma] downregulated QRFP, but only in macrophages. Macrophage-conditioned medium reduced QRFP expression in adipocytes and this effect was blocked with an IFN-[beta] neutralizing antibody. The effect of IFN-[beta] on QRFP expression was blocked by PI3K inhibitor (LY294002) and by p38 MAPK inhibitor (SB203580). Conclusions: LPS induces IFN-[beta] release from macrophages which reduces QRFP expression in both macrophages and adipocytes in an autocrine/paracrine dependent manner, suggesting QRFP as a potential biomarker in ME.[br][br]Sources of Research Support: This work was supported by Aeterna Zentaris.[br][br]Nothing to Disclose: CJ, RG, MGS, SM, HO 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 366 52 216 SAT-103 PO27-02 Saturday 155 2012


156 ENDO12L_SAT-104 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) GRK2 Contribution to the Regulation of Energy Expenditure and Brown Fat Function Sonia Fernandez-Veledo, Rocio Vila-Bedmar, Lucia Garcia-Guerra, Iria Nieto-Vazquez, Federico Mayor, Jr, Cristina Murga IISPV Universitat Rovira i Virgili, Tarragona, Spain; CSIC-UAM, Madrid, Spain; CSIC-UAM, Madrid, Spain; Complutense University, Madrid, Spain Obesity is a major health problem and an important risk factor for the development of multiple disorders. Previous studies in our laboratory have revealed that downregulation of GRK2 decrease age-related adiposity (1), but the physiological and molecular mechanisms underlying this outcome remain unclear. We evaluate whether the lean phenotype results from a direct effect of GRK2 on energy homeostasis. The study of WAT in WT and GRK2+/- littermates showed a reduced expression of lipogenic enzymes and enhanced lypolitic rate in adult GRK2+/- mice. Moreover, hemizygous mice display higher energy expenditure and lower respiratory exchange ratio. Analysis of BAT from adult GRK2+/- mice showed a less deteriorated morphology associated to age compared to WT, which is correlated with a higher basal core temperature. BAT from young GRK2+/- mice showed an increase in gene expression of thermogenesis-related genes. Accordingly, hemyzigous mice display better thermogenic capacity and exhibited a more oxidative phenotype in both BAT and WAT than WT littermates. Overexpression of GRK2 in brown adipocytes corroborated the negative impact of this kinase in BAT function and differentiation. Collectively, our data point at GRK2 inhibition as a potential tool for the enhancement of brown fat activity, which may have important therapeutic implications for the treatment of obesity and associated metabolic disorders.[br][br](1) Garcia-Guerra L et al., Diabetes 2010; 2407-17.[br][br]Sources of Research Support: Ministerio de Educacion y Ciencia (SAF2008-00552 to F.M and BFU2008-04043 to S.F-V); Fundacion Ramon Areces; Comunidad de Madrid (S-SAL-0159-2006);Ministerio Sanidad y Consumo-Instituto Carlos III via The Cardiovascular Network (RECAVA, RD06-0014/0037 to F.M. and PS09/0128 to C.M) and Fondo de Investigacion Sanitaria (FIS) [ldquo]Miguel Servet[rdquo] Grant CP10/00438 to S.F-V.[br][br]Nothing to Disclose: SF-V, RV-B, LG-G, IN-V, FM, CM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1137 52 217 SAT-104 PO27-02 Saturday 156 2012


157 ENDO12L_SAT-105 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) The Expression in and Role of Oxytocin Receptor on Adipose Tissue Lipid Metabolism under Different Feeding Conditions in Mice Sanggun Roh, Kyoungha So, Yutaka Suzuki, Chen Chen, Kazuo Katoh Tohoku University, Sendai, Japan; University of Queensland, Brisbane, Australia Oxytocin, synthesized in hypothalamic neurons, transported to the posterior pituitary gland and released into peripheral circulation, cannot re-enter the brain because of the blood-brain barrier. Through specific and high affinity oxytocin receptor (OXTR), the behavioral effects of oxytocin are thought to reflect only release from centrally projecting oxytocin neurons, separated from hypothalamic ones that project to the posterior pituitary gland. Peripheral actions of oxytocin are mediated by OXTR, which ubiquitously expressed in peripheral tissues including mammary gland. It has been reported that oxytocin intraperitoneal injection induced lipolysis and improved glucose tolerance. However, the role of changing OXTR expression in adipose tissue to have balanced catabolic effects under different metabolic conditions is not clear. The aim of this study was to investigate the expression of OXTR in adipose tissues and possible role in regulating lipid metabolism. To clarify whether OXTR was expressed in isolated adipocytes, its RNA level was analyzed in adipocytes and stromal-vascular cells isolated from mouse adipose tissues. OXTR was highly expressed in adipocytes, with a much lower level of expression in stromal-vascular cells. OXTR mRNA expression was markedly up-regulated in adipose tissues of mice fed a high-fat diet compared with those fed a normal-fat diet. However, OXTR mRNA expression was down-regulated in pituitary gland of mice fed by a high-fat diet. In addition, OXTR mRNA was significantly increased in adipose tissues of db/db mice compared with wild type. To investigate differential gene expression of OXTR in adipose tissue in response to changes in nutritional status, mice was deprived of feeding for 16 hr and then allowed to refeed for 4hr. OXTR gene expression was markedly reduced in adipose tissue of mice deprived of feeding compared with ad libitum fed lean mice, but was increased after refeeding. To investigate whether oxytocin modified the expression of OXTR genes, 3T3-L1 adipocytes were treated with oxytocin (100 pg/ml) for 24 hr. Oxytocin treatment significantly up-regulated OXTR mRNA. These results suggest that up-regulation of OXTR during fat accumulation and adipocyte development may intensify the catabolic effects by oxytocin in adipose tissue to cause lipolysis.[br][br]Sources of Research Support: Grant-in-Aid for Scientific Research from the Ministry of Education, Science, Sports and Culture of Japan.[br][br]Nothing to Disclose: SR, KS, YS, CC, KK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1573 52 218 SAT-105 PO27-02 Saturday 157 2012


158 ENDO12L_SAT-106 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Paradoxical Changes in Glucose Tolerance with Aging in Mice Over-Expressing Human IGFBP3: Possible Role of Brown Fat Hoa Khanh Nguyen, Xing Hai Yao, Suresh Mishra, Gregoire Nyomba University of Manitoba, Winnipeg, Canada; University of Manitoba, Winnipeg, Canada [bold]Introduction:[/bold] Three month-old transgenic (Tg) mice overexpressing human IGFBP3 (BP3) and mutant IGFBP3 (mBP3) lacking IGF binding ability have different degrees of glucose intolerance. In this study we have investigated the role of brown fat in the glucose intolerance. [bold]Methods:[/bold] Glucose status of 3, 6 and 12 month-old BP3 and mBP3 mice was studied by glucose (GTT) and insulin (ITT) tolerance tests, and expression of brown fat marker mRNAs was determined by real time PCR in the interscapular fat tissue. [bold]Results:[/bold] At 3, 6 and 12 months, respectively, body weight was 36.5[plusmn]1.0, 42.4[plusmn]1.2, 41.1[plusmn]0.8g in BP3; 40.2[plusmn]0.8, 45.4[plusmn]1.5, 50.1[plusmn]1.3g in mBP3; 42.7[plusmn]1.0, 48.4[plusmn]1.4, 53.3[plusmn]1.9g in wild-type (WT) mice. Glucose AUC was 1726[plusmn]93[sup][amp][/sup], 1854[plusmn]91, 1548[plusmn]146 in BP3; 1743[plusmn]76[sup][amp][/sup], 2241[plusmn]137[sup]#[/sup], 2131[plusmn]117[sup]#[/sup] in mBP3; 1413[plusmn]32, 1442[plusmn]92, 1382[plusmn]57 in WT mice. K[sub]itt[/sub] was 3.66[plusmn]0.42, 4.16[plusmn]0.42[sup][amp][/sup], 4.03[plusmn]0.24 in BP3; 4.14[plusmn]0.70[sup][amp][/sup], 3.21[plusmn]0.5, 0.98[plusmn]0.6[sup][sect][/sup] in mBP3; 2.34[plusmn]0.42, 2.98[plusmn]0.36, 3.35[plusmn]0.29 in WT mice. Percent brown fat was 0.48[plusmn]0.04, 0.52[plusmn]0.04, 0.51[plusmn]0.04 in BP3; 0.55[plusmn]0.03, 0.80[plusmn]0.4[sup][amp]*[/sup], 0.95[plusmn]0.06[sup][sect]$[/sup] in mBP3; 0.55[plusmn]0.03, 0.58[plusmn]0.05, 0.67[plusmn]0.06 in WT mice. Fat accumulation was greater in the interscapular space of mBP3 vs. BP3. At 6 months, mRNA increased for uncoupling protein-1 (UCP1; 6.16[plusmn]1.59[sup][sect][/sup] vs. 1.25[plusmn]0.19), peroxisome proliferator-activated receptor-[gamma] (PPAR[gamma]; 3.90[plusmn]0.56[sup][amp][/sup] vs. 1.62[plusmn]0.40), PPAR-coactivator-1[alpha] (PGC1[alpha]; 3.45[plusmn]0.57[sup][sect][/sup] vs. 1.30[plusmn]0.16), PGC1[beta] (5.63[plusmn]1.29[sup][amp][/sup] vs. 1.45[plusmn]0.30), Twist (3.53[plusmn]0.80[sup][sect][/sup] vs. 0.99[plusmn]0.11) and PRDM16 (3.72[plusmn]0.52[sup][sect][/sup] vs. 1.18[plusmn]0.10) in BP3 vs. WT mice; mBP3 had increases for PGC1[alpha] (2.75 [plusmn]0.55[sup][amp][/sup]), PGC1[beta] (7.72[plusmn]2.01[sup][sect][/sup]), Twist (3.95[plusmn]0.45[sup][sect][/sup]) and PRDM16 (4.15[plusmn]0.89[sup][sect][/sup]), but not PPAR[gamma] (3.08[plusmn]0.62) or UCP1 (3.25[plusmn]1.09). At 12 months, vs. WT mice, UCP1 (2.63[plusmn]0.42[sup][amp][/sup] vs. 1.11[plusmn]0.09) mRNAs were higher in BP3, while PGC1[alpha] (0.78[plusmn]0.18[sup][amp][/sup] vs. 1.35[plusmn]0.36), PPAR[gamma] (0.78[plusmn]0.11[sup][micro][/sup] vs. 1.17[plusmn]0.14) and PRDM (0.57[plusmn]0.07[sup][amp]*[/sup] vs. 1.01[plusmn]0.07) mRNAs were lower in mBP3. [bold]Conclusion:[/bold] Glucose intolerance occurred in both Tg mice, but was worse in mBP3, and improved with age in BP3 but not in mBP3, which developed insulin resistance with age. Thus, mutant IGFBP3, which does not bind IGFs, led to insulin resistance with reduced expression of brown fat specific genes, whereas these genes were higher in the presence of native IGFBP3. This suggests that brown fat explains differences in insulin sensitivity between the two Tg mice via IGF dependent and independent mechanisms.[br][sup][amp][/sup]p[lt]0.05,[sup][sect][/sup]p[lt]0.01, [sup]#[/sup]p[lt]0.001 vs. WT mice;[sup][micro][/sup]p[lt]0.05, [sup]*[/sup]p[lt]0.01,[sup]$[/sup]p[lt]0.001 vs. BP3 mice.[br][br]Sources of Research Support: The study was supported by a grant from the Canadian Institutes of Health Research (MOP-64306).[br][br]Nothing to Disclose: HKN, XY, SM, GN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 73 52 219 SAT-106 PO27-02 Saturday 158 2012


159 ENDO12L_SAT-107 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Fas (CD95) Induces Lipolysis in 3T3-L1 Adipocytes Via Activation of the ERK1/2 Pathway Reto A Rapold, Stephan Wueest, Adrian Knoepfel, Eugen J Schoenle, Daniel Konrad University Children[apos]s Hospital, Zurich, Switzerland; University Children[apos]s Hospital, Zurich, Switzerland; University of Zurich, Zurich, Switzerland [bold]Background and Aims[/bold]: Fas (CD95) belongs to the superfamily of the tumor necrosis factor (TNF) receptors. Besides its key role in apoptosis, Fas contributes to non-apoptotic pathways such as cell proliferation and inflammation. We previously found that Fas-deficient and adipocyte-specific Fas knockout mice are partly protected from high fat diet-induced insulin resistance. The aim of the present study was to elucidate the underlying molecular mechanisms of this protective effect; in particular the impact of Fas activation on lipolysis in adipocytes.[br][bold]Materials and Methods:[/bold] Fully differentiated 3T3-L1 adipocytes were treated with 2 ng/ml membrane-bound Fas ligand (FasL) in the presence or absence of the MEK inhibitor U0126, the Ca2+ chelator BAPTA/AM, the Ca2+/calmodulin dependent protein kinase II (CaM kinase II) inhibitor KN62 or after siRNA mediated p44/42 MAP kinases (ERK1/2) depletion. Protein was collected for western blot and Luminex analysis. Lipolysis was assessed by measuring free glycerol and free fatty acid release.[br][bold]Results:[/bold] Treatment of adipocytes with FasL induced lipolysis in a time dependent manner with an approximately 2-fold increase after 12 hours. In parallel, Fas activation increased phosphorylation of ERK1/2. FasL-induced lipolysis was blunted in the presence of the ERK-inhibitor U0126 or in ERK1/2 depleted adipocytes. In addition, blocking of the Ca2+/calmodulin kinase pathway (either by a Ca2+ chelator or by a CaM kinase II inhibitor) significantly blunted FasL-induced ERK1/2 phosphorylation and glycerol release.[br][bold]Conclusions[/bold]: Fas activation in mature 3T3-L1 adipocytes induces lipolysis in a CaM kinase II/ERK1/2-dependent manner and, thus, may impact on adipocyte and whole-body metabolism.[br][br]Sources of Research Support: Swiss National Science Foundation.[br][br]Nothing to Disclose: RAR, SW, AK, EJS, DK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 470 52 220 SAT-107 PO27-02 Saturday 159 2012


160 ENDO12L_SAT-108 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Effects of Growth Differentiation Factor 3 (GDF3) on Differentiation and Mature Adipocyte Function Maria Namwanje, Chester Brown Baylor College of Medicine, Houston, TX; Baylor College of Medicine, Houston, TX Growth differentiation factor 3 (GDF3) is a member of the TGF-[beta] family expressed in adipose tissue and its expression is up regulated under high fat diet (HFD) conditions. Gdf3 knock out mice are protected against HFD-induced obesity compared to wild type mice. In contrast, adenoviral transfer of GDF3 in mice results in enhanced sensitivity to HFD including increased adipocyte size and adipose tissue mass relative to wild type mice under the same diet conditions. These results suggest a possible role of GDF3 in regulation of differentiation and/or function of adipocytes. We hypothesize that under HFD conditions, loss of Gdf3 function results in increased lipolysis via effects on [beta]-adrenergic receptor signaling or other mechanisms, or defects in lipogenesis resulting in protection from obesity. We differentiated 3T3-L1 cells into mature adipocytes and treated the cells with GDF3 recombinant protein. We used quantitative RT-PCR to assess the expression of mature adipocyte genes and glycerol and non-esterified fatty acid assays to measure lipolytic activity. GDF3 had no significant effect on the amount of free glycerol and non-esterified fatty acids released at basal and stimulated levels in 3T3-L1cells. Also GDF3 has no effect on expression of lipogenic and lipolytic genes.[br]We then examined the cell autonomous effects of GDF3 on differentiated mouse embryonic fibroblasts (MEFs) from wild type and Gdf3-/- mice. On the whole the Gdf3-/- MEFs had a higher expression of Ppar[gamma] and Prmd16 compared to the wild type cells at day 2 and day 4 after induction of differentiation. The expression of Cebp-[alpha], an upstream regulator of Ppar[gamma] during adipogenesis, was not affected during the differentiation of wild type and Gdf3-/- MEFs. We are currently examining the effects of GDF3 on cell proliferation and apoptosis in MEFs as well as primary preadipocytes.[br][br]Nothing to Disclose: MN, CB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1001 52 221 SAT-108 PO27-02 Saturday 160 2012


161 ENDO12L_SAT-109 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Different Responses to Growth Differentiation Factor 3 in Adipogenic Cell Lines Juan Bournat, Chester Brown Baylor College of Medicine, Houston, TX Growth differentiation factor 3 (GDF3) is a member of TGF-beta superfamily whose expression in white adipose tissues is induced by high fat diet (HFD). Body weight and adipose tissue mass normally increase under HFD conditions, and these processes are augmented when adenovirus-mediated gene transfer overexpresses GDF3 systemically. In contrast, Gdf3 knockout mice have less adipose tissue mass than wild type mice under HFD conditions and exhibit higher basal metabolic rates. These results suggest that GDF3 may act as an adipogenic factor under conditions of caloric excess, and that GDF3 deficiency results in the dysregulation of energy expenditure. Interestingly, the brown adipose markers, CPT1b, PGC1-alpha, and UCP1 are overexpressed selectively in the white adipose tissues of GDF3 knockout mice on HFD, suggesting that GDF3 may play a role in modulating brown versus white adipocyte fate decisions. However, the signaling pathway(s) that mediate these effects are unknown. Recent reports indicate that GDF3 binds BMPs and inhibits their signaling during development. BMPs play important roles in adipogenesis, and BMP7 can induce brown adipocyte differentiation. Moreover, myostatin (GDF8) can inhibit adipogenesis by selectively inhibiting BMP7 signaling. Therefore, we are studying the molecular mechanisms underlying the effects of GDF3 alone and on BMP-dependent adipogenesis using pre-adipocyte cell lines, reporter assays, and quantitative PCR (qPCR). Our results show that GDF3 increases the proliferation of 3T3-L1 a committed white pre-adipocyte cell line, but does not affect the proliferation of C3H10T1/2 a multipotent mesenchymal stem cell (MSC) line. We also find that GDF3 selectively blocks both BMP4 and BMP7 signaling in 3T3-L1 cells, however, GDF3 does not have an effect on BMP4 or BMP7 signaling in C3H10T1/2 cells. In addition, GDF3 alone or in combination with BMPs increases the expression and activity of PPAR-gamma in these preadipocyte cell lines. These observations could be due to intrinsic differences between the cells that affect the expression of signaling components (either present or absent) that contribute to the inhibition of BMP signaling, such as receptors, ligands, co-receptors/ligands, extracellular inhibitors, or downstream mediators of canonical or non-canonical signaling pathways, and have provided insight into the stages and mechanisms by which GDF3 can influence adipogenesis.[br][br]Sources of Research Support: This work was supported by National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) RO1 Grant DK073572 to Dr. Chester Brown. Dr Juan Bournat was supported by a research supplement to promote diversity in health related research from the NIDDK.[br][br]Nothing to Disclose: JB, CB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 992 52 222 SAT-109 PO27-02 Saturday 161 2012


162 ENDO12L_SAT-110 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Trichostatin A (TSA) Modulates TZD Suppression of TNF[alpha]-Induced Lipolysis in Adipocytes Juu-Chin Lu, Yu-Tzu Chang, Chun-Ken Lin, Zhong-Sheng Wu Chang Gung University, Tao-Yuan, Taiwan; Chang Gung University, Tao-Yuan, Taiwan; Chang Gung University, Tao-Yuan, Taiwan Tumor necrosis factor [alpha] (TNF[alpha]) has several effects on adipocytes that may contribute to the development of insulin resistance. It increases local inflammation in adipose tissue and induces basal lipolysis of adipocytes, resulting in elevated serum levels of proinflammatory cytokines, chemokines, and free fatty acids, which have been suggested to mediate pathogenesis of obesity-linked chronic diseases such as insulin resistance and type 2 diabetes mellitus. The peroxisome proliferator-activated receptor [gamma] (PPAR[gamma]) is a nuclear receptor and the key protein controlling adipocyte function. It is also the cellular target of insulin-sensitizing drug thiazolidinediones (TZDs), the synthetic agonists of PPAR[gamma] used in treating diabetic patients. TZD activation of PPAR[gamma] has been shown to antagonize several TNF[alpha] actions in adipocytes, including TNF[alpha]-induced lipolysis, which may account for parts of the insulin-sensitizing effects of TZDs. As a transcriptional factor, the action of PPAR[gamma] is closely modulated by coregulators which include coactivators and corepressors. Previous studies have revealed that in macrophages, insulin-sensitizing effect of PPAR[gamma] may involve suppression of proinflammatory gene expression by recruitment of the corepressor complex that contains corepressors and histone deacetylases (HDACs). However, the role of corepressor complex in adipocytes remains largely unknown. We use a pan-HDAC inhibitor trichostatin A (TSA) to determine the role of HDACs in TZD suppression of TNF[alpha]-induced lipolysis in adipocytes. Treatment of 3T3-L1 adipocytes with TSA increased basal lipolysis in adipocytes. Moreover, inhibition of HDAC activity by TSA also reduced the suppression effect of TZD on TNF[alpha]-induced lipolysis. To determine how TSA modulates TZD suppression on TNF[alpha]-induced lipolysis, downstream signaling molecules involved in TNF[alpha]-induced lipolysis were examined. While TNF[alpha] induced ERK1/2 MAPK phosphorylation, known to be involved in TNF[alpha]-induced lipolysis, treatment of TZDs suppressed TNF[alpha]-induced phosphorylation of ERK1/2. Interestingly, treatment with TSA not only increased basal phosphorylation of ERK1/2, but also reduced TZD suppression on TNF[alpha]-induced phosphorylation of ERK1/2. Further experiments will be required to elucidate possible mechanisms, which may improve therapeutic approaches in the future.[br][br]Nothing to Disclose: J-CL, Y-TC, C-KL, Z-SW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 749 52 223 SAT-110 PO27-02 Saturday 162 2012


163 ENDO12L_SAT-111 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Alendronate Inhibits Adipocyte Differentiation in 3T3-L1 Cells through a Vitamin D Receptor[ndash]Mediated Effect Massimiliano Caprio, Matilde Calanchini, Caterina Mammi, Antonella Antelmi, Francesca Cinti, Giuseppe Rosano, Andrea Fabbri IRCCS San Raffaele Pisana, Rome, Italy; University [ldquo]Tor Vergata, Rome, Italy BACKGROUND: Adipocyte and osteoblast derive from the same mesenchimal progenitor. Age-related decrease in bone mass is accompanied by an increase in marrow adipose tissue. Vitamin D3 (VD) inhibits adipogenesis in 3T3-L1 preadipocytes. Recently it has been demonstrated that Alendronate (ALN) inhibits adipogenesis while promoting osteoblast differentiation of mesenchimal stem cells.[br]AIM OF THE STUDY: To evaluate the in vitro role of ALN on 3T3-L1 adipose differentiation and the potential sinergic role of VD co-treatment.[br]PROCEDURES: Murine 3T3-L1 and 3T3-F442A preadipocytes were routinely differentiated for 7 days adding ALN and VD 10-9,10-8 and 10-7M, then stained with Red Oil. We analyzed through RT-PCR the effect of such treatments upon mRNA expression of main molecular markers of differentiation, PPAR[gamma] and C/EBP[alpha], and VD Receptor (VDR).[br]RESULTS: ALN displayed a marked anti-adipogenic effect on 3T3-L1 cells. VD showed a clear dose-dependent anti-adipogenic effect. Interestingly co-incubation of ALN 10-8M and VD 10-9M did not show synergic effect in inhibition of adipogenesis. PPAR[gamma] mRNA expression was significantly reduced by ALN and VD. mRNA expression of C/EBP[alpha] was reduced only by the highest doses of VD. Interestingly a concomitant increase in VDR mRNA expression was observed in the presence of ALN and VD, suggesting that VDR may represent the molecular target of the anti-adipogenic effect of ALN.[br]To confirm this hypothesis, we explored the effects of ALN and VD on 3T3-F44 cells, which are in a more advanced differentiation stage in adipogenesis than 3T3-L1 cells; interestingly, we found that expression of VDR mRNA was much lower in 3T3-F44 than in 3T3-L1 cells. Moreover, adipose differentiation in this cell model was not affected by ALN nor by VD, differently from what observed in 3T3-L1 cells.[br]CONCLUSIONS: These data represent an indirect evidence of the role of VDR in mediating the anti-adipogenic effect of ALN. Further studies are required to clarify this mechanism.[br][br]Nothing to Disclose: MC, MC, CM, AA, FC, GR, AF 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 802 52 224 SAT-111 PO27-02 Saturday 163 2012


164 ENDO12L_SAT-112 POSTER SESSION: The Adipocyte [amp] Biology of Adipose Tissue (1:30 PM-3:30 PM) Cheonghunhwadam-Tang Inhibits Differentiation to Adipocytes in 3T3-L1 Mouse Fibroblasts Hoyoung Lee, Ji-Hye Lim, Myeong Soo Lee, Min-Ho Cha Korea Institute of Oriental Medicine, Deajeon, Republic of Korea Cheonghunhwadam-tang(CHT), a herbal formula, is have been used in oriental medicine for many centuries as a therapeutic agent of fire and phlegm. It has been composed with 16 herbs including Pinellia ternate (Thunb.) Breit, Citrus unshiu Markovich, Poria cocos Wolf and Atractylodes macrocephala Koidzumi etc. In this study, we examined the efficacy of CHT on adipogenesis. 3T3-L1 cells were differentiated for adipocytes given 1 mg/mL insulin, 0.5 mM isobutylmethylxanthine (IBMX) and 0.25 [mu]M Dexamethasone (DEX) for 7days. After differentiation, cells were stained with Oil-Red-O to detect oil droplets in adipocytes and measured for triglyceride (TG) and glycerol-3-phosphate dehydrogenase (GPDH) activity. We confirmed that PPAR-[gamma], C/EBP-[alpha], Fas and aP2 mRNA expression by RT-PCR and analyzed the gene by microarray in treatment of 3T3-L1 with CHT.[br]These finding showed that CHT suppressed GPDH activity and TG accumulation in a dose-dependent manner significantly. Also, the treatment of CHT reduced the levels of PPAR-[gamma], C/EBP-[alpha], Fas and aP2 mRNA. In microarray analysis, the most down/up expressed pathways were cell cycle and lipid regulation signaling. These results suggest that CHT may prevent obesity and abdominal obesity.[br][br]1) Marie Thompson G etal., Anal Biochem 2004; 21-28. 2) Gregory R etal., Gene 2004; 167-185. 3) Tersuro O etal., Food Chem 2010; 239-244.[br][br]Sources of Research Support: This research was supported by a grant from the Korea Institute of Oriental Medicine (K12130).[br][br]Nothing to Disclose: HL, J-HL, MSL, M-HC 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 635 52 225 SAT-112 PO27-02 Saturday 164 2012


165 ENDO12L_SAT-113 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Body Mass Index and Age Determine the Acrophase of Prolactin in the Human Ferdinand Roelfsema, Hanno Pijl Leiden University Medical Center, Leiden, Netherlands Prolactin has many effects in animals and man. For example, it regulates fat mass in fish, birds and mammals. In particular, the timing of the serum PRL acrophase in relation to the light-dark cycle, or to serum cortisol in constant light conditions, determines whether the fat mass increases or decreases, as part of the adaptation to seasons (1). The role of PRL in this respect has been less well studied in man. Hyperprolactinemic patients have increased fat which decreases after bromocriptine. Other studies, in small groups, have demonstrated (slightly) increased PRL levels in obese subjects, which may be attributed to defective D2R signaling (2), but no detailed studies on PRL acrophase in large groups are available, nor the effect of age on the phase setting. We hypothesized that the timing of the PRL acrophase may be correlated with fat mass (or BMI as proxy) in the human, as observed in animals. We therefore retrospectively analyzed our 24h blood sampling studies in healthy volunteers, in whom both PRL and cortisol were measured. Seventy-four subjects were available, mean age 43 yr (range 22-77 yr) and BMI 26.8 kg/m[sup]2[/sup] (range 18.7-38.4 kg/m[sup]2[/sup]).The PRL acrophase occurred at 03:22 h [plusmn] 13 min (mean [plusmn] SEM) and that for cortisol at 10:03 h [plusmn] 15 min. In the forward selection multivariable regression analysis BMI (P=0.01) and age (P=0.012) were independent negative predictors of the acrophase, but gender was not a significant factor. The cortisol acrophase correlated negatively with age (R=0.308, P[lt]0.009), but not with BMI or gender. The time difference between the cortisol and PRL acrophase was positively correlated with BMI (R=0.418, P[lt]0.001). In various species a wide gap between the cortisol acrophase and that of PRL leads to fat storage. Our findings are consistent with this observation, although we had to use BMI as proxy. If an advance shift of the PRL acrophase in relation to that of cortisol is also responsible for increased fat mass in man, manipulating the PRL phase may open an alternative route for treating obesity.[br][br](1)Meier AH and Cincotta AH, Diabetes Rev 4:464-87, 1996. (2)Kok P et al, JCEM 89:4445-9, 2004.[br][br]Nothing to Disclose: FR, HP 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 34 53 226 SAT-113 PO28-02 Saturday 165 2012


166 ENDO12L_SAT-114 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Regulation of Adiponectin by Free Fatty Acids in Lean and Obese Subjects Kyung Wook Kim, Sy Nam, CW Ahn, KR Kim, JM Yu, Bjoern Richelsen Yongin Severance Hospital, Yongin-si Gyeonggi-do, Republic of Korea; Gangnam Severance Hospital, Seoul, Republic of Korea; Hallym University Kangnam Medical Center, Seoul, Republic of Korea; Aarhus University Hospital, Aarhus, Denmark [italic]Objective[/italic]: Among the adipocyte-derived cytokines, adiponectin possesses an enhancer of insulin sensitivity, anti-inflammatory and antiatherogenic properties. It is suggested that adiponectin plays an important role in lipid metabolism, however, whether free fatty acids (FFA) regulate adiponectin levels remains unclear. The present study was thus undertaken to evaluate the effects of FFAs on the regulation of plasma adiponectin and adipose tissue adiponectin mRNA expression in lean and obese subjects.[br][italic]Methods[/italic]: Seven obese men (BMI 31.3 [plusmn] 1.1 kg/m[sup]2[/sup]) and eight lean men (BMI 21.3 [plusmn] 0.4 kg/m[sup]2[/sup]) were studied. Subjects received an Intralipid 20% (Greencross, Korea) infusion at a rate of 1.5 mL/min over 180-minute period together with sodium heparin at a rate of 1600 IU/h (priming dose 200 IU). Blood sampling and abdominal subcutaneous adipose tissue biopsy through percutaneous mini-liposuction were taken before and after 180-minute Intralipid/heparin infusion.[br][italic]Results[/italic]: The FFA concentration was increased 6 fold after Intralipid/heparin infusion in both lean and obese groups. Adipose tissue adiponectin mRNA was significantly reduced by Intralipid/heparin infusion (38.0 [plusmn] 7.7 [italic]vs.[/italic] 21.1 [plusmn] 3.0.0, [italic]p[lt]0.05[/italic]), and the plasma level of adiponectin was reduced from 5.72 to 5.37 which, however, was a non-significant reduction ([italic]p=0.16[/italic]). If the two groups were separated, in the lean subjects, adipose tissue adiponectin mRNA was significantly lower after Intralipid/heparin infusion (48.3 [plusmn] 12.5 [italic]vs.[/italic] 23.2 [plusmn] 5.0, [italic]p[lt]0.05[/italic]), whereas there was no difference in plasma adiponectin before and after (5.89 [plusmn] 0.6 [italic]vs.[/italic] 5.84 [plusmn] 0.8, [italic]p=0.88[/italic]). In obese subjects, plasma adiponectin concentration was significantly lower after Intralipid/heparin infusion (5.5 [plusmn] 0.8 [italic]vs.[/italic] 4.8 [plusmn] 0.8, [italic]p[lt]0.05[/italic]), whereas there was no difference in adipose tissue adiponectin mRNA expression (26.0 [plusmn] 6.3 [italic]vs.[/italic] 18.7 [plusmn] 3.1, [italic]p=0.24[/italic]).[br][italic]Conclusions[/italic]: Albeit the other data on the effect of FFAs on adiponectin concentrations are not consistent, we could get that FFAs suppress the expression of adiponectin in lean and obese human. The issue of possible FFA-induced regulation of adiponectin warrants further investigations.[br][br]Nothing to Disclose: KWK, SN, CWA, KRK, JMY, BR 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 59 53 227 SAT-114 PO28-02 Saturday 166 2012


167 ENDO12L_SAT-115 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Morbidly Obese Hypothyroid Patients Require an Increase in Weight-Based Thyroid Hormone Dose Following Bariatric Surgery Umasankari Sundaram, Valerie K Shostom, Jane L Meza, Corrigan L McBride, Melissa M Monzu-Sparks, Whitney S Goldner University of Nebraska Medical Center, Omaha, NE; University of Nebraska Medical Center, Omaha, NE; University of Nebraska Medical Center, Omaha, NE [bold]Background[/bold][br]The relationship between morbid obesity and thyroid hormones is complex and obesity itself can be accompanied by changes in thyroid function. In addition, hypothyroidism is common in morbidly obese patients. Previous studies have been inconsistent and have reported an increase, decrease, and no change in total thyroid hormone dose following bariatric surgery. We sought to evaluate the change in thyroid hormone dose following bariatric surgery in hypothyroid morbidly obese patients and the factors that influenced the change.[br][bold]Methods[/bold][br]We retrospectively analyzed 231 hypothyroid morbidly obese patients on levothyroxine replacement (LT4) who underwent Roux-en-Y-gastric bypass, lap banding or sleeve gastrectomy at The University of Nebraska Medical Center(UNMC) between January 2000 and December 2010. We analyzed for change in LT4 dose (both actual dose and weight based dose (mcg/kg)) and correlation between change in dose and confounding variables including change in weight, BMI, TSH, and vitamins B12 and 25OHD.[br][bold]Results[/bold][br]The prevalence of hypothyroidism in morbidly obese patients undergoing bariatric surgery at UNMC was 19% (231/1221). Complete pre and post-operative thyroid data was available on 74 patients. Overall, 19/74 (26%) had a decrease, 41/74 (55%) had no change, and 14/74 (19%) had an increase in actual LT4 dose. Pre and post-operative median TSH was not significantly different (pre: 2.64 mcIU/ml, post: 2.0 mcIU/ml) and median LT4 dose was also not different pre and post-operatively (125 mcg for both) between groups. However, the weight based LT4 dose following surgery was significantly increased post-operatively (pre: 0.95 mcg/kg) versus (post: 1.14 mcg/kg) (p=[lt]0.01). There was no significant difference between patient[apos]s minimum vitamin B12 and 25OHD levels and change in actual and weight based dose, however, there was a trend for the lowest levels of B12 and 25OHD being inversely associated with both actual and weight based LT4 dose.[br][bold]Conclusion[/bold][br]We observed an increase in weight based LT4 dose following bariatric surgery. However, actual LT4 dose changes were variable, and the majority had no change. This suggests malabsorption as a major contributor to post-operative changes in thyroid hormone dose rather than weight loss alone as the main contributor to thyroid hormone dose changes following bariatric surgery.[br][br]Nothing to Disclose: US, VKS, JLM, CLM, MMM-S, WSG 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 234 53 228 SAT-115 PO28-02 Saturday 167 2012


168 ENDO12L_SAT-116 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Testosterone Therapy with Testosterone Undecanoate Long-Acting Intramuscular Injections in Hypogonadal Men Is Associated with Reduction in Obesity Aksam Yassin, Youssef El Douaihy, Ridwan Shabsigh, Aiman Yassin, Farid Saad Segeberger Kliniken, Norderstedt, Germany; Dresden International University, Dresden, Germany; Gulf Medical University School of Medicine, Ajman, United Arab Emirates; Maimonides Medical Center, Brooklyn, NY; Klinikum Braunschweig, Braunschweig, Germany; Bayer Pharma AG, Berlin, Germany; Gulf Medical University School of Medicine, Ajman, United Arab Emirates [bold]Introduction[/bold]: Obesity and the metabolic syndrome are frequently associated with late onset hypogonadism (LOH). We sought to study the effects of testosterone replacement therapy on weight, body mass index (BMI) and waist circumference (WC) in patients with LOH.[br][bold]Materials and Methods[/bold]: As of November 2004 130 patients with LOH (diagnosis criteria: total testosterone [le]3.5ng/dl and documented erectile dysfunction (ED) according to Sexual Health Inventory for Men score (SHIM; [le] 21) were included in a prospective cohort study to investigate the effect of long-acting testosterone undecanoate (TU) 1000 mg intramuscular injection. Treatment was initiated at day 1 (T1), then the second dose was administered 6 weeks later (T2). TU was injected at 3 months intervals thereafter. The parameters weight and waist circumference were measured at baseline and at every treatment visit. Serial BMI, WC and percentage of weight change from baseline were calculated.[br][bold]Results[/bold]: Median follow-up time was 4.7 years. The mean weight decrease from baseline to the last visit was 14.3 [plusmn] 8.7 kg (minimum -5, maximum 44), the mean percentage weight loss was 13.0 [plusmn] 6.8% (minumim -5.3, maximum 34.4), the mean of BMI points decreased was 4.5 [plusmn] 2.7 (minimum -1.6, maximum 13.4), and the mean drop in WC was 11 cm [plusmn] 6 (minimum -13, maximum 24). There is a strong linear relationship between weight loss, decrease in BMI and decrease in WC and time on TU treatment.[br][bold]Conclusion[/bold]: Long-term testosterone treatment with TU results in significant reduction of all parameters of obesity.[br][br]Sources of Research Support: Data entry was supported by Bayer Pharma AG.[br][br]Disclosures: AY: Speaker, Bayer Schering Pharma. RS: Speaker, Lilly USA, LLC. FS: Employee, Bayer Schering Pharma. Nothing to Disclose: YED, AY 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 477 53 229 SAT-116 PO28-02 Saturday 168 2012


169 ENDO12L_SAT-117 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) 15 Years of Experience with Intramuscular Testosterone Undecanoate for Substitution in Male Hypogonadism [mdash] Beneficial Effects on the Metabolic Syndrome and High Safety Profile Michael Zitzmann, Farid Saad University Clinics, Muenster, Germany; Gulf Medical University, Ajman, United Arab Emirates; Bayer Health Care, Berlin, Germany [bold]Background:[/bold][br]A reliable form of androgen substitution therapy in terms of favorable kinetics and tolerance as well as effective restoration of androgenicity is paramount for hypogonadal men. The intramuscular injection of the long-acting ester testosterone undecanoate (TU) offers a convenient modality for testosterone substitution.[br][bold]Methods:[/bold][br]We report data from 334 patients (147 with primary, 100 with secondary hypogonadism and 87 with late-onset ([ldquo]mixed[rdquo] or [ldquo]metabolic[rdquo] hypogonadism) aged 15 to 72 years (mean 42[plusmn]15 years) receiving altogether 6596 intramuscular injections of 1000 mg of TU during a maximal treatment time of 15 years, overall corresponding to 1403 treatment years.[br]Components of the metabolic syndrome were assesed in 269 men receiving 4296 injections.[br][bold]Results:[/bold][br]Individual dosing intervals ranged from 10 to 14 weeks Serum T concentrations increased from 5.8 to stable 16.1 nmol/L within the first year of treatment. The proportion of men fulfilling the new Harmonized Criteria for definition of the Metabolic Syndrome decreased from initially 88% to 52% within 2 years (Chi-square for trend: p[lt]0.001). During the maximal duration of treatment, an overall favorable change from baseline was visible for a multitude of parameters related to androgen effects/metabolic risk (see Table). Prostate size increased from 16.1[plusmn]5.2 to 21.1[plusmn]5.2 ml (p[lt]0.001), whilst PSA levels increased moderately (1.8[plusmn]0.4 to 1.9[plusmn]0.4[micro]g/l, p=0.001). No case of prostate cancer was observed. Hematocrit was significantly elevated during treatment but remained within the normal range (40.9[plusmn]2.1 to 46.2[plusmn]2.5%, p[lt]0.001). One patient suffered from deep vein thrombosis, one from stroke. No case of prostate cancer was observed.[br][bold]Parameter [rarr] Unit [rarr] Baseline [rarr] Endpoint 15 years [rarr] p[/bold] (ANOVA)[br]BMI [bold][rarr][/bold] kg[sup]x[/sup]m[sup]-2[/sup] [bold][rarr][/bold] 31.8[plusmn]5.2 [bold][rarr] [/bold]24.4[plusmn]3.2 [bold][rarr] [/bold][lt]0.001[br]Waist circ. [bold][rarr] [/bold]cm [bold][rarr] [/bold]114[plusmn]10.5 [bold][rarr] [/bold]94.1[plusmn]8.7 [bold][rarr] [/bold][lt]0.001[br]Weight [bold][rarr] [/bold]kg [bold][rarr] [/bold]103.0[plusmn]16.3 [bold][rarr] [/bold]79.1[plusmn]12.6 [bold][rarr] [/bold][lt]0.001[br]LDL-Cholesterol [bold][rarr][/bold] mg/dl [bold][rarr] [/bold]157[plusmn]29 [bold][rarr] [/bold]110[plusmn]19 [bold][rarr] [/bold][lt]0.001[br]HDL-Cholesterol [bold][rarr] [/bold]mg/dl [bold][rarr] [/bold]38.4[plusmn]9.7 [bold][rarr] [/bold]53.6[plusmn]11.7 [bold][rarr] [/bold][lt]0.001[br]Triglycerides [bold][rarr] [/bold]mg/dl [bold][rarr] [/bold]198[plusmn]33 [bold][rarr] [/bold]145[plusmn]21 [bold][rarr] [/bold][lt]0.001[br]Fasting glucose [bold][rarr] [/bold]mg/dl [bold][rarr] [/bold]118.1[plusmn]29.7 [bold][rarr] [/bold]91.2[plusmn]15.2 [bold][rarr] [/bold][lt]0.001[br]RR systolic [bold][rarr] [/bold]mmHg [bold][rarr] [/bold]148[plusmn]14 [bold][rarr] [/bold]128[plusmn]11 [bold][rarr] [/bold][lt]0.001[br]RR diastolic [bold][rarr] [/bold]mmHg [bold][rarr] [/bold]98[plusmn]11 [bold][rarr] [/bold]81[plusmn]10 [bold][rarr] [/bold][lt]0.001[br]Pulse [bold][rarr] [/bold]bpm [bold][rarr] [/bold]89[plusmn]9 [bold][rarr] [/bold]75[plusmn]8 [bold][rarr] [/bold][lt]0.001[br][bold]Conclusion:[/bold][br]Intramuscular injections of TU represent a feasible, safe and well tolerated modality of androgen substitution in hypogonadal men, substantiated by 15 years of experience, facilitating a decrement of metabolic/cardiovascular risk factors.[br][br]Disclosures: FS: Employee, Bayer Schering Pharma. Nothing to Disclose: MZ 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1470 53 230 SAT-117 PO28-02 Saturday 169 2012


170 ENDO12L_SAT-118 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Restoring Testosterone to Normal Levels in Elderly Men Is Efficacious in Weight Reduction. A Follow-Up Study over 5 Years Farid Saad, Ahmad Haider, Louis Gooren Bayer Pharma AG, Berlin, Germany; Gulf Medical University School of Medicine, Ajman, United Arab Emirates; Private Practice, Bermerhaven, Germany; VUMC Amsterdam, Amsterdam, Netherlands [bold]Introduction:[/bold] Obesity is associated with reduced testosterone, and low testosterone induces weight gain. This study analysed the effects of normalization of serum testosterone in mainly elderly, hypogonadal men.[br][bold]Methods:[/bold] Open-label, single-center, cumulative, prospective registry study of 255 men (aged 38 [ndash] 83 years, mean 60.6 [plusmn] 8.0 years), with testosterone levels between 1.7 [ndash]3.5 ng/mL (mean: 2.87 [plusmn] 0.4). Cut-off point for testosterone treatment was serum testosterone [le] 3.5. ng/mL). 215 men were studied for at least 2 years, 182 for 3 years, 148 for 4 and 116 for at least 5 years. They received parenteral testosterone undecanoate 1000 mg/12 weeks after an initial interval of 6 weeks.[br][bold]Results:[/bold] After 5 years the following changes were observed: weight (kg) decreased from 106.22 [plusmn] 16.93 (minimum: 70, maximum: 139) to 90.07 [plusmn] 9.51 (min 74.00, max 115). The statistical significance was p[lt]0.0001 vs baseline and vs the previous year over 5 years indicating a continuous weight loss over the full observation period. Waist circumference (cm) declined from 107.24 [plusmn] 9.14 (min 86, max 129) to 98.46 [plusmn] 7.39 (min 84, max 117) (p[lt]0.0001 vs baseline and vs the previous year over 5 years). Body mass index (BMI, m/kg[sup]2[/sup]) declined from 33.93 [plusmn] 5.54 (min 21.91, max 46.51) to 29.17 [plusmn] 3.09 (min 22.7; max 36.71) (p[lt]0.0001 vs baseline and vs the previous year over 5 years). The mean per cent weight loss after 1 year was 4.12 [plusmn] 3.48%, after 2 years 7.47 [plusmn] 5.01%, after 3 years 9.01 [plusmn] 6.5%, after 4 years 11.26 [plusmn] 6.76% and after 5 years 13.21 [plusmn] 7.24%. At baseline, 96% of men had a waist circumference of [ge] 94 cm. This proportion decreased to 71% after 5 years.[br][bold]Conclusions:[/bold] Raising serum testosterone to normal produced loss of body weight, waist circumference and BMI. These improvements were progressive over the full 5 years of the study.[br][br]Sources of Research Support: Data entry and statistical analysis were supported by Bayer Pharma AG.[br][br]Disclosures: FS: Employee, Bayer Schering Pharma. AH: Principal Investigator, Bayer Schering Pharma. LG: Speaker, Bayer Schering Pharma. 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 495 53 231 SAT-118 PO28-02 Saturday 170 2012


171 ENDO12L_SAT-119 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Relationship between Cortisol and Components of the Metabolic Syndrome in Obese Children and Adolescents Chiara Guzzetti, Patrizia Zavattari, Sabrina Pilia, Anastasia Ibba, Maria Rosaria Casini, Luigi Minerba, Sandro Loche Ospedale Microcitemico, Cagliari, Italy; Universita[apos] degli Studi di Cagliari, Cagliari, Italy; Universita[apos] degli Studi di Cagliari, Cagliari, Italy [bold]Background.[/bold] A relationship between serum cortisol (F) and insulin resistance has been shown in obese adults and children, suggesting a role for F in the development of the metabolic syndrome (MS). In this regard, F could be an important predictor factor for MS.[br][bold]Objective.[/bold] The aim of this study was to evaluate the relationship between F and components of MS in obese children and adolescents.[br][bold]Subjects and Methods.[/bold] This is a retrospective study in 1027 obese children and adolescents (BMI-SDS 2.62[plusmn]0.51) referred to our endocrine unit in the last 8 y. Waist circumference (WC), systolic and diastolic blood pressure (SP, DP), morning serum concentrations of F, glucose (Glyc), cholesterol HDL, tryglicerides (TG) and HOMA (Glyc(mmol/L)xInsulin(mU/L)/22.5) were evaluated in all subjects. MS was defined according to the IDF criteria (1). Patients were subdivided into 3 age groups: 6-10, 10-16 and [gt]16 y.[br][bold]Results.[/bold] In univariate regression analysis including the entire cohort, F was weakly associated with SP (r=0.17, P[lt]0.001), DP (r=0.13 P[lt]0.001), Glyc (r=0.16 P[lt]0.001) and HOMA (r=0.15, P[lt]10[sup]-5[/sup]), also when adjusted for age, gender, puberty and BMI-SDS. 14/372 patients aged 10-16 y had MS. Univariate regression analysis in these 372 subjects showed that F was associated with SP (r=0.11, P=0.047), DP (r=0.16 P=0.003) and Glyc (r=0.21 P[lt]0.0001) but not with HOMA. When adjusted for age, gender, puberty and BMI-SDS, F was no longer correlated with SP but was weakly correlated with HOMA (r=0.12, P=0.02). In multivariate regression analysis including age, sex, puberty, BMI-SDS and F as independent variables and one of the component of the MS as the dependent variable, the only models where F was a weak predictor of the variability were one with DP (B=0.02, P=0.009, adjusted R[sup]2[/sup]=0.13) and one with Glyc (B=0.03, P=0.003, adjusted R[sup]2[/sup]=0.12) as dependent variables. Patients were then subdivided into 4 groups according to the WC centile: WC[lt]90, WC[gt]90, WC[gt]90 and 1 of the other components of the MS, WC[gt]90 and 2 of the other components of the MS. Mean F concentrations in the 4 groups was significantly different (F=7.12, P=0.0001), even adjusted for age, gender, puberty and BMI-SDS (F=4.53, P=0.004). Patients with one or more components of the MS had higher F concentrations.[br][bold]Conclusions.[/bold] In this cohort of obese children and adolescents, F was weakly associated with components of the MS. These findings suggest that F has a minor role, if any, in the development of MS.[br][br](1)Zimmet et al., Pediatric Diabetes 2007; 8:299[ndash]306.[br][br]Nothing to Disclose: CG, PZ, SP, AI, MRC, LM, SL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 478 53 232 SAT-119 PO28-02 Saturday 171 2012


172 ENDO12L_SAT-120 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Body Fat Is Inversely Correlated with Urine Free Cortisol in Human Subjects Nicola M Neary, Brent S Abel, Xiongce Zhao, Michael S Ring, Gail R Hall, Rana Malek, Kong Y Chen, Lynnette K Nieman, Monica C Skarulis National Institutes of Health, Bethesda, MD; National Institutes of Health, Bethesda, MD; University of Maryland School of Medicine, Baltimore, MD [bold]Introduction: [/bold]Patients with anorexia nervosa are known to have increased plasma and urine free cortisol (UFC). We recently reported that UFC was elevated over 4-fold in a male with low body fat (1). In these situations, elevated cortisol may protect against further loss of visceral fat. We postulated that cortisol plays a role in fat regulation over a wider range of adiposities and hypothesized that body fat is inversely related to UFC in adults. The relationships between body fat as a percent of body weight (BF%) and AM and PM cortisol were also examined.[br][bold]Methods:[/bold] BF% was measured by DEXA in volunteers enrolled in a longitudinal study of overweight and obese adults. Inpatient assessment of the HPA axis included 24h UFC and mean diurnal serum cortisol obtained at 07:30, 08:00 and 23:30, 23:59. Patients treated with exogenous steroids, including creams and inhalers, or with diagnoses or treatment of depression, diabetes or obstructive sleep apnea or weight instability were excluded. For statistical analysis, log serum cortisol and square root UFC were used.[br][bold]Results: [/bold]212 subjects (89 males, 113 females) aged 18-69 (median 40, IQR 28-52) years, BMI 18-52 (median 28.7, IQR 24.1-34.7) kg/m[sup]2[/sup], of white (n=106, 50%), African American (n=83, 39%) and other (n=23, 11%) racial origin participated. UFC was median 21 (IQR 14-34) [mu]g/24h [normal range (nr) 3-45], AM cortisol: 10.9 (IQR 8.8-14.1) [mu]g/dl (nr 5-25), PM cortisol 2.8 (IQR 2.1-4.3), and BF% 35.4 (IQR 27.2-44.7). In univariate analysis, there was a weak but significant inverse correlation between BF% and UFC, R[sup]2[/sup]=0.08, p[lt]0.0002. After adjusting for race, sex, and age, BF% was inversely associated with AM cortisol (p=0.0321), but not with PM cortisol or UFC. Females had higher BF% than males (40.2[plusmn]0.9 (mean[plusmn]SEM) vs. 28.4[plusmn]1.0; p[lt]0.0001) but lower UFC (22.1[plusmn]1.4.2 vs. 29.7[plusmn]1.8, p=0.0002). Similarly, African American subjects had higher BF% than white subjects (37.8[plusmn]1.2 vs. 32.7[plusmn]1.1; p=0.0062) but lower UFC (20.9[plusmn]1.4 vs. 29.5[plusmn]1.9; p=0.0010).[br][bold]Summary: [/bold]In univariate analysis, BF% was weakly inversely correlated with UFC Adjusting for age, sex and race, BF% was inversely correlated with AM serum cortisol. There were significant differences in UFC between sexes and between racial groups that could relate in part to differences in body fat. Thus glucocorticoids may play a physiological role in fat preservation when the HPA axis is intact in addition to their critically adverse effect in excess as seen in Cushing[apos]s syndrome.[br][br](1)Neary NM et al., ENDO 2011; P3-226.[br][br]Sources of Research Support: National Institute of Diabetes and Digestive and Kidney Diseases; The Eunice Kennedy Shriver National Institute of Child Health and Human Development.[br][br]Nothing to Disclose: NMN, BSA, XZ, MSR, GRH, RM, KYC, LKN, MCS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1576 53 233 SAT-120 PO28-02 Saturday 172 2012


173 ENDO12L_SAT-121 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Estradiol Levels in Morbidly Obese Patients: Relationship with Metabolic and Inflammatory Markers Diana Teixeira, Diogo Pestana, Carla Sa, Sonia Norberto, Manuela Meireles, Gil Faria, Paula Freitas, Conceicao Calhau, Rosario Monteiro Faculty of Medicine, University of Porto, Porto, Portugal; S Jo[atilde]o Hospital, Faculty of Medicine, University of Porto, Porto, Portugal; S Jo[atilde]o Hospital, Faculty of Medicine, University of Porto, Porto, Portugal Obesity-related inflammation is a subject of great interest, not only for the exponential growth of incidence this pathology but also because obesity-driven inflammation may be an important instigator of the metabolic abnormalities that accompany the obese state. The dysregulation of estrogen metabolism has been associated with the susceptibility to obesity. However, the influence of estrogens on obesity-related inflammation has been little explored.[br]The aim of this study was to evaluate the relationship between total estradiol and inflammatory and metabolic markers in a sample of obese (BMI [gt] 40) patients undergoing bariatric surgery.[br]Twenty six men (age: 42.9 [plusmn] 9.3; BMI 44.6 [plusmn] 9.3; estradiol: 46.5 [plusmn] 33.9) and 221 women (age: 41.8 [plusmn] 10.6; BMI: 44.6 [plusmn] 4.9; estradiol 57.33 [plusmn] 63.90) divided by menopausal status (90 premenopausal: age: 38.0 [plusmn] 8.9; BMI 44.6 [plusmn] 4.3; estradiol: 66.6 [plusmn] 68.9; and 41 postmenopausal: age: 51.3 [plusmn] 8.7; BMI 44.9 [plusmn] 5.3; estradiol: 36.1 [plusmn] 40.8).[br]Body mass index (BMI), waist and hip circumference, blood pressure, lipid profile, plasma glucose, insulin, HgbA1c, estradiol, morning cortisol, sex hormone binding globulin (SHBG), testosterone, and hs-CPR were measured during standard clinical evaluation. Adipose tissue (subcutaneous and visceral) samples were collected during surgery at the Hospital of S. Jo[atilde]o, Portugal (protocol approved by the Ethics Committee of hospital) to determine size and number of adipocytes.[br]Plasma estradiol levels were positively correlated with BMI and HgbA1c in men and negatively correlated with morning cortisol levels. When all women were analyzed, estradiol levels were negatively correlated with: age, SHBG, testosterone and adipocyte number in visceral adipose tissue. In premenopausal women, estradiol levels were negatively associated with BMI, hip circumference, SHBG and positively correlated with alteration in HDL ([gt] 50 mg/dL) and IR-HOMA. There are no such significant correlations in postmenopausal women. No significant associations were found between estradiol and hs-CRP.[br]In conclusion, estradiol levels in men seem to be a marker of adiposity, and in women, seem to predict metabolic dysfunction, at least when all premenopausal women are analyzed as a whole.[br][br]Sources of Research Support: This work was supported by FCT (POCI, FEDER, Programa Comunit[aacute]rio de Apoio, PTDC/QUI/65501/2006; SFRH/BD/64691/2009; SFRH/BD/46640/2008, SFRH/BD/78367/2011,and SFRH/BPD/40110/2007).[br][br]Nothing to Disclose: DT, DP, CS, SN, MM, GF, PF, CC, RM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1446 53 234 SAT-121 PO28-02 Saturday 173 2012


174 ENDO12L_SAT-122 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Effects of GH Withdrawal in Obese Premenopausal Women with Abdominal Adiposity Eleanor Lin, Miriam A Bredella, Anu V Gerweck, Andrea L Utz, Karen K Miller Massachusetts General Hospital, Boston, MA; Vanderbilt, Nashville, TN [bold]Context[/bold]: Abdominal adiposity is associated with decreased growth hormone (GH) secretion. We previously reported results from a 6-month randomized, placebo-controlled study, which demonstrated that GH replacement in abdominally obese premenopausal women decreased total abdominal adipose tissue (TAT), subcutaneous abdominal adipose tissue (SAT), trunk fat, tissue plasminogen activator (tPA), apolipoprotein B (apoB), low-density lipoprotein (LDL), and high-sensitivity C-reactive protein (hsCRP), and increased thigh muscle and total lean mass. This was accompanied by a mild increase in both fasting and 2-hour glucose after oral glucose tolerance test (OGTT).[br][bold]Objective[/bold]: To determine whether GH replacement for 6 months has long-term effects on cardiovascular risk biomarkers, body composition, or glucose tolerance in obese women.[br][bold]Design[/bold]: Seventy-nine abdominally obese premenopausal women were allocated to GH (Genentech, Inc.) or placebo for 6 months. Thirty-nine completed a subsequent 6-month withdrawal observation period.[br][bold]Results[/bold]: Mean age (37[plusmn]1y) and BMI (34.9[plusmn]0.9kg/m[sup]2[/sup]) were comparable between the 39 subjects who completed the 6-month withdrawal period and the larger group that was initially randomized to GH or placebo. Insulin-like growth factor 1 (IGF-1) standard deviation scores (SDS) within the GH group were -1.7[plusmn]0.1 (pretreatment), -0.1[plusmn]0.3 (after 6 mos of GH) and -1.7[plusmn]0.1 (6 mos post GH withdrawal). Six months after GH withdrawal, TAT, SAT, total fat, trunk fat, trunk/extremity fat, hsCRP, apoB, LDL, and tPA increased ([italic]p[/italic][lt]0.05) by repeated-measures analysis of variance. There was a trend toward an increase in total cholesterol and extremity fat and a decrease in 2-hour OGTT glucose ([italic]p[/italic][lt]0.10). All body composition and cardiovascular risk marker outcome measures that had improved with GH returned to baseline levels by 6 months after GH discontinuation ([italic]p[/italic][gt]0.05). Fasting and 2-hour OGTT glucose returned to pre-treatment levels.[br][bold]Conclusion[/bold]: The effects of GH administration to abdominally obese premenopausal women have a short time-course. The beneficial effects on body composition and cardiovascular risk outcome measures, and the side effect of altered glucose tolerance, reverse after GH withdrawal; all return to pretreatment levels. Importantly, there was no indication of suppression of endogenous GH levels, as IGF-1 levels returned to baseline after withdrawal of GH therapy.[br][br]Sources of Research Support: This work was supported in part by National Institutes of Health Grants R01 HL-077674, UL1 RR-025758, K23 RR-23090, and T32 DK007028. Study medication and placebo only were supplied by Genentech, Inc.[br][br]Nothing to Disclose: EL, MAB, AVG, ALU, KKM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 679 53 235 SAT-122 PO28-02 Saturday 174 2012


175 ENDO12L_SAT-123 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Longitudinal Changes in Body Mass Index and Body Composition among Adult Survivors of Childhood Cancer Karin Blijdorp, Marry M van den Heuvel-Eibrink, Rob Pieters, AJ van der Lely, Sebastian JCMM Neggers Erasmus University Medical Centre Roterdam, Roterdam, Netherlands; Sophia Children[apos]s Hospital/Erasmus University Medical Centre Roterdam, Rotterdam, Netherlands Objective[br]Obesity, represented by high body mass index (BMI), is a major complication after treatment for childhood cancer. High amount of total and visceral fat and low lean body mass are described as more reliable determinants, predicting the development of cardiovascular disease. In this study longitudinal changes of BMI and body composition in adult childhood cancer survivors were evaluated.[br]Design Retrospective single center cohort study, evaluating longitudinal changes in BMI and body composition.[br]Subjects Data of 417 adult childhood cancer survivors, who had visited the late effects clinic twice, were analyzed. Median follow up time was 16 years (interquartile range 11-21) and time between visits was 3.2 (2.9-3.6). At both time points BMI was measured and body composition was assessed by dual X-ray absorptiometry (Lunar Prodigy). BMI and body composition measures were compared with those of healthy Dutch references and calculated as standard deviation scores (SDS).[br]Growth hormone deficient subjects treated with growth hormone replacement at time of follow up were excluded from further analyses.[br]Results[br]BMI SDS at 1st assessment was only significantly higher in female cranial radiotherapy (CRT) survivors as compared to healthy Dutch references (SDS=0.40, p=0.02). Increase of BMI over time (expressed as units per year) was only significantly higher in male survivors (0.27 versus 0.02 in controls (p[lt]0.001)). Percentage fat was significantly higher than controls in both men (SDS 1.37, P[lt]0.001) and women (SDS 1.05, P[lt]0.001) in all therapy groups, with the highest SDS after CRT (mean SDS 1.73 in men, 1.48 in women, P[lt]0.001). Only in men, increase in total fat percentage was significantly higher as compared to controls ([Delta]SDS=0.22, P[lt]0.001). Lean body mass did not significantly change over time.[br]Conclusion[br]Significantly greater increase of BMI and total fat percentage as compared to healthy references was found, especially in adult male survivors of childhood cancer.[br][br]Nothing to Disclose: KB, MMvdH-E, RP, AJvdL, SJCMMN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 627 53 236 SAT-123 PO28-02 Saturday 175 2012


176 ENDO12L_SAT-124 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Comparison of Fat Mass Index and Percent Body Fat as Measures of Adiposity in Youth David R Weber, Renee H Moore, Mary B Leonard, Babette S Zemel Children[apos]s Hospital of Philadelphia, Philadelphia, PA; University of Pennsylvania, Philadelphia, PA; Children[apos]s Hospital of Philadelphia, Philadelphia, PA; Children[apos]s Hospital of Philadelphia, Philadelphia, PA Background: Body Mass Index (BMI) of [ge]85th percentile is widely used to identify individuals at risk for excess adiposity and metabolic disease; however the ability of BMI to measure adiposity in youth may differ by racial group. Recent studies have suggested percentage body fat (PBF) as a superior index of adiposity in youth. The use of PBF as the gold standard of adiposity is an incomplete solution because it fails to account for the effect of height (HT). We hypothesize that fat mass index (fat mass/ht[sup]2[/sup], FMI) will more accurately represent adiposity in youth.[br]Methods: The LMS method was used to generate reference curves and Z-scores for FMI using total fat mass obtained from dual energy x-ray absorptiometry(DXA) scans in 8983 (3775 F) subjects aged 8-25 in 1999-2004 as a part of the National Health and Nutrition and Examination Survey. Comparisons of BMI and PBF were performed in 7123 (2903 F) subjects aged 8-20. Age and sex specific Z-scores for BMI (BMIZ) and HT (HTZ) were calculated using 2000 CDC growth reference data. PBF percentiles were defined using published reference data(1). Excess adiposity was defined as FMI or PBF [ge]75th percentile. Two-sample t-tests were used to compare continuous variables; chi-square tests for proportions.[br]Results: The positive predictive value (PPV) of BMI [ge]85th percentile to identify adiposity defined by FMI was lower in blacks (M:36%, F:30%) compared to whites (M:65%, F:52%), and Mexican Americans (M:73%, F:68%); (p[lt]0.001). Among subjects differentially classified by FMI and PBF, mean HTZ (M:0.71, F:0.48), BMIZ (M:1.54, F:1.62) and proportion black (M:17%, F:33%) were higher in subjects identified as having excess adiposity by FMI but normal by PBF, compared to HTZ (M:-0.04, F:-0.24), BMIZ(M:0.54, F:0.73) and proportion black (M:5%, F:5%) in subjects identified as having excess adiposity by PBF but normal by FMI (p[lt]0.02).[br]Conclusion: In a representative cohort of youth, the PPV of BMI [ge]85th percentile to identify excess adiposity defined by FMI is markedly lower in blacks. Subjects classified as having excess adiposity by FMI but normal adiposity by PBF are taller, have higher BMI, and are more likely to be black. We hypothesize that this represents a population of youth with high lean mass and high fat mass who may be at risk of metabolic disease but would be missed if using PBF to identify excess adiposity. Further studies are planned to investigate the effects of lean and fat mass on metabolic risk.[br][br](1)Ogden CL, Li Y, Freedman DS, Borrud LG, Flegal KM, Smoothed percentage body fat percentiles for US children and adolescents, 1999-2004. Natl Health Stat Report 2011;43:1-7.[br][br]Nothing to Disclose: DRW, RHM, MBL, BSZ 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 632 53 237 SAT-124 PO28-02 Saturday 176 2012


177 ENDO12L_SAT-125 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Proteomic Identification of Morbidities in Overweight and Obese Prepubertal Children Nabila Roohi, Ateek Ahmad University of the Punjab, Lahore, Pakistan; Lahore General Hospital, Lahore, Pakistan [bold]Background and aims: [/bold]Childhood obesity is associated with significant morbidity and mortality in adult life. Proteomic identification of plasma signatures may be a valid tool for predicting and tracking the progression of comorbidities in overweight/obese children.[br][bold]Subjects and Methods:[/bold] We assessed plasma proteome of prepubertal boys (Mean age: 10.76[plusmn]0.23 years) with BMI 5[sup]th [/sup]to [lt]85[sup]th[/sup] percentile (healthy weight; n=12), 85[sup]th [/sup]to [lt]95[sup]th[/sup] percentile (overweight; n=20) and [ge]95[sup]th[/sup] percentile (obese; n=20) by two dimensional gel electrophoresis using Protean IEF coupled with II XL Cell System. The resolved proteins were identified by Swiss 2DPAGE database.[br][bold]Results: [/bold]By comparative proteome analysis, eight spots; five up-regulated and three down-regulated were differentially expressed in overweight and obese children in comparison to their normal weight counterparts. The expressions of Apolipoprotein (Apo) A-I, ApoA-IV and ApoE were significantly suppressed (p[lt]0.05) in 45%, 25% and 25% of overweight, whereas, 60%, 30% and 35% of obese children, respectively. ApoB-100, C-reactive protein (CRP), Retinol binding protein (RBP), Fibrinogen beta chain and Plasminogen activator inhibitor-1 (PAI-1) indicated marked expressions (p[lt]0.05) in 40%, 25%, 30%, 35% and 20% of overweight, while, 55%, 30%, 35%, 45% and 30% of obese children, respectively. The altered expressions of ApoA-I, ApoA-IV, ApoE, CRP, Fibrinogen and PAI-1 predict that overweight, in particular, obese children are at a higher risk of future progression towards cardiovascular disorders, particularly, atherosclerosis. Further, the over-expressions of CRP and RBP, in these children, demonstrae the risk for the development of insulin resistance and type 2 diabetes, in their later life.[br][bold]Conclusions: [/bold]Prominent alterations in the expressions of plasma proteins, in obese and overweight prepubertal children, may serve as novel diagnostic and prognostic biomarkers of morbidities in adulthood. Overweight, not only the obese, children should be screened for biomarkers. Extensive weight control strategies, for the prevention and reversal of overweight/obesity in children, may be warranted at population level.[br][br]Nothing to Disclose: NR, AA 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 602 53 238 SAT-125 PO28-02 Saturday 177 2012


178 ENDO12L_SAT-126 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) High Second-Trimester Body Mass Index and Offspring Neurocognitive Development Wendy Y Craig, Glenn E Palomaki, Louis M Neveux, James E Haddow Foundation for Blood Research, Scarborough, ME; Women [amp] Infants Hospital/Alpert Medical School of Brown University, Providence, RI Among women and their offspring serving as controls for a separate nested case-control study, we observed an unexpected inverse association between second trimester maternal body mass index (BMI) and neurocognitive development at two years old. We explored this association further in that dataset, and in a second control group from an earlier nested case-control study that measured neurocognitive development in eight year old offspring. Both study groups were recruited from the Maine pregnancy population; demographic and pregnancy data were obtained at the time of prenatal screening and from the birth record. We tested 101 children (mothers pregnant 2004-6) using the Bayley Scales of Infant Development (BSID-III), and 118 children at age eight years (mothers pregnant 1987-90), using the Wechsler Intelligence Scale for Children (WISC-III). Frequency of maternal BMI [ge]30 kg/m[sup]2[/sup] increased from 10% to 30% in the 15 years between studies (p[lt]0.001), and the socioeconomic gradient with BMI became more pronounced (measured by the Hollingshead score at the time of neurocognitive testing). Regression analysis showed an inverse relationship between BSID-III language scores and log BMI in two year old offspring, after adjustment for potential effect modifiers (r = -0.16, slope = -26.810, p = 0.05; intercept = 138.8594); there was no significant relationship between BMI and either cognitive or motor scores. The inverse relationship between WISC-III performance subscale IQ scores and log BMI at eight years remained significant after adjustment (r = -0.20, slope = -34.873, p= 0.023; intercept = 148.534); there was no significant relationship between BMI and either full scale IQ or verbal subscale scores. Maternal obesity may be an independent risk factor for children[apos]s neurocognitive development. Confirmatory studies are needed.[br][br]Nothing to Disclose: WYC, GEP, LMN, JEH 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 670 53 239 SAT-126 PO28-02 Saturday 178 2012


179 ENDO12L_SAT-127 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Relationship of Body Composition with Bone Mass in Korean Adults Kyung-Soo Kim, Soo-Kyung Kim, Se-Hwa Kim, Seok Won Park, Yong-Wook Cho CHA Bundang Medical Center, CHA University, Seongnam, Republic of Korea; Kwandong University College of Medicine, Goyang, Republic of Korea A relationship between bone mineral density (BMD) and body mass index or weight has been established. However, the effect of body composition such as fat or muscle mass on bone mass remains controversial. The aim of the present study was to examine the relationship of body composition with bone mass in Korean adults based on a large population-based survey.[br]Cross-sectional analysis were carried out on 3643 Korean adults (1928 men, 705 premenopausal women, 1010 postmenopausal women) who were 40 years or older and participated in the Fourth Korea National Health and Nutrition Examination Survey conducted in 2009. Total and regional fat mass, lean mass, and bone mineral content (BMC), BMD were measured by dual energy X-ray absorptiometry.[br]In normal (18.5[le]BMI[lt]23) and overweight (23[le]BMI[lt]25) group of premenopausal women, lumbar spine BMC was significantly higher in low waist circumference (WC) ([lt]80cm) than in high WC ([ge]80cm) after adjustment of age and weight (P[lt]0.05). Also in overweight group of postmenopausal women, lumbar spine BMC was significantly higher in low WC than in high WC after adjustment of age and weight (P[lt]0.05). In men, lumbar spine BMC was high in low WC in overweight group but in high WC in obese (BMI[ge]25) group. We showed both total fat mass and total lean mass were positively correlated with BMD (femur neck, lumbar spine) in all three groups. However, after controlling for age and weight, the association between total fat mass and BMD (femur neck, lumbar spine) became negative (P[lt]0.05). Furthermore, after controlling for age and weight, there were no significant association between appendicular fat mass and femur neck BMD although truncal fat mass still showed a negative association in premenopausal and postmenopausal women. In contrast, there was a positive association between total and regional lean mass (arm and leg) and BMD (femur neck and lumbar spine) after adjustment of age and weight in all three groups.[br]In conclusion, fat mass was negatively correlated with BMD (femur and lumbar spine) and lean mass was positively after removing the mechanical loading effect in Korean adults. Especially, in femur neck BMD, truncal fat mass was more important than appendicular fat mass in women.[br][br]Nothing to Disclose: K-SK, S-KK, S-HK, SWP, Y-WC 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 688 53 240 SAT-127 PO28-02 Saturday 179 2012


180 ENDO12L_SAT-128 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Gender Difference in Secular Trends of Anthropometric Parameters among Korean Adolescents, 1998[sim]2008 Ki Eun Kim, Shin Hye Kim, Sang Shin Park, Mi Jung Park Gangnam CHA Hospital, CHA University, School of Medicine, Seoul, Republic of Korea; Sanggye Paik Hospital, Inje University, College of Medicine, Seuol, Republic of Korea; College of Veterinary Medicine and Biomedical Sciences, Texas A[amp]M University, College Station, Canyon, TX; Sanggye Paik Hospital, Inje University, College of Medicine, Seoul, Republic of Korea Objectives[br]The aim of this study was to investigate national trends of body physique [height, weight, body mass index (BMI), and waist circumference] in Korean adolescents.[br]Methods[br]We analyzed data for 2,822 subjects aged 10-18 yr (1,478 boys and 1,344 girls) from the Korean National Health and Nutrition Examination Surveys conducted between 1998 and 2008.[br]Results:[br]The final height of boys increased from 172.5 [plusmn] 6.6 cm in 1998 to 175.5 [plusmn] 5.1 cm in 2008 (P=0.008), while there was no significant change in the final height of girls. Over the study period, mean values of body weight, BMI and waist circumference were significantly increased in boys; however no significant changes were observed in girls. The proportion of boys with normal weight was decreased from 82.8% to 71.4% (P[lt]0.0001), while the proportion of overweight (from 6.3% to 14.7%, P[lt]0.0001) and obese boys (from 4.7% to 8.2%, P=0.004) were significantly increased. No significant difference in proportions of normal weight, overweight and obese girls was seen over the study period, however increasing trend in proportion of underweight girls was observed in elementary school girls (from 4.8% to 9.4%, P=0.05).[br]Conclusions[br]Secular trend of anthropometric parameters in Korean adolescents was marked by increasement of mean final height, obesity prevalence in teen aged boys, and emergence of underweight in elementary school girls. Age and gender targeted measures to prevent obesity and underweight are warranted in health policies for Korean youth.[br][br]Nothing to Disclose: KEK, SHK, SSP, MJP 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 708 53 241 SAT-128 PO28-02 Saturday 180 2012


181 ENDO12L_SAT-129 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Normative Values of Anthropometric Indices among Residents of Calabar, South East Nigeria Ofem Egbe Enang, Okon Ekwere Essien, Olufemi Adetola Fasanmade, Augustine Efedaye Ohwovoriole University of Calabar Teaching Hospital, Calabar, Nigeria; Lagos University Teaching Hospital, Lagos, Nigeria Background: Ethnic specific anthropometric values have been proposed by the International Diabetes Federation (IDF) to aid in the diagnosis of the metabolic syndrome. No such values are available for Sub-Saharan Africa including Nigeria.[br]Objectives: To determine and propose normative values of indices of anthropometry among the inhabitants of a coastal Nigerian city.[br]Methods: In a cross sectional survey, a sample comprising 1134 subjects (645 males and 489 females) representative of the entire population of Calabar aged 15[ndash]79 was studied. A multistage sampling method was applied and 50 house-holds from each of the four wards were selected using the table of random numbers, out of which eligible individuals aged between 15 years and 79 years from the 200 households selected were recruited.Using a modification of WHO STEPS instrument the information obtained included anthropometric indices (height in meters, weight in kilogram, waist circumference in centimetre, Hip circumference in centimetre). Anthropometric indices were expressed as mean (standard deviation). The normative values of indices of nutriture were determined using mean at 95% confidence interval, and the level of significance was taken at P [lt] 0.05.[br]Results: The mean (95% CI) values of BMI for males and females were 27.0kg/m2 (95% CI 26.5-27.2) and 28.5kg/m2 (95% CI 28.0-29.0) respectively, P [lt]0.01. The mean (95% CI) value of WC males was 91cm (95% CI 90.2-91.8), of females was 89.8cm (95% CI 88.8-90.8), P [lt] 0.01. The mean (95% CI) values of WHR males was 0.90 (95% CI 0.88-0.89) while that of females was 0.85 (95% CI 0.87-0.89) P [lt] 0.01. The mean (95% CI) values of height of males was 1.70m (1.70-1.72) and of females was 1.60m (1.64-1.65), P [lt]0.01.[br]Conclusion: The normative values of anthropometry in the study population are different from those from other parts of Nigeria and other parts of the world with lower BMI, higher WC and WHR.More studies on anthropometric indices, involving other regions of the country are needed, to aid in the establishment of nationwide specific cut-off values for Nigeria.[br][br]Nothing to Disclose: OEE, OEE, OAF, AEO 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 315 53 242 SAT-129 PO28-02 Saturday 181 2012


182 ENDO12L_SAT-130 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) IGF-1, IGFBP-1, IGFBP-3 and IGFBP-4 Levels and Their Associations to Cardiovascular Risk Factors in Obese Children before and after Weight Loss Thomas Reinehr, Joachim Woelfle, Nina Lass, Yvonne Uysal, Christian Roth Vestische Children[apos]Hospital, University of Witten/Herdecke, Datteln, Germany; University of Bonn, Bonn, Germany; University of Washington, Seattle, WA [bold]Background[/bold]: Insulin-like growth factor (IGF) -1 and its binding proteins (IGFBP-1,-3,-4) are postulated to be associated to obesity, insulin resistance and its associated comorbidities. However, studies in children are controversial and most importantly longitudinal studies are scarce.[br][bold]Methods: [/bold]We analyzed IGF-1, IGFBP-1, IGFBP-3, and IGFBP-4 in 60 obese children (mean age 10.8[plusmn]2.4, 52% female, mean BMI 28.2[plusmn]4.1kg/m[sup2]) at onset and end of a one-year lifestyle intervention. Anthropometrical markers, waist circumference (WC), systolic (SBP) and diastolic blood pressure (DBP), leptin, adiponectin, fasting glucose, insulin, triglycerides, uric acid, LDL- and HDL-cholesterol concentrations were determined at baseline and 1 year later.[br][bold]Results[/bold]: At baseline, IGF-1 correlated significantly to SBP (r=0.39), DBP (r=0.21), HDL-cholesterol (r=-0.30), and triglycerides (r=0.37). IGFBP-1 correlated significantly to BMI (r=-0.48), WC (r=-0.44), SBP (r=0.37), DBP (r=0.26), insulin (r=0.25), HOMA (r=-0.33), leptin (r=-0.37), and adiponectin (r=0.34). IGFBP-3 correlated significantly to WC (r=-0.39), SBP (r=-0.37), DBP (r=-0.30), HDL-cholesterol (r=-0.32), LDL-cholesterol (r=0.33), triglycerides (r=0.28), uric acid (r=0.25), insulin (r=0.25), HOMA (r=0.24), leptin (r=0.21), and adiponectin (r=-0.40). IGFBP-4 correlated significantly to BMI (r=0.34), WC (r=0.39), uric acid (r=0.39), insulin (r=0.32), HOMA (r=0.34), leptin (r=0.30), and adiponectin (r=-0.28).[br]In the course of 1y, the changes of IGF-1 were significantly related to changes of triglycerides (r=0.35) and uric acid (r=0.27). The changes of IGFBP-3 correlated significantly to changes of WC (r=0.29), SBP (r=0.29) and DBP (r=0.20). The changes of IGFBP-4 were significantly related to changes of BMI (r=0.21), uric acid (r=0.26), insulin (r=0.21), and HOMA (r=0.22).[br]In the 30 children with reduction of BMI (in mean -2.9kg/m[sup2]) all cardiovascular risk factors improved significantly, while IGF-1, IGFBP-3, and IGFBP-4 did not change significantly. In the 30 children with increase of BMI (in mean +1.3kg/m[sup2]), IGF-1 and IGFBP-4 increased significantly, IGFBP-1 decreased significantly, while IGFBP-3 and all cardiovascular risk factors remained stable.[br][bold]Conclusion[/bold]: The cross-sectional and longitudinal relationships between IGF-1 and its binding proteins to cardiovascular risk factors suggest a potential role of the IGF-1 axis in the pathogenesis of insulin resistance and its associated comorbidities.[br][br]Sources of Research Support: This work was supported by an investigator-initiated, unrestricted research grant from Merck Serono GmbH.[br][br]Nothing to Disclose: TR, JW, NL, YU, CR 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 10 53 243 SAT-130 PO28-02 Saturday 182 2012


183 ENDO12L_SAT-131 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Waist Circumference Is Superior to Weight and BMI in Predicting Sexual Symptoms, Voiding Symptoms and Psychosomatic Symptoms in Men with Late-Onset Hypogonadism Aksam Yassin, Youssef El Douaihy, Ridwan Shabsigh, Aiman Yassin, Farid Saad Segeberger Kliniken, Norderstedt, Germany; Dresden International University, Dresden, Germany; Gulf Medical University School of Medicine, Ajman, United Arab Emirates; Maimonides Medical Center, Brooklyn, NY; Klinikum Braunschweig, Braunschweig, Germany; Bayer Pharma AG, Berlin, Germany; Gulf Medical University School of Medicine, Ajman, United Arab Emirates [bold]Introduction[/bold]: We investigated whether three measurements of obesity: weight, BMI, and waist circumference correlate with the International Index of Erectile Function (IIEF), the Aging Males[apos] Symptoms questionnaire (AMS), and the International Prostate Symptom Score (IPSS).[br][bold]Materials and methods[/bold]: As of November 2004, 130 patients were enrolled in a prospective study on late onset hypogonadism (LOH defined as total testosterone [le] 3.5 ng/mL and symptoms). All men were treated with i.m. testosterone undecanoate (TU) 1000mg. The treatment was initiated on day 1, then 6 weeks thereafter and was continued at 3 months intervals. The mean time of treatment and follow up was 4.7 years. Baseline demographic data were recorded including hormones, lipids, and glucose. Blood samples were collected at every or every other visit. In addition patients were asked to fill in three standardized questionnaires IIEF, IPSS and AMS. The Sexual Health Inventory for Men (SHIM), a short version of the IIEF was used. SPSS was used to plot weight, BMI and waist circumference versus the recorded scores on the three different questionnaires. ANOVA analysis was used to look at any possible statistical significance.[br][bold]Results[/bold]: The cohort[apos]s means and standard deviations for the IIEF, AMS and IPSS scores were 7.3 ([plusmn]3.11), 54.63 ([plusmn]7.06) and 11.65 ([plusmn]4.41) respectively. The estimated curves and the ANOVA regression for continuous variables showed that waist circumference is inversely proportional to IIEF and directly proportional to AMS and IPSS with statistical significance. Weight was inversely proportional to IIEF, and directly proportional to IPSS both with statistical significance. Weight did not show enough linearity with AMS to give statistical significance. BMI and all three measurements of size had no proportionality.[br][bold]Conclusion[/bold]: Among weight, BMI and waist circumference, the latter is the best predictor of health related quality of life in men with LOH including sexual symptoms, voiding symptoms and other psychosomatic symptoms.[br][br]Sources of Research Support: Data entry has been supported by Bayer Pharma AG.[br][br]Disclosures: AY: Speaker, Bayer Schering Pharma. RS: Speaker, Lilly USA, LLC. FS: Employee, Bayer Schering Pharma. Nothing to Disclose: YED, AY 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 469 53 244 SAT-131 PO28-02 Saturday 183 2012


184 ENDO12L_SAT-132 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Depressive Symptoms in Adolescents Referred for Obesity Management: What Is the Relationship with Insulin Resistance? Tamara S Hannon, Ann M Lagges, Jordan N Huber, Aaron E Carroll, Sandeep K Gupta Indiana University School of Medicine, Indianapolis, IN; Indiana University School of Medicine, Indianapolis, IN; Indiana University School of Medicine, Indianapolis, IN; Indiana University School of Medicine, Indianapolis, IN Studies in adults have shown depression and psychosocial stress to be associated with metabolic syndrome and type 2 diabetes; however, few studies in pediatrics have examined these relationships. We aimed to determine whether depressive symptoms are related to measures of glucose tolerance (fasting glucose, glycosylated hemoglobin [A1C]) and insulin resistance (fasting insulin, HOMA-IR) in obese adolescents referred to a hospital-based obesity program (Pediatric OverWeight Education and Research, POWER). Methods: The Children[apos]s Depression Inventory[sup]1[/sup] (CDI) was used to examine self-reported depressive symptoms (anhedonia, self-esteem, ineffectiveness, interpersonal problems, negative mood) in 284 children referred for treatment of obesity. Of the 284 patients, 260 had complete CDI and laboratory data (64% female; 54% white, 26% black, 5% Hispanic, 3% other race; mean age 12.9[plusmn]2.7 y, BMI 37.3[plusmn]8.1 kg/m[sup]2[/sup]). Results: BMI was associated with insulin resistance (r=0.37, p[lt]0.001 for fasting insulin; r=0.32, p[lt]0.001 for HOMA-IR) but not with glucose tolerance (r=[minus]0.09, p=0.20 for fasting glucose; r=0.08, p=0.21 for A1C). CDI t-scores (mean=49; range 31-100) were not indicative of increased prevalence of depression in this population and were not associated with BMI (r=0.04, p=0.54). CDI t-scores were, however, positively correlated with insulin resistance (r=0.15, p=0.02 for fasting insulin and r=0.16, p=0.11 for HOMA-IR), but not with glucose tolerance. These relationships were largely due to associations with interpersonal problems (difficulties interacting with people, social avoidance and isolation) (r=0.16, p=0.01 for fasting insulin; r=0.13, p=0.03 for fasting glucose; r=0.16, p=0.01 for HOMA-IR). Multiple linear regression analysis was performed controlling for sex, race, age, and BMI to determine the degree to which interpersonal problems predicted insulin resistance. The total variance explained by the model was 20%, F=10.4, p[lt]0.001. The interpersonal problem t-score contributed significantly to the model, R[sup]2[/sup] change=0.11, F change=17.5, p[lt]0.001. Conclusions: Among adolescents referred for obesity treatment, endorsement of interpersonal problems is associated with insulin resistance. The relationships between interpersonal problems and risk factors for the development of diabetes in youth should be investigated further to evaluate the mechanisms and determine if early recognition and therapy could lessen the risk of type 2 diabetes.[br][br](1) Kovacs, M. (2003) Children[apos]s Depression Inventory. Western Psychological Services, Los Angeles, CA.[br][br]Nothing to Disclose: TSH, AML, JNH, AEC, SKG 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 943 53 245 SAT-132 PO28-02 Saturday 184 2012


185 ENDO12L_SAT-133 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Effects of Different Bariatric Surgery Procedures on Oligoelements and Antioxidants in Human Obesity Antonio Mancini, Roberto Festa, Sebastiano Raimondo, Chantal Di Segni, Serena Marchitelli, Andrea Silvestrini, Elisabetta Meucci, Alfredo Pontecorvi, Roberto Maria Tacchino Catholic University of the Sacred Heart, Rome, Italy; Polytechnic University of the Marche, Ancona, Italy; Catholic University of the Sacred Heart, Rome, Italy; Catholic University of the Sacred Heart, Rome, Italy In previous works we have demonstrated that surgically-induced weight loss, obtained by biliopancreatic diversion (BPD), is associated with an increase in insulin sensitivity, but a decrease in some molecules, such as Coenzyme Q[sub]10[/sub], that act as powerful antioxidant system. The mechanism of this phenomenon (weight loss per se, lipid malabsorption, metabolic or hormonal variations after surgery) are not yet clear. Since antioxidants are very important in preventing metabolic complications of obesity, in order to delve into the physiopathology of this condition we have studied 19 subjects (6 males and 13 females, age 41.4 [plusmn] 11.0 years, BMI 48.5 [plusmn] 7.9 Kg/m[sup]2[/sup]), before and at a long time (12-18 months) from surgery, treated by different surgical procedures: BPD, gastric bypass (GB), gastric minibypass (GM), evaluating oligoelements and antioxidants in plasma.[br]After surgery they reached a mean % decrease in body weight of 41.7 [plusmn] 0.1, without differences with the different surgical techniques. They were treated with the same schedule of integrative therapy. We studied metabolic parameters (glycemia, insulinemia with the calculation of HOMA insulin-resistance index, total- LDL- HDL-cholesterol, triglycerides, uric acid, albumin, transaminases), hormones (fT3, fT4, TSH, IGF-1, cortisol, ACTH) and serum values of copper, zinc and Vitamin E. Total antioxidant capacity was evaluated by the system metmyoglobin-H[sub]2[/sub]O[sub]2[/sub], as a source of radicals, and the chromogen ABTS, whose radical cation is spectroscopically revealed: the latency time (sec) in the appearance of radical species is a measurement of the antioxidant content of the sample. Analysis of variance was used for statistical evaluation.[br]We observed significant differences (p[lt]0.05) in mean variation of: HOMA (-80[plusmn]11% in BPD, -55[plusmn]31% in GB, -84[plusmn]15% in GM), HDL-cholesterol (+3[plusmn]21%, +26[plusmn]30%, +32[plusmn]17%, respectively), IGF-1 (-41[plusmn]21%, +9[plusmn]37%, -50[plusmn]32%, respectively). LAG values did not significantly differed in the three groups (-12[plusmn]14%, -18[plusmn]14%, -13[plusmn]21%, respectively).[br]These data, although preliminary, suggest a different pattern of response to bariatric surgery, irrespective of weight loss, with a more favourable response in gastric bypass and minibypass in comparison to BPD. They also suggest that previously demonstrated loss of antioxidant could be due to lipid malabsorption as main mechanism.[br][br]Nothing to Disclose: AM, RF, SR, CDS, SM, AS, EM, AP, RMT 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 946 53 246 SAT-133 PO28-02 Saturday 185 2012


186 ENDO12L_SAT-134 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) How Does Treating Sleep-Disordered Breathing Affect the Lipid Profile in Obese Children and Adolescents? Zarlasht Amini, Seema Kumar, Suresh Kotagal, Robin Lloyd, Christine Lohse Mayo College of Medicine, Rochester, MN; Mayo College of Medicine, Rochester, MN; Mayo College of Medicine, Rochester, MN [bold]Background: [/bold]Childhood obesity is frequently associated with hyperlipidemia, hypertension, insulin resistance and cardiovascular risk. Obstructive Sleep Apnea (OSA) seen commonly in obese children appears to contribute to greater risk of these comorbidities independent of obesity. However the impact of treatment of OSA on metabolic parameters in children is not known. The objective of the study was to assess the impact of treating OSA with Positive Airway Pressure breathing (PAP) or Adenotonsillectomy (T[amp]A) on the fasting glucose and lipid profile in obese children and adolescents.[br][bold]Methods: [/bold]Retrospective medical record review of subjects aged 2-18 years with BMI[gt] 85[sup]th[/sup] percentile for age and gender who underwent polysomnography at the Center for Sleep Medicine for evaluation of Sleep Disordered Breathing (SDB) between January 1[sup]st[/sup], 2000 and December 31[sup]st[/sup], 2010. Associations among variables were evaluated using Spearman rank correlation coefficients and changes in variables from baseline following treatment were evaluated using paired t and Wilcoxon signed rank tests.[br][bold]Results: [/bold]Forty seven patients with OSA (defined as Apnea Hypopnea Index (AHI) of [ge] 1 per hour on overnight polysomnography) had a fasting lipid panel consisting of total cholesterol, triglycerides, high-density lipoprotein (HDL) cholesterol and calculated low density lipoprotein (LDL) cholesterol measured within three months prior to the diagnosis of OSA. Forty four patients had a fasting glucose measured within three months prior to the diagnosis of OSA. OSA was treated with PAP or T[amp]A. Twenty four subjects had a repeat lipid panel and 18 subjects had a repeat fasting glucose drawn at a mean of 19 months (median 12.5; range 3 [ndash] 46) following treatment of OSA.[br][bold]Results:[/bold] AHI was significantly positively correlated with fasting glucose (r= 0.45, p=0.002) and with systolic blood pressure (r=0.24, p=0.023). There was a statistically significant decrease in total cholesterol and in LDL cholesterol following treatment of OSA (median change -12.5 mg/dL; p[lt]0.001 and -4.5 mg/dL; p=0.021, respectively). No significant change in HDL-cholesterol, triglycerides or fasting glucose was noted.[br][bold]Conclusion[/bold]: Treatment of OSA in obese children is associated with improvement in levels of total cholesterol and LDL-cholesterol levels. Screening for OSA in obese children and its treatment should be an integral component of programs aimed at decreasing cardiovascular risk in these children.[br][br]Nothing to Disclose: ZA, SK, SK, RL, CL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 958 53 247 SAT-134 PO28-02 Saturday 186 2012


187 ENDO12L_SAT-135 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Effect of CPAP on Weight in Patients with Sleep Apnea: A Meta-Analysis Zunaira Z Chaudhry, Ali Hamza, Carmella Evans-Molina, Rashid Nadeem, Janice Gilden, Saleha Z Chaudhry, Abdul Rehman Rishi, Muhammad A Rishi Indiana University, Indianapolis, IN; Captain James A Lovell Federal Health Care Center, North Chicago, IL; Mercy Hospital Medical Center, Chicago, IL; MacNeal Hospital Medical Center, Berwyn, IL Background: Obesity is a common cause of obstructive sleep apnea syndrome (OSAS). Impact of treatment of sleep apnea with continuous positive airway pressure (CPAP) on body mass index (BMI) is not well established with various studies producing differing results.[br]Methods: We conducted a meta-analysis of controlled trials to evaluate the effects of CPAP on BMI, reported as either a primary or secondary end point, among patients with OSAS. Studies were retrieved by searching Medline (January 1980 to Nov 2011), the Cochrane Database of Systematic Reviews, conference abstracts, and bibliographies of review and original articles. From 345 relevant reports, 10 randomized control clinical trials were selected that compared CPAP to control among participants with OSAS, had a minimum treatment duration of 4 weeks, and reported BMI changes during the intervention or control period. Data on sample size, participant characteristics, study design, intervention methods, duration, and treatment results were independently abstracted by 2 investigators using a standardized protocol and examined using a fixed-effects model.[br]Results: Mean net change in BMI in patients with OSAS treated with CPAP compared with control was 0.78 kg/m[sup2] (95% confidence interval; CI 0.65-0.91).[br]Conclusion: This meta analysis indicates that CPAP use in patients with OSAS may lead to a modest increase in BMI. Further studies are needed to elucidate its cause.[br][br]Nothing to Disclose: ZZC, AH, CE-M, RN, JG, SZC, ARR, MAR 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2358 53 248 SAT-135 PO28-02 Saturday 187 2012


188 ENDO12L_SAT-136 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Abdominal Adiposity and Obstructive Airway Disease: Testing Insulin Resistance and Sleep-Disordered Breathing Mechanisms Matthew Timothy Haren, Gary Misan, Tracey-Jayne Paterson, Richard E Ruffin, Janet F Grant, Jonathan D Buckley, Peter RC Howe, Jonathan Newbury, Anne W Taylor, Robyn A McDermott University of South Australia, Adelaide, Australia; University of South Australia and The University of Adelaide, Whyalla, Australia; University of South Australia, Whyalla, Australia; University of Adelaide, Adelaide, Australia; University of South Australia, Adelaide, Australia [bold]Background:[/bold] This study examined associations of abdominal adiposity with lung function, asthma symptoms and current doctor-diagnosed asthma, and mediation by insulin resistance (IR) and sleep disordered breathing (SDB).[br][bold]Methods:[/bold] A random sample of 2500 households was drawn from the community of Whyalla, South Australia (The Whyalla Intergenerational Study of Health, WISH February 2008 - July 2009). Seven-hundred twenty-two randomly selected adults (18 years and over) completed clinical protocols (32.2% response rate). Lung function was measured by spirometry. Post-bronchodilator FEV[sub]1[/sub]/FVC was used to measure airway obstruction, and reversibility of FEV[sub]1[/sub] was calculated. Current asthma was defined by self-reported doctor-diagnosis and evidence of currently active asthma. Symptom scores for asthma (CASS) and SDB were calculated. Intra-abdominal fat (IAF) was estimated using dual-energy x-ray absorptiometry (DXA). IR (HOMA2-IR) was calculated from fasting glucose and insulin concentrations.[br][bold]Results:[/bold] The prevalence of current doctor-diagnosed asthma was 19.9% (95% CI 16.7, 23.5%). The ratio of observed to expected cases given the age and sex distribution of the population was 2.4 (95%CI 2.1, 2.9). IAF was not associated with current doctor-diagnosed asthma, FEV[sub]1[/sub]/FVC or FEV[sub]1[/sub] reversibility in men or women but was positively associated with CASS independent of IR and SDB in women. A 1% increase in IAF was associated with decreases of 12mL and 20mL in FEV[sub]1[/sub] and FVC respectively in men, and 4mL and 7mL respectively in women. SDB mediated 12% and 26% of these associations respectively in men only.[br][bold]Conclusions:[/bold] In this population with an excess of doctor-diagnosed asthma, IAF was not a major factor in airway obstruction or doctor-diagnosed asthma, although women with higher IAF perceived more severe asthma symptoms which did not correlate with lower FEV[sub]1[/sub]. Higher IAF was significantly associated with lower FEV[sub]1[/sub] and FVC and in men SDB mechanisms may contribute up to one quarter of this association.[br][br]Sources of Research Support: The South Australian Premiers Science Research Fund; University Departments of Rural Health Program (Australian Government Department of Health and Ageing); National Health and Medical Research Council (NHMRC) of Australia Post-doctoral Training Fellowship (Public Health) 511345 awarded to MTH; NHMRC Public Health Practitioner Fellowship awarded to RAM.[br][br]Nothing to Disclose: MTH, GM, T-JP, RER, JFG, JDB, PRCH, JN, AWT, RAM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1315 53 249 SAT-136 PO28-02 Saturday 188 2012


189 ENDO12L_SAT-137 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) The Ability of Body Mass Index To Predict Abnormal Waist Circumference: ROC Analysis Marcio W Lauria, Livia P Moreira, George M Coelho, Silvia N Freitas, Raimundo Marques, Adauto V Ramos, Maria Marta S Soares UFMG, Belo Horizonte, Brazil; Hospital Felicio Rocho, Belo Horizonte, Brazil; UFOP, Ouro Preto, Brazil Background: Body mass index (BMI) is widely utilized as an anthropometric estimate of general adiposity; however, differences in body composition and body fat distribution limit its usefulness. In recognition that visceral fat accumulation increases the risk for metabolic disease, waist circumference (WC) was promoted as an alternative surrogate measure of obesity. However, WC is subject to significant inter-examiner variation.[br]Objectives: To correlate BMI and WC measures in a group of Brazilian adults searching for the most accurate BMI values in predicting abnormal WC.[br]Methods: BMI and WC were measured in 1,184 volunteers (45.6[plusmn]17.3 yrs; 69% women) according to standard procedures. Abnormal WC was defined as [gt] 88 cm in women and [gt] 102 cm in men. Statistics analysis was done using Pearson[acute]s correlation coefficient and receiver operating characteristic (ROC) curve.[br]Results: Anthropometric measures: BMI- women=27.1[plusmn]5.9 kg/m2; WC-women=88.9[plusmn]15.1 cm; BMI-men=25.4[plusmn]4.8 kg/m2; WC-men=89.9[plusmn]14.4 cm. BMI had a strong correlation with WC (women: r=0.87, p[lt]0.0001, area under ROC curve=0.93[plusmn]0.01; men: r=0.89, p[lt]0.0001, area under ROC curve=0.94[plusmn]0.01). The most accurate BMI cut offs for abnormal WC were 26.8kg/m2 for women (sensibility=82%; specificity=89%) and 27.1 kg/m2 for men (sensibility=96%; specificity=81%).[br]Conclusion: Patients with BMI[gt] 27 kg/m2 usually have abnormal WC. In this group, WC measurement may reasonably be waived. For patients with lower BMI, WC measurement still remains informative.[br][br]Nothing to Disclose: MWL, LPM, GMC, SNF, RM, AVR, MMSS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 995 53 250 SAT-137 PO28-02 Saturday 189 2012


190 ENDO12L_SAT-138 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Association between Mitochondrial DNA Content, Mitochondrial Function, and Childhood Obesity and Insulin Resistance Zohreh Shoar, Michael J Goldenthal, Francesco De Luca, Elizabeth A Suarez St Christopher[apos]s Hospital for Children, Drexel University College of Medicine, Philadelphia, PA; St Christopher[apos]s Hospital for Children, Drexel University College of Medicine, Philadelphia, PA BACKGROUND[br]In adults, low levels of mitochondrial DNA (mtDNA) and mitochondrial enzyme activity have been associated with obesity; in addition, altered mitochondrial function is thought to play a causal role in the metabolic syndrome, as a defective oxidative phosphorylation may be involved in visceral fat gain and development of insulin resistance. However, there is little evidence on the correlation between mtDNA content, mitochondrial function, and adiposity in children.[br]OBJECTIVES[br]Our goal was to compare mtDNA content and mitochondrial enzyme activity between obese and non-obese children and to correlate mtDNA content with BMI.[br]DESIGN/METHODS[br]Obese (BMI[ge]95% for age and sex) and non-obese children, ages 2-18 yrs were recruited. Anthropometric measurements were taken and BMI Z-score and percentile were calculated. Buccal swabs for mitochondrial enzymes (Complex I and Citrate Synthase) were collected and their activity was measured by using a dipstick methodology combined with spectrophotometric analysis. In obese children only, serum levels of fasting glucose and insulin were measured, HOMA-IR was calculated and nuclear/mtDNA from mononuclear cells was isolated and quantitated by real-time PCR.[br]RESULTS[br]54 obese (30 girls, 24 boys, mean age of 12.3 [plusmn] 3.8 yrs) and 25 control (15 girls, 10 boys, mean age of 10.9 [plusmn] 4.6 yrs) children were recruited. In obese children, complex I and Citrate Synthase levels were lower than those of controls (79.5[plusmn]49.6 unit/min/mg protein vs. 92.1[plusmn]43.1and 11.3[plusmn]4.9 nmol/min/mg protein vs.12.6[plusmn]4.3, respectively); however, this difference did not reach statistical significance (p=0.29 and 0.3 respectively). We found a positive correlation between BMI z-score and the ratio of mt/nuclear DNA (Spearman rho=0.43, p=0.035). No correlation was found between mtDNA and HOMA-IR. Lastly, children with the BMI in the highest quartile exhibited a higher mt/nuclear DNA ratio (1.9[plusmn]1.0) compared to those with a BMI in the lowest quartile (0.7[plusmn]0.7), but again the difference was not statistically significant.[br]CONCLUSIONS[br]Our preliminary findings in obese children suggest that mtDNA content in mononuclear cells is positively correlated with degree of adiposity. Although not statistically significant, the difference of the selected mitochondrial enzyme activity between obese and control children suggests a decreased oxidative phosphorylation in the former. Further studies in a larger population sample are needed to confirm these findings.[br][br]Nothing to Disclose: ZS, MJG, FDL, EAS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1153 53 251 SAT-138 PO28-02 Saturday 190 2012


191 ENDO12L_SAT-139 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Increased Circulating Placental Growth Factor (PLGF) Concentrations in Children and Adolescents with the Metabolic Syndrome Panagiota Pervanidou, Athanassios Akalestos, Despoina Bastaki, George P Chrousos Athens University Medical School, Athens, Greece Background: Childhood obesity is associated with an increased risk for early onset endothelial dysfunction and atherosclerosis. This association is mediated by pathogenetic mechanisms related to the development of the Metabolic Syndrome (MetS) and associated changes in endothelial and inflammation functions. Placental growth factor (PLGF), a member of the vascular endothelial growth factor family, plays an important role in a range of physiologic and pathologic processes,, including stimulating angiogenesis and atherogenic migration of monocytes/macrophages into the arterial wall, and contributions to preeclampsia, cardiovascular disease and tumor progression,. The aim of this study was to investigate differences in PLGF concentrations between children with obesity/metabolic syndrome and age-matched normal weight controls.[br]Materials and Methods: Forty-four (34 males) obese and 33 lean (normal body mass index, BMI) children were assessed at the Childhood Obesity Clinic of our department. Mean age was 11.65 [plusmn]2.01 years. Obesity and metabolic syndrome were defined using the IOTF and IDF criteria, respectively. PLGF was measured using electrochemiluminescence.[br]Results: Obese children (N=44) had similar PLGF concentrations [mean value 11.82 [plusmn]2.97 pg/ml] to those of children with normal BMI [mean value 11.87[plusmn] 3.09pg/ml], while no significant correlations were found between PLGF and BMI z-score. Among the obese children, those with the full MetS (8 children, 7 males) had significantly higher (p=0.014) PLGF [14.47[plusmn]3.47 pg/ml] concentrations than those without the MetS (11.32 [plusmn] 2.63 pg/ml).[br]Conclusions: Although no apparent differences were found in PLGF concentrations between obese and lean children, obese children with the MetS had higher concentrations of PLGF than obese children without the MetS. These findings suggest PLGF may participate in the process of atherogenesis in obese children and adolescents with the MetS.[br][br]Nothing to Disclose: PP, AA, DB, GPC 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2016 53 252 SAT-139 PO28-02 Saturday 191 2012


192 ENDO12L_SAT-140 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Using Structural Equation Modeling To Assess the Associations between Obesity and Pancreatic Beta-Cell Function in Adolescents Alice PS Kong, Kai Chow Choi, Ronald CW Ma, Juliana CN Chan The Chinese University of Hong Kong, Hong Kong, Hong Kong; The Chinese University of Hong Kong, Hong Kong, Hong Kong [bold]Aim[/bold][br]Obesity, insulin resistance and lipotoxicity may contribute to the decline in pancreatic beta-cell function (BCF), which in turn leads to glucose intolerance. Here, we aim to explore the complex inter-relationships between these factors in adolescents using structural equation modeling (SEM).[br][bold]Methods[/bold][br]We identified 225 Chinese adolescents aged 12-19 year-old from a territory-wide, population recruited cohort surveyed in Hong Kong. Anthropometric measurements, fasting blood sampling for lipid profile, insulin (to estimate insulin resistant state by calculating homeostasis model assessment, HOMA-IR) and oral glucose tolerance test (OGTT) were performed. The SEM analyses were performed using LISREL 8.8 (Scientific Software International, Inc.) and the parameters were estimated by maximum likelihood method. The following fit indices were chosen to assess the goodness-of-fit of the path: (1) Chi-square statistic to degree of freedom ratio (c[sup]2[/sup]/[italic]df[/italic]), (2) standardized root mean square residual (SRMR), (3) root mean square error of approximation (RMSEA), (4) adjusted goodness-of-fit index (AGFI), (5) comparative fit index (CFI) and (6) non-normed fit index (NNFI).[br][bold]Results[/bold][br]Mean age of the study cohort was 15.0(+/-1.9) years. 45.8% were boys. 65.3% had normal glucose tolerance. The c2 of the final model is 53.5 with df=25, p[lt]0.001. The goodness-of-fit indices (c2/df =2.1, RMSEA = 0.07, SRMR=0.04, AGFI = 0.94, CFI = 0.98, and NNFI=0.96) indicate that the final model is a reasonably good fit to the data. All the path coefficients are significant (p [lt] 0.05). The model explains for 28%, 29%, 44% and 8% of the variance of lipid profile, HOMA-IR, pancreatic beta cell function and two-hour plasma glucose from OGTT respectively.[br][bold]Conclusion[/bold][br]Obesity leads to decline in BCF through interactions with lipids and insulin resistance in adolescents, culminating to the development of glucose intolerance.[br][br]Sources of Research Support: This study was supported by funding from the General Research Fund from the Research Grants Council (CUHK 4055/01M and 467410), Hong Kong.[br][br]Nothing to Disclose: APSK, KCC, RCWM, JCNC 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1193 53 253 SAT-140 PO28-02 Saturday 192 2012


193 ENDO12L_SAT-141 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Effect of Body Fat Mass on Fasting and Glucose-Stimulated Insulin Secretory Dynamics in Healthy Men Ali Iranmanesh, Donna M Lawson, Johannes Veldhuis Salem VA Medical Center, Salem, VA; Mayo Clinic, Rochester, MN Background: Insulin secretion in fasting and fed states is episodic, with circulating concentrations mostly contributed by the pulsatile events.[br]Objective: To assess the effect of total body fat and body fat distribution on circulating insulin levels and insulin secretory parameters in normal men during short-term fasting and after oral glucose intake.[br]Subjects: 57 men ages 19-78 yrs and BMIs 20-39Kg/m[sup]2[/sup][br]Study design: Each man studied on 2 separate occasions, and after an overnight fast.[br]Intervention: Ingestion of either 75 grams of dextrose solution or equal volume of water. Sessions were 6.5-hr long, starting at of 0800-0900 hr. Blood was collected at 10-min intervals for glucose and insulin measurements. DXA and single-slice CT of abdomen were used for the assessment of body composition and abdominal fat area.[br]Results: Mean 6.5 hr serum glucose (g) and insulin (i) concentrations were positively correlated (R/P) with % body fat both on water (g: 0.42/0.001; i: 0.42/0.001), and dextrose (g: 0.49/0.0001; i: 0.43/0.0008) days. Deconvolution analysis of insulin time series identified combined contributions of basal (b) and pulsatile (p) events to total (t) insulin secretion, with positive effects of % body fat in fasting (b: 0.42/0.001, p: 0.27/0.04, t: 0.43/0.0009) and fed (0.35/0.008, 0.42/0.001, 0.43/0.0007) states. While not a determining factor in fasted state, body fat had a positive effect on frequency of insulin secretory bursts after dextrose intake. Backward stepwise multiple regression statistics with % body fat, visceral abdominal fat (VAF), and subcutaneous abdominal fat (SAF) as independent variables [underline]identified SAF as a stronger predictor (R/P)[/underline] of circulating insulin concentrations, and corresponding basal and total insulin release both on water (0.46/0.0003, 0.44/0.0006, 0.46/0.0003) and oral dextrose (0.46/0.003, 0.40/0.002, 0.47/0.0003) days. Pulsatile insulin secretion maintained a higher correlation with SAF during fasting (0.38/0.003), but with % body fat (0.42/0.001) after dextrose intake.[br]Summary: Excess body fat in healthy men augments pulsatile and basal insulin release both during short-term fasting and after glucose ingestion, with resulting increases in circulating insulin concentrations. The latter is further augmented by increased number of insulin secretory bursts, but only in the fed state. By virtue of association, SAF rather than VAF appears to be a stronger factor in fat-induced alterations in insulin/glucose homeostasis in men.[br][br]Nothing to Disclose: AI, DML, JV 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1722 53 254 SAT-141 PO28-02 Saturday 193 2012


194 ENDO12L_SAT-142 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) Endurance Training Increases Serum Uncarboxylated Osteocalcin Level and Decreases HOMA-IR Via Reduction of Serum Leptin in Obese Young Patients Eun Jin Kwon, Yu Sik Kim, Shinae Kang, Dong Wook Yeo, Min Kyung Kim, Jiyoon Ha, Youngmi Lee, Jiwoon Kim, Tae Woong No, Jong Suk Park, Kyung Rae Kim, Sang Hoon Suh, Chul Woo Ahn Yonsei University Colleage of Medicine, Seoul, Republic of Korea Recently, serum osteocalcin (OC) secreted from osteoblast has been reported to enhance glycemic control. On the other hand, moderate increase of leptin has been known to stimulate osteoblastic functions. Therefore, it can be assumed that leptin may regulate glucose homeostasis by stimulating the secretion of OC. Between two forms of OC (uncarboxylated OC (unOC) and carboxylated OC (cOC)), unOC is the predominant form in the circulation. Interestingly, leptin is known to increase the expression of osteotesticular protein tyrosine phosphatase, which in turn is known to decrease OC secretion by inducing OC carboxylation. This suggests that reduction of leptin may result in increased osteoblastic activity, OC production/secretion, and consequently, glycemic control. This study investigated whether endurance training may regulate circulating levels of ucOC and parameters of glycemic control via leptin reduction. 39 class I obese, normoglycemic young Korean males were randomly assigned to control (C, n=10) or exercise group (Ex, n=29). The subjects in Ex group underwent 8 week-supervised-endurance exercise training (2400Kcal/week). Weight, BMI, waist circumference, % body fat, leptin, fasting plasma insulin, and HOMA-IR decreased significantly. Whereas ucOC and total OC increased in Ex, without significant change of cOC. There was also negative correlation between the change in leptin and ucOC. Total adiponectin decreased but the ratio of high molecular weight adiponectin to total adiponectin increased by 6.6 fold. Fasting plasma glucose and AUC from oral glucose tolerance test showed no significant change. Bone alkaline phosphatase and bone mineral density were unaffected by the exercise training. These results suggest that endurance training may contribute to increased circulating OC and decreased HOMA-IR without any significant change in bone metabolism. Considering that this study was limited to the normoglycemic obese individuals, further study in the prediabetic and diabetic individuals may be warranted.[br][br]Nothing to Disclose: EJK, YSK, SK, DWY, MKK, JH, YL, JK, TWN, JSP, KRK, SHS, CWA 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1350 53 255 SAT-142 PO28-02 Saturday 194 2012


195 ENDO12L_SAT-143 POSTER SESSION: Obesity Topics II (1:30 PM-3:30 PM) The Association between 25(OH)D and Adiposity Indices among Preadolescent Children (Aged 7[ndash]9 Years). Hye Ah Lee, Young Ju Kim, Eun Hee Ha, Hwayoung Lee, Hye Sun Gwak, Eun Ae Park, Sujin Cho, Se Young Oh, Eun Hee Ha, Hyesook Park, Hae Soon Kim School of Medicine, Ewha Womans University, Seoul, Republic of Korea; School of Medicine, Ewha Womans University, Seoul, Republic of Korea; School of Medicine, Ewha Womans University, Seoul, Republic of Korea; School of Medicine, Ewha Womans University, Seoul, Republic of Korea; School of Medicine, Ewha Womans University, Seoul, Republic of Korea; Kyung Hee University, Seoul, Republic of Korea [bold]Background and aims:[/bold] The lower 25-Hydroxyvitamin D(25(OH)D) in obese child is being considered as a important public health problem, but there is still under debate for its relationship. Therefore, we aimed to investigate the association between 25(OH)D and adiposity indices [body mass index(BMI), body fat mass(BFM), percent body fat(PBF), waist circumference(WC), upper arm circumference, and skin-fold thickness] among preadolescent child (aged 7-9 years).[br][bold]Methods: [/bold]We followed up subjects at 2011 aged 7 to 9 child, who were part of Ewha Birth [amp] Growth Cohort study, which is a prospective cohort established 2001-2006. The 25(OH)D concentration in blood were measured using Radio immunoassay. We assessed the magnitude of association with vitamin D level and adiposity indices using multivariate regression analysis adjusted for sex, age, birth weight and calories.[br][bold]Results: [/bold]38(17.2%) of the total 221 child were deficiency ([lt]20ng/mL) for vitamin D level, and its prevalence was more higher in boys (25.2%) than girls (8.1%). 25(OH)D level was negatively associated with BMI([italic][beta][/italic]=-0.08, [italic]p[/italic]=0.03), BFM([italic][beta][/italic]=-0.10, [italic]p[/italic]=0.04), and WC([italic][beta][/italic]=-0.24, [italic]p[/italic]=0.02) in any given for sex, age, birth weight and calories, but was not associated with PBF, upper arm circumference. Regarding the risk of overweight ([ge]BMI 85%tile), those who were in lower 25(OH)D group had significantly higher risk of overweight than sufficient group ([italic]p[/italic][sub]trend[/sub]=0.01).[br][bold]Conclusions: [/bold]The lower 25(OH)D level is associated with risk of adiposity indices in general preadolescent child. This study may contribute to add the evidence of an association between Vitamin D level and adiposity in general population.[br][br]Sources of Research Support: Acknowledgements: This work was supported by National Research Foundation of Korea Grant funded by the Korean Government (2010-0026225).[br][br]Nothing to Disclose: HAL, YJK, EHH, HL, HSG, EAP, SC, SYO, EHH, HP, HSK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1382 53 256 SAT-143 PO28-02 Saturday 195 2012


196 ENDO12L_SAT-154 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) GAD65 and IA-2 Autoantibodies Identify a Population of Type 2 Diabetes Patients with a Significant Impairment in [beta]-Cell Function Angela R Subauste, Anne Chang, Cindy Plunkett, Suman Tandra, Massimo Pietropaolo University of Michigan, Ann Arbor, MI Type 2 diabetes mellitus (T2DM) represents a group of heterogeneous metabolic diseases, ranging from abnormalities related to glucose-stimulated insulin secretion, insulin resistance and autoimmune abnormalities of pancreatic [beta]-cells. The objective of this study was to evaluate indexes of insulin secretion and visceral adiposity in clinically diagnosed T2DM patients with and without the presence of GAD65 and IA-2 autoantibodies (Ab). A total of 18 subjects with T2DM diagnosed over the age of 40 participated in this study: 10 GAD65 and IA-2 Ab negative and 8 GAD65 and/or IA-2 Ab positive. These patients were subdivided in two groups: Ab positive: insulin and/or metformin and Ab negative: insulin and/or metformin. None of these individuals had family history significant for T1DM.These subjects underwent an arginine challenge test to asses [beta]-cell function and a DEXA scan to asses body composition. Both groups were matched for age and sex. No statistically significant differences were found between Ab positive vs Ab negative in terms of age (years) (54[plusmn]3 vs 57[plusmn]3), duration of T2DM (years) (11.6[plusmn]2.2 vs 14.7[plusmn]2.0), BMI (33[plusmn]3 vs 30[plusmn]2) or HgbA1c (8.2[plusmn]0.6 vs 7,5[plusmn]0.5). Both groups had similar body composition with DEXA central fat (g) estimated as 3055 [plusmn]299 for GAD65/IA-2 positive and as 3045[plusmn]671 for GAD65/IA-2 negative. DEXA free fat mass (kg) was estimated at 54[plusmn]5 and 54[plusmn]4 respectively. Ab positive T2DM patients exhibited severely impaired [beta]-cell function as demonstrated by lower AIRmax (33 vs. 197 p=0.0431) compared to the Ab negative. In conclusion, GAD65 and IA-2 Ab identify a pathogenetically distinct subpopulation of T2DM patients with similar body mass composition and seemingly immune-mediated injury of pancreatic [beta]-cells.[br][br]Sources of Research Support: NIH grant DK056200 awarded to MP.[br][br]Nothing to Disclose: ARS, AC, CP, ST, MP 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2150 54 257 SAT-154 PO11-02 Saturday 196 2012


197 ENDO12L_SAT-155 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Steps toward Adopting Universal Gestational Diabetes Mellitus Criteria: A Review of GDM Practice at the Centre Hospitalier de l[apos]Universite de Montreal (CHUM) Thi Hoang Lan Nguyen, Elisabeth Codsi, Ariane Godbout Centre de Recherche du Centre Hospitalier de l[apos]Universit[eacute] de Montr[eacute]al (CRCHUM), Montreal, Canada; Centre Hospitalier de l[apos]Universit[eacute] de Montr[eacute]al (CHUM), Montreal, Canada Background: New recommendations for gestational diabetes mellitus (GDM) have been recently proposed (1). Review of local rates of adverse pregnancy outcome was warranted before revision of GDM diagnostic criteria in our center. Objective: To establish local demographic profile, review current care provided and analyze the impact of new diagnostic criteria on our population. Methods: An observational retrospective study was carried out among pregnant women with GDM diagnosed at the CHUM between 2005 and 2009. GDM was diagnosed if at least one of the two values on 75g oral glucose tolerance test was above local criteria: fasting plasma glucose [ge]5.0 or 2-hr post [ge]7.8mmol/L. Rates of adverse pregnancy outcomes were established for macrosomia, defined as birth weight (BW) [ge]4kg, [ge]4,5kg and as large for gestational age (BW [gt]90th percentile for gestational age); for intra-uterine growth restriction (IUGR), defined as low BW ([lt]2,5 kg) and as small for gestational age (BW [lt]10th percentile for gestational age); and for caesarean C-section (CS). Results: A total of 1354 pregnant women were included, representing 1407 pregnancies and 1437 newborns. Women mean age and BMI were 31.9yr and 26.6 kg/m2. Overall, 44.2% were Caucasians and reported family history of diabetes, past GDM or previous macrosomia represented respectively 36.5, 19.1 and 10.0% of subjects. All patients were followed by a nutritionist and 62.9% necessitated insulin therapy. For newborns, mean term and BW were 39wks and 3.4kg. Rates of macrosomia defined as BW[ge]4kg, [ge]4.5kg and LGA were respectively 9.3, 1.3 and 9.9%. Rates of low BW and SGA were 6.0 and 7.3%. Primary CS was needed for 17.7% of deliveries and assisted vaginal birth for 10.3%. Shoulder dystocia complicated 1.7% of birth. Overall, GDM prevalence was 15,3% in our cohort. Application of new criteria would decrease prevalence below 12% but the 308 patients that would have been undiagnosed with GDM did not present better pregnancy outcomes than the rest of the cohort. Conclusion: Despite the high-risk population treated at the CHUM, this study demonstrates lower complications rates in our cohort then in others reported GDM populations (2,3) and results were comparable to general population (4-8) suggesting benefice of even lower diagnostic criteria. Universal standardization of diagnostic criteria could improve GDM care but new recommendations have to be adapted to our high-risk population before validation. Further analyses are needed.[br][br](1) International Association of Diabetes and Pregnancy Study Groups Consensus Panel. International Association of Diabetes and Pregnancy Study Groups Recommendations on the Diagnosis and Classification of Hyperglycemia in Pregnancy. Diabetes Care 2010; 33:676-682. (2) Johns K, Olynik C, Mase R, Kreisman S, Tildesley H. Gestational Diabetes Mellitus Outcome in 394 patients. JOGC February 2006, 122-127. (3) Crowther CA, Hiller JE, Moss JR, et al. Effect of treatment of gestational diabetes on pregnancy outcomes. N Engl J Med 2005; 352:2477-86. (4) Statistics Canada. Birth-related indicators (low and high birth weight, small and large for gestational age, pre-term births), by sex, three-year average 2005-2007 in Canada and Quebec. (5) Canadian Institute for Health Information. Too Early, Too Small: A Profile of small babies Across Canada (2009) p. 1-101. (6) Canadian Institute for Health Information. Giving Birth in Canada: Regional Trends From 2001-2002 to 2005-2006, p. 1-48. (7) Canadian Institute for Health Information. Highlights of 2008-2009 Selected Indicators Describing the Birthing Process in Canada, p. 1-7. (8) The Society of Obstetricians and Gynaecologists of Canada (SOGC) website. Quarisma (Quality of Care, Management of Obstetrical Risks and Birthing Mode in Quebec). www.sogc.org/projects/quarisma_e.asp.[br][br]Nothing to Disclose: THLN, EC, AG 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 269 54 258 SAT-155 PO11-02 Saturday 197 2012


198 ENDO12L_SAT-156 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Racial Differences in Maternal Metabolic Profiles in Gestational Diabetic (GDM) Women at One Year Postpartum Karen E Elkind-Hirsch, Martha S Paterson, Donna L Shaler, Brett L Schelin, Beverly W Ogden Woman[apos]s Hospital, Baton Rouge, LA; Woman[apos]s Hospital, Baton Rouge, LA [bold]Objective[/bold]: Women who develop gestational diabetes mellitus (GDM) have an increased risk for later glucose intolerance and type 2 diabetes (DM2). We examined whether race impacts the incidence of metabolic disturbances 1 year postpartum in previous GDM (pGDM) women.[br][bold]Design[/bold]: From April 2009-October 2011, 170 pGDM (130 white [W] and 40 non-white [NW]) women underwent metabolic assessment at 2-3 months postpartum including evaluation of glucose tolerance by a 75-g oral glucose tolerance test (OGTT) with glucose and insulin concentrations measured at 0, 30, 60, and 120 min. Participants returned at 12-months postpartum for reassessment. Fasting and mean blood glucose (FBG, MBG), cholesterol (C), HDL- and LDL-C, triglycerides (TRG), liver enzymes, TSH, blood pressure (BP), body mass index (BMI), and waist circumference (WC) were measured. Insulin sensitivity and secretion indices were derived from fasting (HOMA-IR and HOMA-B) and glucose-stimulated levels (Matsuda index [SI[sub]OGTT[/sub]] and [Delta]I[sub]0[ndash]30[/sub]/[Delta]G[sub]0[ndash]30[/sub] [IGI])[sup]1-4[/sup]. A measure of [beta]-cell response to glucose challenge was calculated using the insulinogenic index divided by HOMA-IR to adjust for degree of insulin resistance (IGI/IR).[sup]5,6[/sup][br][bold]Results[/bold]: A total of 113 pGDM women (92 W; 21 NW) returned for follow-up; 21 women on medical therapy for impaired glucose regulation (IGR) diagnosed early (2-3 months) postpartum were excluded from the analyses. Fifty-seven pGDM women were not tested at 12 months (7 DM2; 11 pregnant). The study cohort comprised 92 (73 W; 19 NW) pGDM women studied early and 1 year postpartum. At follow-up, dysglycemia was more prevalent in NW women with 47.4% having IGR compared to 24.7% of W women (p[lt].044). Postpartum weight and WC gain, BP, MBG, and IR were significantly greater whereas [beta]-cell secretory response was significantly decreased in NW pGDM women. W pGDM women had a less favorable lipid profile (higher TRG and LDL-C; p[lt].04). W pGDM women were more likely have breastfed compared with their NW pGDM counterparts (p[lt]02). Patient age, treatment of GDM, parity, FBG, TSH, and liver enzymes were not racially disparate.[br][bold]Conclusions:[/bold] Between 3 and 12 months, pGDM women who progressed to IGR manifest low insulin sensitivity, insulin response and [beta]-cell responsivity. Postpartum weight gain, particularly increased central adiposity, was strongly associated with deterioration of [beta]-cell compensation for insulin resistance which may be a key factor in the greater prevalence of glycemic impairment in NW pGDM women.[br][br](1) Matthews DR et al., Diabetologia. 1985; 28: 412 [ndash]419. (2) Matsuda M et al., Diabetes Care 1999;22: 1462[ndash]1470. (3) Sluiter WJ et al., Diabetes 1976; 25:241-244. (4) Wareham, NJ et al., Diabet. Med., 1995; 12, 931. (5) Kosaka K et al., Diabet Med. 1996; 13(9): S109-19. (6) Hanson RL et al., Am J Epidemiol 151:190[ndash]198.[br][br]Sources of Research Support: Irene W. and C.B. Pennington Foundation and Blue Cross/Blue Shield of Louisiana Impact Grant. (ClinicalTrials.gov number, NCT00849849 [ClinicalTrials.gov]).[br][br]Nothing to Disclose: KEE-H, MSP, DLS, BLS, BWO 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 14 54 259 SAT-156 PO11-02 Saturday 198 2012


199 ENDO12L_SAT-157 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Glycemic Control in Familial Versus Sporadic Type 1 Diabetes Patients over 5 Years Sujana Reddy, Steven Reinert, Wendy Plante, Geetha Gopalakrishnan, Charlotte M Boney, JB Quintos The Warren Alpert Medical School of Brown University, Rhose Island Hospital, Providence, RI Background: Previous studies showed that familial type 1diabetes patients (FTID; patients with an affected parent and/or sibling) have less severe metabolic derangement at presentation compared to sporadic patients (ST1D; without affected parent or sibling). Glycemic control was similar at 1 yr follow-up but data on long term metabolic control is lacking.[br]Objective/Hypothesis: 1) FT1D compared to ST1D have less severe presentation at diagnosis and better HbA1c (A1c) over 5 yrs due to parent[apos]s experience in diabetes management 2) In FT1D parent-offspring group, A1c in the child will correlate with parent and 3) in sib-pair group, A1c of the second affected sibling (SP2) will correlate with the first affected sibling (SP1).[br]Methods[bold]:[/bold] Data was collected from T1D children age 6 months to 18 yrs over 20 yr period. Cohort includes 33 parent-offspring and 19 sib-pairs; 33 sporadic patients matched by age, sex, ethnicity, pubertal status, and insulin regimen were selected as controls. We collected A1c, pH, HCO3, glucose at presentation, and A1c, ER and clinic visits and DKA episodes 5 yrs post-diagnosis. Paired T-test and Pearson correlation were used for statistical analysis.[br]Results[bold]: [/bold]At diagnosis, mean age was FT1D (7.7+/- 4.9 yrs) vs ST1D (7.6 +/-4.5 yrs). The metabolic parameters were not different for FT1D vs ST1D (mean A1C: 9.6 vs 10.7%, HCO3: 21 vs 18 meq/l, glucose 428 vs 463 mg/dl and pH 7.35 vs 7.36 (p= ns). In the 5 yr follow up, mean A1C for FT1D vs ST1D (8.9 vs 8.8%; p=0.81), mean clinic (12 vs 12.5, p=0.68) and ER visits (0.48 vs 0.24, p=0.10).[br]Mean glucose at presentation between sib-pair groups (SP1: 433 vs SP2: 347 mg/dl, p=0.49 and HCO3 (SP1:18 vs SP2: 24 meq/l (p[lt]0.01).[br]Mean A1C for SP1 and SP2 was 9.1%. A1C for SP2 correlated positively with that of SP1 (r=0.67, p[lt]0.01). Mean A1C for offspring vs parents (9.3% vs. 8.6 %, p=0.34). A1C for offsprings correlated positively with parents but was not significant (r= 0.57, p=0.18).[br][bold]Conclusion: [/bold]Metabolic control at diagnosis of T1D and the mean A1C over 5 yrs were similar in familial and sporadic patients. In sib-pairs, the second affected sibling had milder clinical presentation compared to the first affected sibling. However, A1C over 5 yrs of SP2 correlates with the SP1. We found only a trend in the correlation of glycemic control between parents and offsprings.Therefore, based on our small sample size, parental experience in diabetes does not positively influence control in their children.[br][br]Nothing to Disclose: SR, SR, WP, GG, CMB, JBQ 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2109 54 260 SAT-157 PO11-02 Saturday 199 2012


200 ENDO12L_SAT-158 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) The Effect of Parental Type 1 Diabetes on Glycemic Control in Children with Type 1 Diabetes Alexandra Martin, Michael B Johnson, Whitney Brown, Marielisa Rincon TC Thompson Children[apos]s Hospital, Chattanooga, TN; The University of Tennessee at Chattanooga, Chattanooga, TN; Palmetto Health Children[apos]s Hospital, Columbia, SC [bold]Background:[/bold] Complex psychosocial factors affect control of type 1 diabetes (T1D) in children. Parental T1D has not been identified as an independent predictor of glycemic control. To date, no published data exist comparing control of T1D in children with and without diabetic parents.[br][bold]Objective:[/bold] Demonstrate whether parental T1D is an independent predictor of poor glycemic control in children withT1D, defined as higher hemoglobin A1c (HbA1c), increased incidence of emergency department (ED) visits, increased incidence of diabetic ketoacidosis (DKA), decreased follow-up, and/or decreased frequency of home blood glucose (BG) testing.[br][bold]Design/Methods:[/bold] Data was collected retrospectively over a 3 year period (November 2007-October 2010) from the charts of preadolescent patients with duration of T1D [gt]12 months who were living full time with a parent with T1D. Patients with parental type 2 diabetes were excluded, as were patients age [gt]12 in order to avoid the confounding effects of adolescent behavior. Controls matched for age and sex were randomly selected from patients with T1D who did not have parents or sibling with T1D. Data collected included average HbA1c, ED visits/year, DKA episodes/year, follow-up visits/year, and BG tests/day.[br][bold]Results:[/bold] 15 subjects and 30 matched controls met criteria for inclusion in this study. Patients with parental T1D had trends toward higher mean HbA1c (0.35% higher, p=0.37), greater incidence of DKA (additional 1.44 episodes/year, p=0.16), greater frequency of ED visits (additional 1.99 visits/year, p=0.05), less follow-up (1.64 fewer clinic/visits/year, p=0.11), and decreased home BG testing (1.76 fewer tests/day, p=0.09) as compared to controls. ANCOVA analysis showed that these results were independent of socioeconomic status or duration of T1D.[br][bold]Conclusions:[/bold] All variables showed trends in the hypothesized direction, with parental T1D being associated with a trend toward higher HbA1c, greater incidence of DKA and ED visits, and poorer compliance with follow-up and BG testing. Larger studies are needed to demonstrate statistical significance, and thereafter, to clarify why diabetic children of diabetic parents have poorer control.[br][br]Nothing to Disclose: AM, MBJ, WB, MR 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 116 54 261 SAT-158 PO11-02 Saturday 200 2012


201 ENDO12L_SAT-159 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Accuracy of Continuous Glucose Monitoring in Pediatric Patients with Diabetic Ketoacidosis Niyati Skaria, Thomas Wilson, Katherine Messina, Andrew Lane Stony Brook University Hospital, Stony Brook, NY Background[br]Continuous glucose monitors (CGM) are increasingly used in the outpatient setting. Patients may make clinical decisions based on CGM readings, including during development of diabetic ketoacidosis (DKA). Volume contraction during DKA contributes to acidosis via lactic acid production. Because CGM devices rely on measurement of generated hydrogen ions, it is unknown whether DKA affects CGM accuracy.[br]Objective[br]We aim to determine whether a commercially available CGM (Dexcom) measures blood glucose measurements accurately compared to a standard of care bedside glucometer in the setting of diabetic ketoacidosis.[br]Design/Methods[br]Succesive eligible patients ages 2 to 21 years old admitted to our hospital in DKA (venous pH [lt] 7.25) were invited to participate. Venous blood gases and chemistry panels were drawn as per standard PICU protocol. Exclusion criteria: other known metabolic conditions, or dermatologic conditions preventing sensor placement. The CGM was placed subcutaneously at presentation. After warm-up, CGM was calibrated to the bedside glucometer initally, and every 12 hours. CGM values were analyzed by comparison to bedside and serum gluocose values.[br]Results[br]In this ongoing study, 91 paired reference vs. CGM values have been obtained to date from 5 patients, ages 12 to 15 years old. The mean absolute relative difference (MARD) between glucometer and CGM values is 29% (SD=25.7). The MARD between serum glucose and CGM values is 26.7% (SD=24). The MARD between mild, moderate, and severe acidosis groups was not statistically different (p[gt].05). In a Clark Error Grid Analysis- 45% of values fell in category A - desirable correlation and 44% in category B - acceptable correlation. A total of 89% fell in category A and B. Of the rest of the values, 9% fell in category C - risk for inappropriate treatment, 2% in categories D and E - high risk.[br]Conclusion[br]Other investigators studying correlations between CGM and reference values in healthy individuals with diabetes have demonstrated MARDs of 13.2 to 21.5%, and 98% of values falling within Clarke Error Grid category A and B. In contrast, our data suggest that during DKA, there is a much higher MARD between CGM and reference values, and more values fall outside Clarke categories A and B. Our results have important implications in counseling patients and caretakers regarding reliance on CGM values at times of risk for the development of diabetic ketoacidosis.[br][br]Nothing to Disclose: NS, TW, KM, AL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 558 54 262 SAT-159 PO11-02 Saturday 201 2012


202 ENDO12L_SAT-160 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) The Incidence of Vitamin D Deficiency in Children with Type 1 Diabetes Mellitus (T1DM) in Northeast Ohio Berrin Ergun-Longmire, Asuman Bilgin, Lillian Prince Akron Children[apos]s Hospital, Akron, OH [bold]Background:[/bold] Vitamin D deficiency is highly prevalent worldwide although it is largely unrecognized. Several epidemiological studies indicate lower serum 25-hydroxy vitamin D (25OHD) levels in diabetic individuals compared to non-diabetic individuals.[br][bold]Objective:[/bold] The Northeast Ohio is one of the cloudiest places in the world. The aim of our study is to investigate the incidence of vitamin D deficiency in children with T1DM and its correlation with their glycemic control.[br][bold]Methods:[/bold] We conducted a retrospective chart review to identify serum 25OHD levels in patients with T1DM between January 1, 2010 and December 31, 2011. Vitamin D deficiency was defined if serum 25OHD level was [le]20 ng/mL. Vitamin D insufficiency was defined if serum 25OHD level was between 21-31 ng/mL. Levels of HbA1C and vitamin D were stratified in four seasons. The four seasons are defined as follows: Winter, December to February; Spring, March to May; Summer, June to August; and Fall, September to November. We used the SAS 9.2 statistical software for data analyses.[br][bold]Results:[/bold] Data were available from 200 children with T1DM (96 females, 104 males). The mean age, in years, was 12.51[plusmn]3.93. The mean duration of T1DM was 5.12[plusmn]3.31 years. The overall incidence of vitamin D deficiency was 12.5% and insufficiency was 46%. HbA1C was found to be higher in patients with vitamin D deficiency and insufficiency. When analyzing for a correlation using Pearson correlations (r) between HbA1C and serum 25OHD levels, correlations were not significant.[br]A one-way between subjects ANOVA was conducted to compare the effect of the four seasons on serum 25OHD levels. There was a significant effect of the seasons on the Vitamin D25 levels at the p[lt]0.05 level [F(3, 196) = 10.10, p = 0.0001]. The Bonferroni post-hoc comparison test indicated that the mean Vitamin D25 level for the Summer season (34.95[plusmn]9.68) was significantly different than all of the other seasons. These results suggest that highest levels of Vitamin D25 are indeed found in the Summer season. Specifically, the results confirm that the levels decrease during seasons of less sunlight. However, there was no significant difference in the mean Vitamin D25 levels for the pair-wise comparisons between the other seasons.[br][bold]Conclusion: [/bold]Although vitamin D deficiency/insufficiency is common in children with T1DM, there is no statistically significant correlation between their glycemic control and serum vitamin D levels.[br][br]Nothing to Disclose: BE-L, AB, LP 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1772 54 263 SAT-160 PO11-02 Saturday 202 2012


203 ENDO12L_SAT-161 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Type 2 Diabetes Mellitus Re-Emergence Post Gastric Bypass Surgery Yessica Ramos, Sophie Bersoux, Lori R Roust, Yu-Hui H Chang Mayo Clinic Arizona, Scottsdale, AZ Roux-en-Y gastric bypass surgery (RYGB) is an effective intervention for improvement and remission of type 2 diabetes mellitus (T2DM). Limited data exist about re-emergence of T2DM after this procedure. A retrospective analysis of patients who underwent RYGB from January 2000 to December 2007 at our institution was performed. 138 patients with documented T2DM were identified. Of these, 72 patients (52%) had minimum 3-year continuous follow-up at our institution. The 72 patients were assessed pre- and post-operatively at various time intervals. Patients[apos] baseline demographics, weight, body mass index (BMI), HbA1C, fasting glucose and diabetic pharmacological treatment were documented. The purpose of the analysis was to determine the persistence, remission and re-emergence of T2DM. Remission of T2DM was defined as HbA1c of less than 6.5% with no pharmacological treatment. Re-emergence was defined by HbA1c of equal to or [gt]6.5%, fasting glucose [gt] 7mmol/L or re-initiation of anti-diabetic medications. The patients[apos] mean pre-operative BMI was 45 kg/m2. The majority were Caucasian (93%), mean age was 49.5 +/- 9.3 years, and 54% were female patients. The mean duration of T2DM preoperatively was 4.9 +/- 4.4 years. 66 patients (91.6%) were in remission at a minimum of one time point while 6 patients (8.4%) had persistence of T2DM throughout follow-up. Of the 66 patients that had remission, 14 patients (21.2%) had recurrence of T2DM. Five of these patients developed T2DM at two years post RYGB, 3 at 3 years, 3 at 4 years and 3 at 5 years. Duration of T2DM prior to RYGB was significantly associated with recurrence of T2DM (p=0.0021). The odds ratio was 1.26 indicating that longer duration of pre-op T2DM was associated with higher probability of recurrence. No significant association between pre-op BMI or weight regain was identified with a higher recurrence rate. Conclusion: The recurrence rate of T2DM after RYGB is 21%. Duration of diabetes prior to RYGB was associated with a higher recurrence rate of T2DM. This suggests that early surgical intervention in the type 2 diabetic obese population may improve durability of remission of T2DM.[br][br]Nothing to Disclose: YR, SB, LRR, Y-HHC 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1881 54 264 SAT-161 PO11-02 Saturday 203 2012


204 ENDO12L_SAT-162 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) FPG, OGTT, HbA1c: Which Combination Best Detects Diabetes Mellitus? Daryl Wei Chong Tan, Daniel Ek Kwang Chew Tan Tock Seng Hospital, Singapore, Singapore Background: Previous methods of diagnosing diabetes mellitus in Singapore were the fasting plasma glucose (FPG) and the 2-h 75g oral glucose tolerance test (OGTT). However, the American Diabetes Association (ADA) has recommended the use of HbA1c as an alternative test for diabetes mellitus with a threshold of [ge] 6.5% in 2010. These diagnostic tests detects differing pathophysiology of dysglycemia amongst diabetes subjects and as such do not demonstrate complete concordance. This study aims to determine which combination of the three tests best detects diabetes mellitus.[br]Methods: Data of subjects who had HbA1c, FPG and OGTT tests done in the same seating between January 2007 and March 2010 at Tan Tock Seng Hospital Diabetes Centre were collected and analysed. Subjects with FPG values [ge] 7.0mmol/l, OGTT values [ge] 11.1 mmol/l, or HbA1c [ge] 6.5% were classified as diabetes mellitus.The records of 250 subjects were used.[br]Results: Of the 250 subjects in the study, 133 (53.2%) had newly diagnosed diabetes mellitus defined by a high FPG or OGTT or HbA1c. OGTT has the highest detection rate of diabetes mellitus (82.7%), followed by HbA1c (62.4%) and then FPG (52.6%). The combination of tests with the highest detection rate is that of HbA1c plus OGTT (97.7%), higher than the combination of FPG plus OGTT (89.5%) or FPG plus HbA1c (72.9%).[br]Conclusion: The combination of HbA1c and OGTT has a highest detection rate of diabetes mellitus out of the three tests FPG, OGTT and HbA1c.[br][br]Nothing to Disclose: DWCT, DEKC 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 295 54 265 SAT-162 PO11-02 Saturday 204 2012


205 ENDO12L_SAT-163 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Vitamin D Deficiency across the Spectrum of Glucose Intolerance in an Adult Population at a Northeast Florida Clinic Arnold Vera, Paul Yosmin Casanova-Romero, Sean Patrick Hassett Vera Endocrine Associates, Inc, Daytona Beach, FL Background: The United States is facing a high prevalence of glucose intolerance and diabetes mellitus (DM). Although obesity is clearly a predominant factor in the pathogenesis of diabetes, other modifiable lifestyle factors and certain nutritional factors, such as vitamin D deficiency, are also believed to play a role. The aim of this study was to investigate[bold] the proportion of individuals with vitamin D 25-(OH) insufficiency/deficiency (VitD-)[/bold] across the spectrum of glucose status in an adult population referred for endocrine evaluation at a Northeast Florida clinic.[br]Methods: We performed a comprehensive history, physical and laboratories on consecutive subjects referred for an endocrine evaluation to determine the prevalence of VitD- and its relationship with age, BMI, glucose status, renal status by GFR categories, serum PTH, calcium, phosphorus and magnesium. A Chi-squared test was used to test the proportions for each group.[br]Results: 1663 consecutive subjects aged 18-98 and a mean BMI of 31.42 were evaluated. 51.9 % were female. 1077 individuals had a glucose abnormality: 28.0 % had impaired fasting glucose (IFG) or impaired glucose tolerance (IGT), 55.8 % were diagnosed with Type 2 DM and 16.2 % with Type 1 DM. Females diagnosed with IFG/IGT were found to have a significantly higher proportion of VitD- (40.7 %) than subjects with diagnosis of Type 1 (25.3 %) or Type 2 DM (29.0 %), independent of their age, gender and BMI (Chi-square 7.33 df 2 p[lt]0.05). Adults [lt] 55 year/old with Type 2 DM have a higher proportion of VitD- (42.6%) than subjects 55 to 75 year/old (33.9 %) or [gt] 75 year/old (21.1 %) (Chi-square 12.44 df 2 p[lt]0.001). Obese individuals diagnosed with IFG/IGT (73.3 %) and Type 2 DM (35.3 %) have the highest proportion of VitD- compared with normal or overweight subjects, independent of GFR categories (Chi-square 11.7 df 1 p[lt]0.001; Chi-square 6.9 df 1 p[lt]0.01, respectively).[br]Conclusions: Our data suggests that there is an increased prevalence of vitamin D insufficiency/deficiency in the Southeast female population of the USA with diagnosis of IFG/IGT. Obese adults [lt] 55 year/old with IFG/IGT experience the highest prevalence of VitD- independent of their renal status. We conclude that there is a need to do an early analysis and adjustment of vitamin D levels in adult obese individuals with diagnosis of pre-diabetes; future studies will determine if vitamin D treatment will delay the onset of diabetes.[br][br]Nothing to Disclose: AV, PYC-R, SPH 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1530 54 266 SAT-163 PO11-02 Saturday 205 2012


206 ENDO12L_SAT-164 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Assessment of Pain and Treatment Satisfaction in Patients with Painful Diabetic Peripheral Neuropathy Latha Dulipsingh, Susan Zailskas, Teresa McInnis, Aniello Marotta The Hospital of Central Connecticut, New Britain, CT; Pfizer,Inc, New Haven, CT The purpose of this project was to describe pain experienced from painful diabetic peripheral neuropathy (pDPN) and evaluate current treatment satisfaction in patients followed through an endocrinology clinic.[br]Patients [ge] 18 years of age with a diagnosis of diabetes, with or without diabetic peripheral neuropathy, were offered a voluntary self-administered questionnaire, prior to their examination by the healthcare provider. Only patients who had nerve pain were asked to complete the entire questionnaire. In addition to basic information related to diabetes, patients were asked if they experienced nerve pain, the characteristics of the nerve pain, and questions related to treatment and satisfaction with current therapy.[br]A total of 98 questionnaires were collected with 53.1% of the patients being female in gender. The mean age was 55.1 years, 75.6% had type 2 diabetes and the mean duration of diabetes was 16.3 years. Thirty-one patients (31.6%) reported neuropathic pain with 83.3% having experienced nerve pain for one year or greater and the majority (67.7%) reporting nerve pain in their feet. The percentage of patients who reported moderate to severe nerve pain, score of [ge]4 on the 0-10 numeric pain rating scale, over the past week was 54.8% with 32.3% reporting nerve pain [ldquo]All Day [ldquo] in the past 24 hours. Impact of nerve pain on quality of life was noted by 61.3% of patients although 64.5% of patients reported that nerve pain did not cause them to avoid activities. Fifteen (48.4%) patients reported receiving medication for their nerve pain of which 66.7% reported being satisfied with their treatment. Of the satisfied patients, 40% reported severe nerve pain over the past week with 30% reporting pain [ldquo]All Day[rdquo].[br]Painful diabetic peripheral neuropathy is a common complication of diabetes mellitus and can have debilitating consequences with a significant impact on quality of life. Based on our questionnaire we found about a third of patients with long standing diabetes having pDPN of which about two thirds of them reported that it affected their quality of life. Surprisingly, almost half of the patients reported being satisfied with their treatment despite severe nerve pain.[br]It is important for healthcare providers to recognize barriers to pain control in patients with pDPN. Our findings will serve as a foundation for a pDPN provider-patient initiative focused at improving education and communication at our clinic.[br][br]Nothing to Disclose: LD, SZ, TM, AM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1569 54 267 SAT-164 PO11-02 Saturday 206 2012


207 ENDO12L_SAT-165 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Comparison of the Adherence to Guidelines of Diabetes Care in Primary Care and Subspecialty Clinics Lily K Sunio, Deepthi T Rao, Ved Gossain Michigan State University, East Lansing, MI Background:[br]Diabetes mellitus, with an estimated US prevalence of 8.3%, is a major public health problem with significant morbidity and mortality. Evidence based guidelines have been proposed to reduce the micro and macrovascular complications, but studies have shown that these goals are not being met.[br]Objective:[br]To compare the adherence to American Diabetes Association guidelines for measurement and control of glycohemoglobin (A1c) blood pressure (BP), lipids(LDL) and microalbuminuria (MA) by subspecialty and primary care clinics in an academic medical center.[br]Methods:[br]390 randomly selected charts of patients with diabetes mellitus from diabetes (DM), Internal Medicine (IM) and Family practice (FP) clinics at Michigan State University were reviewed. We assessed measurement and control of A1C, LDL, BP and MA during the one year period preceding their most recent visit.[br]Results:[br]Our study included 131, 134 and 125 charts from the FP, IM and DM clinics, respectively. Mean age of the subjects was 61[plusmn]11 (FP), 62.11[plusmn]12.88 (IM) and 52.82[plusmn]16.01 (DM) years. DM clinic had a higher percent of type 1 diabetes (34.4%) compared with about 5% in FP and IM clinics. A1C was measured in 99.2, 97.8, 100% of patients and was controlled ([lt]7%) in 38.9, 43.3, 28.8% of patients in the FP, IM and DM clinics, respectively. LDL was measured in 93.9, 91.8, 89.6% of patients and was controlled ([lt]100mg/dL) in 71.8, 64.9, 64% of patients in the FP, IM and DM clinics, respectively. MA was measured in 85.5, 82.8, and 76.8% of patients in the FP, IM and DM clinics, respectively. BP was measured 100% of the time in all clinics. 59.5, 67.2, 52.8 % of patients in the FP, IM and DM clinics, respectively had their BP controlled ([lt]130/80). Insulin use was associated with poor glycemic control. Patients on insulin were more likely to have A1C[ge]7 % (OR=4.37, p=0.00). Patients with uncontrolled BP were more likely to have LDL[ge]100mg/dL (OR=1.8, p=0.02). Patients on diuretics were more likely to have BP[gt]130/80 (OR=1.7, p=0.05).[br]Conclusion:[br]Our data show that the rate of HbA1C, LDL, and BP testing was high, similar to the national average in both subspecialty and primary care clinics. However, the degree of control of these parameters was still suboptimal which could be influenced by several factors. Continued efforts to improve these parameters are still needed.[br][br]Nothing to Disclose: LKS, DTR, VG 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2086 54 268 SAT-165 PO11-02 Saturday 207 2012


208 ENDO12L_SAT-166 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Remission in Minorities with Newly Diagnosed Diabetes Mellitus Treated in a Real World Setting Abhishek Kansara, Yin Oo, Pranav Ghody, Anuja Kriplani, Alejandra Borensztein, Rajeev Sharma, Mikhail Kagan, Mary Ann Banerji State University of New York (SUNY) Downstate Medical Center, Brooklyn, NY Objective[br]Remission in type 2 diabetes is described in African-Americans, Hispanics, Asians and Caucasians with a frequency ranging from 20 to 50 % in a research setting. Our goal was to determine the frequency of remission in newly diagnosed minority diabetes patients treated in a real world clinic setting. We examined clinical factors influencing remission.[br]Research Design and Methods[br]We retrospectively analyzed data of 87 newly diagnosed diabetes patients, seen regularly in diabetes or primary care clinic. Therapeutic decisions were individualized. Remission was defined as achieving normoglycemia with A1C [le] 6.3% on medications. We expect that true remission (off all medications) would come from this subset. Remitters and non-remitters were analyzed by chi square and independent samples[apos] t-tests. Data are mean[plusmn]SD.[br]Results[br]39 out of 87 (45%) achieved normoglycemia while 48 (55%) did not. The baseline characteristics of the remission and non-remission group were not statistically different in terms of age (48[plusmn]12 years), gender (2/3 male) or BMI (26.7[plusmn]12). The initial A1C was lower in the remission group (10.6[plusmn]2.9% vs 12.1[plusmn]2.8%; p=0.015). The initial plasma glucose and clinically relevant microalbuminuria were both significantly higher in the remission vs the non-remission group. The follow-up time was 12.8[plusmn]13.0 months.[br]Patients attending diabetes clinic were more likely to develop remission than those in primary care clinic (67% vs 29%, p=0.014). Treatment using insulin or insulin plus oral anti-diabetic agents was twice as effective as those treated with oral agents alone (63.2% vs 30.6%, p=0.004). Clinicians in diabetes clinic were more likely to prescribe insulin based treatment compared to those in primary care clinic (53% vs 12%, p=0.0001). The time to achieve remission was 4.5 [plusmn]2.3 months.[br]Summary[br]Nearly half (45%) of minority patients in a real world setting achieve near normoglycemic remission. Key determinants of such imporved glucose homeostasis are treatment in a diabetes clinic and the use of insulin at the outset which doubled the remissions compared to primary care clinic. Prolonged near-normoglycemia is known to prevent diabetes complications, thus, inducing remissions is likely to have far reaching public health benefits in reducing morbidity. Our data suggest that early referral of new onset diabetes to a specialist and the use of insulin based treatment optimizes the likelihood of inducing remission.[br][br]Disclosures: MAB: Clinical Researcher, Merck & Co.; Scientific Board Member, Novartis Pharmaceuticals; Speaker Bureau Member, Sanofi-Aventis, Merck & Co. Nothing to Disclose: AK, YO, PG, AK, AB, RS, MK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2374 54 269 SAT-166 PO11-02 Saturday 208 2012


209 ENDO12L_SAT-167 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Deterioration of Diabetes Glucose Control during Civil Unrest M Loutfy Sakkal, Saad Sakkal Metabolic Care Center, Aleppo, Syrian Arab Republic; University of Kalamoon, Deir Atieh, Syrian Arab Republic Context: Civil unrest is stressful to all citizens[apos]. Stress effect diabetes control in general, but no or rare studies showed the influence of civil unrest on diabetes glucose control. We quantitated the effect during the recent Syrian civil unrest.[br]Methods:[br]In our clinic patients are seen quarterly. In each visit we review H[amp]P, laboratory parameters and undercurrents psychosocial factors.[br]In 200 usual clinic patients we determined means FBS, 2HPP, HgA1c during 2 distinct comparable periods: for the 6 months before the civil unrest and after (July to December 2010, 2011).We excluded new onset diabetes patients and other illnesses for a separate reports.[br]Results:[br]In the period before civil unrest July to December 2010 There was a drop in FBS: 37 mg/dl (177 to 140), 2hpp 62(214 to 152), HgA1c:2.5 %(%9.9 to 7.4) and insulin dose by 22 %(50 to38) from baseline. In the period after civil unrest July to December2011 there was a rise in FBS: 47(137 to 184), 2hpp:128(161 to 289), HgA1C:2%%7.6 to 9.6) and insulin dose by 25 %(44 to 55).[br]Conclusion:[br]Diabetes Glucose control is affected by civil unrest, probably as a result of stress, lack of health resources or other factors. We need more studies for diabetes during unfortunate civil unrest events.[br][br]Nothing to Disclose: MLS, SS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 155 54 270 SAT-167 PO11-02 Saturday 209 2012


210 ENDO12L_SAT-168 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Response to Vitamin D Supplementation in Type 2 Diabetes Patients with Pulmonary Tuberculosis Sunil K Kota, Siva K Kota, Svs Krishna, Lalit K Meher, Kirtikumar D Modi Medwin Hospital, Hyderabad, India; Central Security Hospital, Riyadh, Saudi Arabia; MKCG Medical College, Berhampur, India Objective:To evaluate effects of vitamin D supplementation on type 2 diabetes mellitus (T2DM) patients with pulmonary tuberculosis(PTB).[br]Methods:45 subjects (M:F= 34: 1) were screened.Inclusion criteria :Age[gt]15 years,newly diagnosed PTB cases with uncontrolled T2DM, serum vitamin D[lt]20 ng/ml.The patients were randomly assigned to 2 groups.Group 1 subjects received oral cholecalceferol(60000 units/week) and calcium carbonate (1 gm/day) along with anti tubercular treatment(ATT),while group 2 subjects did not. Sputum was checked at interval of 2 weeks for 12 weeks.Primary end point was time to achieve sputum smear conversion.Secondary end points were reduction in ESR and improvement in glycemic parameters.[br]Results:15 patients with vitamin D[gt]20 ng/ml were excluded.So the prevalence of vitamin D deficiency in T2DM with PTB was 30/45 (66.66 %),with 25/34 males (73.5%)and 5/11 females (45.5%)were vitamin D deficent.Mean age of patients (30) was 39.5[plusmn]18.9 years with FBS 230.5[plusmn]30.3 mg/dl,PLBS 320.5[plusmn]45.6 mg/dl,HbA1C 10.4[plusmn]4.4 % and 25(OH)D 12.1[plusmn]4.3 ng/ml. At end of 12 weeks,group 1 patients had significantly higher levels of serum 25(OH)D (25.4[plusmn]6.9 ng/ml in group 1 versus 10.2[plusmn]0.9 ng/ml in group 2).Sputum smear conversion was 6 weeks in group 1 versus 8 weeks in group 2(p= 0.067).Reduction in ESR was significant in group 1 vs group 2 (39.6 [plusmn] 12.4 mm/1st hr vs 24.0 [plusmn] 14.9, p-0.004).Difference in the reduction in FBS, PLBS and HbA1C in the 2 groups did not attain statistical significance.[br]Discussion:Correlations exist between low vitamin D levels with PTB and T2DM separately.Calcitriol modulates response to mycobacterial infection by induction of reactive nitrogen and oxygen intermediate,suppression of matrix metalloproteinase enzymes implicated in pulmonary cavitation, and induction of antimicrobial peptide cathelicidin.Calcitriol modulates immune responses by binding vitamin D receptors expressed by antigen presenting cells and activated lymphocytes. Several case series have reported utility of 25000 IU to 100000 IU vitamin D in improving patients[apos] response to ATT (1). Vitamin D supplementation also increases lymphocyte to monocyte ratio,a biomarker of disease resolution.We provided vitamin D at the dosage of 60000 IU/week, and report a 2 weeks reduction in sputum smear conversion.[br]Conclusion: Vitamin D may be the missing link between emerging epidemic of tuberculosis [amp] diabetes.It can serve as adjuvant treatment of tuberculosis in diabetics with vitamin D deficiency.[br][br]1. Martineau AR, et al., J Steroid Biochem Mol Biol 2007; 103: 793.[br][br]Nothing to Disclose: SKK, SKK, SK, LKM, KDM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1403 54 271 SAT-168 PO11-02 Saturday 210 2012


211 ENDO12L_SAT-169 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Impairment in Glycemic Control and Lipid Profile during Indian Festival Diwali Nitish Mathur, Rachita Mathur, Jairam Rawtani Dr S N Medical College, Jodhpur, India; Dr S N Medical College, Jodhpur, India INTRODUCTION: The effect of cultural factors on glycemic control in Asian Indian diabetics has not been studied in detail. Hindu festival of Diwali is associated with increased consumption of sweets and oil rich fried dishes and likely to influence metabolic control. Therefore, present study aims at estimating the changes in glycemic control and lipid profiles after Diwali.[br]METHOD: The study was conducted on 74 T2DM patients (age 60[plusmn]8.26 years, M:F ratio 43:24), recruited from out-patient department of Dr. S. N. Medical College, Jodhpur, India.T2DM was diagnosed as per ADA criteria, and all the subjects were on oral anti-diabetic drugs. Blood samples were collected 2 weeks before and after the Diwali. The patients were instructed to continue with the same oral anti-diabetic medication during this period. Following parameters were measured from the samples: Fasting and post-prandial plasma glucose, Lipid profile (serum total cholesterol, triglycerides, HDL, LDL and VLDL cholesterol)[br]RESULTS: Out of total 74 patients, 67 patients returned for follow-up. There was a significant increase in fasting (136.85[plusmn]2043 vs.154.02[plusmn]19.16, p = 1.82 [times] 10[sup]-14[/sup]), and post-prandial (185.22[plusmn]30.66 vs. 207.05[plusmn]31.16, p = 8.63 [times] 10[sup]-14[/sup]) plasma glucose, total cholesterol (198.47[plusmn]18.84 vs. 212.98[plusmn]19.06, p = 1.69 [times] 10[sup]-15[/sup]), triglycerides (136.37[plusmn]25.76 vs. 153.44[plusmn]23.95, p = 1.18 [times] 10[sup]-15[/sup]), LDL (130.25[plusmn]16.42 vs. 142.28[plusmn]17.50, p = 4.56 [times] 10[sup]-13[/sup]) and VLDL (27.27[plusmn]5.15 vs. 30.68[plusmn]4.79, p = 1.18 [times] 10[sup]-15[/sup]) cholesterol after Diwali. Whereas, HDL was found to be reduced significantly after Diwali (40.94[plusmn]3.75 vs. 40.01[plusmn]2.78, p = 3.57 [times] 10[sup]-4[/sup]).[br]CONCLUSION: Diwali is associated with significant impairment in glycemic control and lipid profile. Therefore, around the festive season, prescription must be adjusted in accordance with the change in food intake and patients must be provided with specialized diet counseling for the season.[br][br]Nothing to Disclose: NM, RM, JR 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1181 54 272 SAT-169 PO11-02 Saturday 211 2012


212 ENDO12L_SAT-170 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Glucose Control during a Driving Training in Patients with Diabetes Peter Wiesli, Joel Capraro, Beat Frauchiger, Giatgen A Spinas, Roman Truninger Kantonsspital Frauenfeld, Frauenfeld, Switzerland; University Hospital of Zurich, Zurich, Switzerland [bold]Objective[/bold]: To investigate the effect of prolonged acute mental stress by means of a driving training on glucose control in patients with type 1 and type 2 diabetes.[br][bold]Research Design and Methods[/bold]: 39 patients with insulin-treated diabetes (18 type 1, 21 type 2 diabetes) were exposed to mental stress by means of a 2hr-driving training. The training session started 15 min after intake of a standard meal. Plasma glucose, blood pressure, heart rate, salivary cortisol, and stress perception (by visual analog scale) were monitored in regular intervals and compared to a control day either preceding or following the stress testing day, respectively.[br][bold]Results[/bold]: On the stress testing day, blood pressure rose from 142/86 [plusmn] 16/9mmHg at baseline to 157/93 [plusmn] 17/10mmHg (p[lt]0.05), heart rate from 72 [plusmn] 11bpm to 84 [plusmn] 15bpm (p[lt]0.05), stress perception from 1.4 [plusmn] 0.6 to 4.1 [plusmn] 1.9 points (p[lt]0.05), and salivary cortisol concentrations from 5.8 [plusmn] 3.3nmol/l to 7.6 [plusmn] 3.9nmol/l (p[lt]0.05); these measurements remained stable on the control day. In the entire study group, the course of glucose concentrations showed no significant difference on the stress testing day compared to the control day. However, 67% of patients (26/39) displayed a change of glucose concentrations of more than 2mmol/l on the stress testing day as compared to the control day; with 46% of patients (18/39) showing an increase of [gt]2 mmol/l (median 3.9mmol/l, range 2.0 [ndash] 8.9mmol/l, p[lt]0.05), and 21% of patients (8/39) showing a decrease of [gt]2mmol/l (median 4.5mmol/l, range 2.7 [ndash] 11.0mmol/l, p[lt]0.05), respectively. 33% of patients (13/39) experienced no relevant alteration of blood glucose ([lt]2mmol/l).[br][bold]Conclusions[/bold]: Although a majority of patients with diabetes display clinically relevant deviations of blood glucose control during prolonged mental stress, there is no constant pattern of glucose dysregulation.[br][br]Nothing to Disclose: PW, JC, BF, GAS, RT 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 127 54 273 SAT-170 PO11-02 Saturday 212 2012


213 ENDO12L_SAT-171 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Use of Professional Continuous Glucose Monitoring (PCGM) in Outpatients with Diabetes at a Community Hospital Keya Malhotra, Sally M Pinkstaff, Naomi Walpert Sinai Hospital of Baltimore, Baltimore, MD [bold]Background[/bold]: Glucose monitoring is recommended for most patients with diabetes. Two techniques are available: self-monitoring of capillary glucose (SMG), and continuous monitoring of interstitial glucose (CGM). Intermittent use of CGM has been used when glucose control is not achieved. Recent AACE and Endocrine Society guidelines have outlined uses for PCGM, including suspected hypoglycemia (hypo), A1C not at target, postprandial hyperglycemia (hyper), or major medication change. The diabetes center (DC) at Sinai Hospital has used PCGM for 3 years. We conducted this chart review to evaluate the patient population referred for PCGM and the results of the PCGM tracings. Objectives: To assess the validity of the PCGM profiles, to compare the reasons for referral with the guidelines, and to identify the frequency of hypo/hyper excursions.[br]Design: Retrospective chart review.[br]Setting: Outpatient DC.[br][bold]Methods[/bold]: 98 patient profiles from January-June 2010 were downloaded using PCGM software. A valid PCGM profile required: 1)[gt]260 readings on each of 3 consecutive days, 2)[gt]2 paired SMG readings/day, 3) Mean Absolute Difference [lt]28%, and 4) R[gt]0.79. Frequency of hyper ([gt]180 mg/dl) and hypo ([lt]70 mg/dl) was calculated from a consecutive 72 hour tracing. 52 patient records with valid PCGM profiles were reviewed.[br][bold]Results[/bold]: Fifty-two PCGM profiles met the criteria for validity. 12 patients had type 1 diabetes (T1D), 40 had type 2 diabetes (T2D), and 37 had an A1C[gt]7%. 43 patients were on insulin. Testing indications were: elevated A1C, suspected hypo, change in medical regimen, dawn phenomenon, or postprandial hyper. PCGM tracings showed hyper 0-20% of the time in 4 T1D patients and 20 T2D patients. Six T1D patients and 13 T2D patients had hyper 21-50% of the time. Overall, hyper for [gt]20% of the time was detected in 67% of T1D and 50% of T2D. No hypo was detected in 2 of T1D and 25 of T2D patients. Hypo for [gt]10% of the time was found in 50% of T1D and 14% in T2D. 4 and 10 of T1D and T2D patients respectively, had hypo between 1-10% of the time.[br][bold]Conclusions[/bold]: This pilot study shows that use of PCGM in our DC needs revision. 50% of tracings were invalid, suggesting inadequate patient training. Reasons for referral agreed with current guidelines in most patients. PCGM profiles did demonstrate frequent hypo and hyper episodes in many patients and can be used to guide medication changes.[br][br]Nothing to Disclose: KM, SMP, NW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1798 54 274 SAT-171 PO11-02 Saturday 213 2012


214 ENDO12L_SAT-172 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) The Development of a Simplified and Integrated Biosensor for Diabetics Francis A Eusebio, Teagan Adamson, Curtiss B Cook, Jeffrey T La Belle Arizona State University, Tempe, AZ; Mayo Clinic Arizona, Scottsdale, AZ Self-monitoring of blood glucose (SMBG) is standard of care in diabetes management. Current technologies for SMBG are based upon enzymatic electrochemical sensing. To increase the sensitivity and specificity of current devices, we are developing a novel method of detecting glucose using electrochemical impedance spectroscopy (EIS) technology. EIS is a frequency based electrochemical method similar to amperometric methods but with the addition of an alternating current (AC) signal (1). This label free technique is capable of detecting femtoMolar concentrations of the desired marker. To test the ability of EIS methods to detect glucose, the enzyme glucose oxidase (GOx) was fixed to gold electrodes through the means of a specific immobilization process. This process involved a self-assembling monolayer, a 16- mercaptohexadecanoic, an N-hydroxysulfosuccinimide and 1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride coupling intermediate, and a blocking element to prevent the attachment of interferents. Once GOx was fixed to the gold electrode surface, a 5 mV sine wave sweeping frequencies from 100 kHz to 1 Hz was induced at glucose concentrations of 0, 1, 4.5, 8, 10, 25, 60, 85, and 100 mg/dL mixed with a ferricyanide redox mediator. Each frequency in the 100 kHz range sweep was analyzed for highest response and r-squared value. The frequency with both factors optimized is specific for the glucose-GOx binding interaction, and was determined to be 966.8 Hz. Four separate electrodes were constructed and data from each were averaged. The mean lower limit of detection was a glucose concentration of 14.6 mg/dL with a relative standard deviation of 14.7 %. The correlation between the impedance response and concentration was determined to be 11.5*ohm/log(mg/dL) with a r-squared value of 0.8. The above data confirm that EIS offers a new method of glucose detection, and development of a sensor strip utilizing EIS methods offers a potential alternative technology for SMBG that offers improved detection at lower concentrations of glucose. The unique frequency response of individual markers allows for modulation of signals so that several markers could be measured with a single sensor. Future work includes assessment of other diabetes associated biomarkers that can be measured on a single sensor, integration testing and tuning of the biomarkers, impedance-time sensing development, and finally, testing on diabetes (and control) subjects.[br][br](1) La Belle JT, Demirok UK, Patel DR, Cook CB. Development of a Novel Single Sensor Multiplexed Marker Assay. Analyst, 2011 136:1496-1501.[br][br]Disclosures: CBC: Coinvestigator, The Epsilon Group. Nothing to Disclose: FAE, TA, JTLB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 400 54 275 SAT-172 PO11-02 Saturday 214 2012


215 ENDO12L_SAT-173 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Capillary and Venous Blood Samples Analyzed in Glucometers Compared to a Laboratory Hexokinase Method Milena G Teles, Suzimara A Oliveira, Isio Schulz, Nairo M Sumita, Maria Elisabeth Rossi, Marcia Nery Hospital das Cl[iacute]nicas da Faculdade de Medicina da Universidade de S[atilde]o Paulo, S[atilde]o Paulo, Brazil; Hospital das Cl[iacute]nicas da Faculdade de Medicina da Universidade de S[atilde]o Paulo, S[atilde]o Paulo, Brazil Blood glucose meters are widely used for monitoring diabetes. A question common to both health professionals and patients is how much glucose meters values actually correlate with laboratory values. [bold]Aim: [/bold]To evaluate the performance of two brands of glucometer using both capillary and venous samples compared to plasmatic glucose (PG) analyzed in a laboratory. [bold]Patients and[/bold] [bold]Methods[/bold]: 237 capillary and venous blood specimens of 188 women and 45 men (age 14-87 years) were analyzed on both Accu-Chek Active (ACA)-Roche Diagnostics and Optium Xceed (OX)-Abbott Diabetes glucometers and compared with plasma glucose analysis in a clinical laboratory (hexokinase method). All blood samples were collected by the same trained meter operator. Venous samples were obtained by venipuncture and collected in tubes with fluoride. The time between capillary and venous samples collection was [lt] 3 minutes. After antisepsis with alcohol, the first drop of capillary blood was discarded and the second and third drops were analyzed in each meter in alternate days. [bold]Results[/bold]: The PG values range was 42-447 mg/dL. The correlation coefficients (CC) between capillary samples analyzed in the two devices and PG were similar: 0.973 (OX) and 0.950 (ACA). The mean percentage variation (MPV) regarding capillary and plasmatic levels was 7.84 and 8.39 for OX and ACA, respectively. Concerning venous samples analyzed in the meters, the CC was 0.931 (OX) and 0.978 (ACA) and MPV was 15.45 and 5.94 for OX and ACA, respectively. There was no significant statistical difference in relation to the order of drops analyzed in the meters. The mean absolute variation in the two devices between capillary samples and PG was significantly higher in individuals with blood glucose levels [ge] 100mg/dL than in individuals with levels [lt] 100mg/dL (p[lt]0.001), but the percentage variation showed no statistical difference (p[gt]0.05). [bold]Conclusions:[/bold] Capillary measurements in the two meters were comparable to PG levels analyzed in the clinical laboratory (variation [lt]10%). Venous samples analyzed in the meters showed satisfactory correlation coefficients, with higher variation in the Optium Xceed device. In conclusion, we observed a good correlation between capillary blood glucose and plasma glucose levels measured by a standard method compared to the two devices tested. Studies to evaluate the correlation of glucometers results and PG levels analyzed by a standard method may improve decision making in diabetes treatment.[br][br]Nothing to Disclose: MGT, SAO, IS, NMS, MER, MN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1487 54 276 SAT-173 PO11-02 Saturday 215 2012


216 ENDO12L_SAT-174 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Cross-Reactivity of Modern Insulin Analogs Insulin Detemir and Insulin Glulisine with Six Automated Insulin Immunoassays Stephen D Merrigan, William E Owen, Sonia L La[apos]ulu, Sara P Wyness, William L Roberts, Joley A Straseski ARUP Institute for Experimental Pathology, Salt Lake City, UT; University of Utah, Salt Lake City, UT Introduction: Identifying whether recombinant insulin analogs cross-react with commercially available insulin immunoassays is critical for accurate result interpretation. While the cross-reactivity of several commonly used analogs has been published previously, it is medically relevant to evaluate newly developed insulin analog formulations across multiple assay platforms.[br]Methods: We determined the cross-reactivity of insulin detemir (Levemir; Novo Nordisk) and insulin glulisine (Apidra; Sanofi-Aventis) with insulin immunoassays on 6 automated platforms at 4 concentrations (30, 100, 300, and 1000 [micro]IU/mL in BSA). Analogs were assayed per manufacturer instructions in duplicate on the Modular E170 (Roche Diagnostics), Liaison (DiaSorin), UniCel DxI 800 (Beckman Coulter), Advia Centaur and Immulite 2000 (Siemens Diagnostics), and Architect i2000 (Abbott Diagnostics) instruments. The ratio of observed and expected analog concentrations was converted to a percent.[br]Results: The Modular E170 and Liaison assays exhibited no cross-reactivity at any concentration with either analog. Cross-reactivity of 1000 [micro]IU/mL insulin detemir exceeded the upper limit of detection of all remaining assays except Immulite 2000 (25%). Conversely, 1000 [micro]IU/mL insulin glulisine exhibited nominal cross-reactivity (maximum 14% by UniCel DxI and Architect i2000). Insulin detemir concentrations from 30 to 300 [micro]IU/mL cross-reacted an average of 17% by UniCel DxI, 52% by Advia Centaur and 22% by Immulite 2000. Insulin glulisine cross-reactivity at these concentrations averaged 6% by UniCel DxI, [lt]3% by Advia Centaur and 8% by Immulite 2000. The most significant cross-reactivity was observed by the Architect i2000 (insulin detemir averaged 101%, with 300 [micro]IU/mL exceeding the upper limit of detection; insulin glulisine averaged 21%).[br]Discussion: This data highlights how exogenous insulins may be detected by, and thus confound the interpretation of, certain insulin immunoassay results. For those assays exhibiting cross-reactivity, insulin detemir was detected at higher percentages than insulin glulisine. All assays utilize two-site (sandwich) antibody detection, thus epitope structure is critical for recognition. Modifications to epitope sites specific to individual analogs may be responsible for the differences observed. Understanding cross-reactivity of automated immunoassays with recombinant analogs is useful for clinical management and diagnosis in patients administering exogenous insulins.[br][br]Sources of Research Support: ARUP Institute for Clinical and Experimental Pathology.[br][br]Nothing to Disclose: SDM, WEO, SLL, SPW, WLR, JAS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2217 54 277 SAT-174 PO11-02 Saturday 216 2012


217 ENDO12L_SAT-175 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) The Impact of Type 2 DM in Mortality of Hospitalized Female Cancer Patients in Taiwan Jen-Der Lin, Wen-Ko Chiou, Feng-Hsuan Liu, Jawl-Shan Hwang, Wei-Ying Chou Chang Gung Memorial Hospital, Chang Gung University, Taoyuan Hsien, Taiwan; Chang Gung University, Taoyuan Hsien, Taiwan [bold]Aim[/bold] The goal of this study was to determine the relationships between diabetes mellitus (DM), specific cancer histologic types (histotypes), age, and mortality in hospitalized women.[br][bold]Methods[/bold] A total of 67,660 females with a mean age 62.76 [plusmn] 14.48 years admitted to the Chang Gung Medical Center in Linkou, Taiwan were diagnosed with cancer or DM. These patients were categorized into the following three groups: patients with cancer, but without DM (group A); patients with cancer and DM (group B); and patients with DM, but without cancer (group C). The disease states of the patients were identified according to the diagnostic codes of the International Classification of Disease-9.[br][bold]Result [/bold]Of the study subjects, 37,204 (54.98 %) were diagnosed with DM. as follows: groups A, n=30,456; group B, n=5,992; and group C, n=31,212. Group B patients had a high frequency of pancreas (34.06%), liver (27.25%), and renal (23.19%) cancers, and a low frequency of thyroid (8.09%), breast (9.30%), and ovarian (12.34%) cancers. Of the 36,448 cancer patients, 2,906 (7.97%) died. The mortality rate in group B (10.46%) was significantly higher than group A and group C. The percentage of DM in patients with different cancers had a negative association with the DM mortality rate/non-DM mortality rate of the corresponding cancer patients.[br][bold]Conclusions[/bold] DM affects the incidence and therapeutic outcomes in female cancer patients. The impact of DM on the mortality rate of specific cancers was negatively associated with the study case of the specific cancer.[br][br]Nothing to Disclose: J-DL, W-KC, F-HL, J-SH, W-YC 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 15 54 278 SAT-175 PO11-02 Saturday 217 2012


218 ENDO12L_SAT-176 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Glycemic Variability as Predictor of Clinical Outcome in Insulin-Treated Patients with Type 2 Diabetes Farnoosh Farrokhi, Dawn Smiley, Francisco J Pasquel, Limin Peng, Christopher Newton, Guillermo E Umpierrez Emory University School of Medicine, Atlanta, GA; Rollins School of Public Health, Atlanta, GA Glycemic variability (GV) has been shown to be an independent predictor of mortality in critically ill patients; however, the impact of GV in non-critical patients with T2DM is not known. Accordingly, we determined the ability of GV in predicting hospital complications in a prospective, randomized trial of insulin-treated medical and surgical patients with T2DM. A total of 279 patients with a BG of 140-400 mg/dl were randomized to basal bolus (n=146, age: 59[plusmn]11 yr, admission BG: 210[plusmn]88 mg/dl, A1C: 8.7[plusmn]2.5%, [plusmn]SD), or basal plus supplements (n=133, age: 59[plusmn]13 yr, admission BG: 207[plusmn]83 mg/dl, A1C: 8.3[plusmn]2.3%, [plusmn]SD). Patients in basal bolus were started at 0.5 U/kg, given half as glargine once daily and half as glulisine before meals. Basal Plus received 0.25 U/kg of glargine once daily plus correction doses of glulisine before meals for BG[gt]140 mg/dl. GV was calculated from all BG values using mean delta daily BG and mean standard deviation (SD).[br]Results: Basal plus regimen resulted in similar glycemic control as basal bolus group. Mean daily BG and rate of hypoglycemia after day 1 were 156[plusmn]36 mg/dl and 16% in the basal bolus and 163[plusmn]37 mg/dl and 13% in basal plus group (p=NS). There were no differences in length of stay or in-hospital complications between treatment groups (p=NS). There were no differences in GV between basal bolus and basal plus groups measured by delta or SD (72.5[plusmn]36 vs. 69.3[plusmn]34 mg/dl, and 38.5[plusmn]18 vs. 37.1[plusmn]16 mg/dl, both, p=NS). When GV was assessed in medical and surgical patients, we observed that surgical patients[apos] GV was higher with basal bolus than basal plus (delta: 74.9[plusmn]41 vs. 60.3[plusmn]32 mg/dl, SD: 38.2[plusmn]17 vs. 31.4[plusmn]14 mg/dl, both p=0.02), but there were no differences in GV in medical patients (delta: 70.7[plusmn]32 vs. 76.0[plusmn]34 mg/dl; SD: 38.7[plusmn]17 vs. 41.4[plusmn]16 mg/dl respectively, p=NS). Adopting median GV measures as a cut off, patients with high GV tend to have a higher risk of hypoglycemia compared to those with low GV (SD: 17% vs 7%, p=0.005, delta: 18% vs. 6%, p[lt]0.001). GV was associated with hypoglycemia (p=0.02), but no association was found between GV and hospital complications (p=NS).[br]In summary, treatment with glargine once daily plus correction doses of glulisine before meals resulted in similar glycemic variability, mean daily blood glucose and frequency of hypoglycemia compared to a basal bolus regimen. Higher glycemic variability was associated with increased risk of hypoglycemia but not with hospital complications.[br][br]Disclosures: GEU: Researcher, Sanofi-Aventis. Nothing to Disclose: FF, DS, FJP, LP, CN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2054 54 279 SAT-176 PO11-02 Saturday 218 2012


219 ENDO12L_SAT-177 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) The Impact of Hyperglycemia on Clinical Outcome in Elderly Patients with and without Diabetes in Acute Psychiatric Units: A Multicenter Observational Study Farnoosh Farrokhi, Powell Winter, Isabel Anzola, Pedram Aram, Francisco Pasquel, Shadi Sadeghi-Yarandi, Shoheera Khaliqdina, Charles Gillespie, Limin Peng, Dawn Smiley, Christopher Newton, Guillermo Umpierrez Emory University School of Medicine, Atlanta, GA; Emory University School of Medicine, Atlanta, GA; Rollins School of Public Health, Atlanta, GA The association between acute psychosis, use of antipsychotic therapy and diabetes (DM) is well established; however, the effect of antipsychotic agents on glycemic control and the impact of hyperglycemia on clinical outcome in elderly patients with acute psychiatric illnesses are not known. Accordingly, we analyzed the association between DM and hyperglycemia (BG[gt]140 mg/dl) and clinical outcome in 407 elderly ([gt]60 yrs) subjects (age 77[plusmn]9 yrs, 63% female, BMI 26[plusmn]7 kg/m[sup]2[/sup]) admitted to 2 acute psychiatric units between 1/1/2008 to 12/31/2008. The most common admission diagnoses were acute psychosis (33%), depression (29%), suicidal ideation/attempt (15%) and substance abuse/delirium (23%). A history of DM was present in 127 (31%) patients and 23 (6%) had newly diagnosed hyperglycemia on admission. Compared to nonDM patients, DM had higher BMI (29[plusmn]9 vs 25[plusmn]6 kg/m[sup]2[/sup], p[lt]0.001) and comorbidities: hypertension (87% vs 73%), dyslipidemia (59% vs 38%), and coronary artery disease (31% vs 18%), all p[le]0.002. The admission BG in patients with DM and hyperglycemia (157[plusmn]66 and 162[plusmn]19 mg/dl) was higher than in patients with normoglycemia (101[plusmn]16 mg/dl, p[lt]0.001). There were no differences on admission or daily hospital BG among DM and hyperglycemia patients treated with typical (163[plusmn]67 and 149[plusmn]39 mg/dl), atypical (156[plusmn]60 and 142[plusmn]36 mg/dl) or combination of antipsychotic agents (151[plusmn]75 and 137[plusmn]44 mg/dl), all p=NS. DM patients were treated with diet (9 %), sliding scale regular insulin (SSI, 9 %), oral antidiabetic drugs (OAD) alone (18 %) or in combination with insulin (43%), or basal insulin (22%). A total of 37% had [ge]1 episode of BG [lt]70 mg/dl. Among those with hypoglycemia, 20% were treated with OAD, 44% with OAD and insulin, 31% basal, and 5% with SSI alone. There were no differences in length of stay (LOS) or complications between patients with hyperglycemia or DM compared to those with normoglycemia.[br]In summary, hyperglycemia is a common finding in elderly patients with and without diabetes in acute psychiatric units. Our results indicate that a history of hyperglycemia and DM are not associated with higher risk of hospital complications compared to patients with normoglycemia. In addition, we found no differences in BG concentrations among patients treated with typical, atypical or with combination of antipsychotic agents. Randomized studies are needed to determine the optimal approach to manage hyperglycemia and DM in acute psychiatric disorders.[br][br]Nothing to Disclose: FF, PW, IA, PA, FP, SS-Y, SK, CG, LP, DS, CN, GU 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1922 54 280 SAT-177 PO11-02 Saturday 219 2012


220 ENDO12L_SAT-178 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Serum Sodium Level on Admission Was a Predictor for Mortality in Hyperglycemic Crises: A 5-Year Study in Thailand Pimjai Anthanont, Thana Khawcharoenporn, Thipaporn Tharavanij Faculty of Medicine Thammasat University, Pathum Thani, Thailand; Faculty of Medicine Thammasat University, Pathum Thani, Thailand [bold]Background:[/bold] Hyperglycemic crises, diabetic ketoacidosis (DKA) and hyperosmolar hyperglycemic state (HHS), are the most serious acute complications of diabetes mellitus (DM) which result in significant morbidity and mortality. However, existing and recently updated data on incidence and outcomes of hyperglycemic crises in Southeast Asia are lacking.[br][bold]Methods: [/bold]A retrospective study of DKA and HHS in adults with type 1 or type 2 DM admitted to Thammasat Hospital between 2006 and 2010 was performed. Incidences, precipitating causes, clinical and laboratory characteristics and treatment outcomes of these hyperglycemic crises were obtained via medical record review. Multivariate logistic regression analysis was used to determine predictors for mortality.[br][bold]Results: [/bold]Eighty-three patients were eligible and included. The mean age was 54.9[plusmn]17.7 years old. Most subjects had type 2 DM (85.5%). The 5-year incidence of hyperglycemic crises was 7.46%. Diabetic ketoacidosis occurred more frequently than HHS (4.67% VS 1.71%). During the hyperglycemic episodes, the mean plasma glucose on admission was 741.3[plusmn]320.8 mg/dL. Infections were the most common precipitating factor [61/83 (73.5%)], followed by non-compliance with treatments [35/83 (42.2%)]. Treatment complications included recurrent hyperglycemia (69.9%), hypokalemia (48.2%), hypernatremia (21.7%) and hypoglycemia (15.7%). The overall mortality rate of hyperglycemic crises was 8.4% (5.8% in DKA, 15.8% in HHS and 8.3% in the overlap of both conditions). The most common causes of death were infections [5/7(71.4%)]. By multivariate analysis, serum sodium level on admission was independently associated with mortality (adjusted odd ratio 1.08, 95% CI 1.01-1.16, [italic]P[/italic]= 0.03).[br][bold]Conclusion:[/bold] Hyperglycemic crises were not uncommon in our setting. Diabetic ketoacidosis occurred more frequently but had a lower mortality rate than HHS. Complications from hyperglycemic crisis treatment could be prevented by close monitoring, while high serum sodium level on admission was a predictor for mortality. Strategies to prevent infections and improve treatment compliance are needed to reduce the incidence of hyperglycemic crises among patients with DM.[br][br]Nothing to Disclose: PA, TK, TT 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 304 54 281 SAT-178 PO11-02 Saturday 220 2012


221 ENDO12L_SAT-179 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Time to Capillary Blood Glucose Retest in Hypoglycemic Hospitalized Patients Eberta Tan, Richard Chen, Li-Wei Cho, Jen-Min NG Changi General Hospital, Singapore, Singapore Background: Hypoglycemia is common in hospitalised patients with diabetes mellitus due to concomitant illnesses, use of medical therapies or disruption of nutritional intake.(1) Hypoglycemia in hospital can lead to neurological damage, sudden cardiac events and has been associated with higher mortality within 1 year.(2) Prompt recognition and reversal of hypoglycaemia are paramount to prevent such undesirable outcomes. The development of an effective hypoglycaemia prevention and management program requires a good understanding of event patterns in the hospital.[br]Aim: This study intends to assess the time to a capillary blood glucose retest in patients found to be hypoglycemic (defined by capillary blood glucose reading [le]63mg/dl) and if it is affected by whether the patient is in a medical or surgical ward.[br]Methods: A retrospective cross-sectional analysis of all patients in the general ward and intensive care units from 1 to 30 November 2011 was conducted. All patients with capillary blood glucose [le]63mg/dl were identified for analysis. The time from the first capillary blood glucose of [le]63mg/dl to the next biochemical reassessment via capillary blood glucose (time to CGR) was determined for each event. Data showing a time to CGR of more than 2 standard deviations from the mean was excluded from analysis. We analysed time to CGR by ward location. Results are presented as Mean[plusmn]SD and time differences were compared using the two sample T-test.[br]Results: 401 patients were identified for analysis (406 surgical, 671 medical). There were a total of 1077 hypoglycemic events. Collectively, the mean time to CGR was 105[plusmn]86 minutes. There was a significant difference in time to CGR between patients who developed hypoglycaemia in surgical wards and medical wards (117[plusmn]102 versus 98[plusmn]74) (p[lt]0.05).[br]Conclusion: There were more patients who developed hypoglycemia in medical wards compared to surgical wards. Patients who developed hypoglycemia in medical wards were more likely to be retested sooner for biochemical correction of the low blood sugar as compared to patients admitted to surgical wards. Postulated reasons for this include a lack of familiarity with managing diabetes cases and different workloads. Despite these differences, the overall time to correction of hypoglycaemia was still less than ideal. Due to the high prevalence of hypoglycaemia in the acute care setting, it is important to ensure that all hospital care providers are competent in basic diabetes management.[br][br](1) Varghese P, Gleason V, Sorokin R, Senholzi C, Jabbour S, Gottlieb JE. Hypoglycemia in hospitalized patients treated with antihyperglycemic agents. J Hosp Med 2007;2(4):234-40. (2) Turchin A, Matheny ME, Shubina M, Scanlon JV, Greenwood B, Pendergrass ML. Hypoglycemia and clinical outcomes in patients with diabetes hospitalized in the general ward. Diabetes Care 2009;32:1153[ndash]1157.[br][br]Nothing to Disclose: ET, RC, L-WC, J-MN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1891 54 282 SAT-179 PO11-02 Saturday 221 2012


222 ENDO12L_SAT-180 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Survey of Barriers to Effective Inpatient Glucose Control Amita Kathuria, John N Clore, Linda Thurby-Hay Virginia Commonwealth University, Richmond, VA Hyperglycemia in hospitalized patients is a common problem, often contributing to health care costs and outcomes. However, management of hyperglycemia in the hospitalized patient remains a complex matter with multiple factors creating a dynamic state adversely impacting effective glycemic control.[br]The present study was designed to assess current site-specific strategies for inpatient glucose management as determined by a national survey of endocrine fellows.[br]A web-based survey was created using the REDCAP instrument. The survey was then distributed electronically to fellows in endocrinology identified via a listing from the Endocrine Fellows[apos] Foundation, as well as through Association of Program Directors in Endocrinology and Metabolism.[br]A total of 206 responses were received, which is estimated to be about half of the endocrine fellows across the nation. Approximately 53.8% of responses were from large hospitals with 500+ beds. 54.8% of the respondents identified a management protocol at their institution, whereas the remainder (45.1%) reported no management protocol at all. Of those that do not have a standing hospital protocol, strategies for glycemic control included: basal/bolus insulin (61.3%), continuation of home regimen (22.6%), or sliding scale insulin (13.9%). The vast majority of respondents (78.1%) indicated that decisions regarding inpatient glycemic management were heavily dependent upon individual clinical assessment (clinical presentation, prior glycemic control, medical adherence, physician preference). In assessing various barriers impacting inpatient hyperglycemia management, the major impediment appeared to be fear of hypoglycemia, followed by constraints related to timing of meals and procedures in relation to insulin administration. Additionally, while 69.9% of respondents reported obtaining a hemoglobin A1c on inpatients who had not had a recent one in the last 3 months, 30.1% reported no hemoglobin A1c was obtained.[br]In conclusion, the present data suggest that a structured in-hospital glycemic strategy is infrequent. A national randomized control trial is sorely needed and would address a critical factor in hospital outcomes.[br][br]Sources of Research Support: UL 1RR031990.[br][br]Nothing to Disclose: AK, JNC, LT-H 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2311 54 283 SAT-180 PO11-02 Saturday 222 2012


223 ENDO12L_SAT-181 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Frequency of Autoimmune Diseases in Children and Adolescents with Type 1 Diabetes Paulo Cesar Alves Silva, Genoir Simoni, Edson Cechinel, Rose Marie Muller Linhares, Juliana Sande Lee, Ana Carolina Salerno Del Menezzi Tessari, Marilza Leal Nascimento Hospital Infantil Joana de Gusm[atilde]o, Florian[oacute]polis, Brazil; Hospital Universit[aacute]rio da Universidade Federal de Santa Catarina, Florian[oacute]polis, Brazil Introduction: Children and adolescents with type 1 diabetes mellitus (DM1) are at increased risk of developing other autoimmune diseases. Hashimoto[apos]s thyroiditis and celiac disease are the most common disorders found in these patients.[br]Objective: To evaluate the frequency of other autoimmune diseases in outpatients with DM1 treated at Hospital Infantil Joana de Gusm[atilde]o, a reference center in South Brazil.[br]Method: Retrospective study of medical records of 290 patients with DM1 to evaluate the presence of other autoimmune diseases, and this data was analyzed by descriptive statistics.[br]Results: The mean age was 7.4 years old, 52.4% were male. Other autoimmune diseases were observed in 16.5% of cases. The most frequent was Hashimoto[apos]s thyroiditis with 9.3%, followed by celiac disease with 2.4%. There was one case of each of the following diseases: Graves[apos] disease, psoriasis and vitiligo.[br]Conclusion: The high frequency of autoimmune diseases associated with DM1 emphasizes the importance of attention to the signs and symptoms of these diseases and screening for adequate control of these patients.[br][br]Nothing to Disclose: PCAS, GS, EC, RMML, JSL, ACSDMT, MLN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 336 54 284 SAT-181 PO11-02 Saturday 223 2012


224 ENDO12L_SAT-182 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Hemoglobin A1c: The New Gold Standard in Type 2 DM Screening? Dawn M Weiler, Leonie Sutherland, Shawn R Simonson, Jonathan Glogowski Boise State University, Boise, ID; Boise State University, Boise, ID; Duke Children[apos]s Hospital [amp] Medical Center, Durham, NC [bold]Purpose:[/bold] Compare traditional fasting plasma glucose (FPG) and the recently published hemoglobin A1c (HbA1c) diagnostic criteria in screening for type 2 diabetes in a population of Latino adults.[br][bold]Background:[/bold] Diabetes is one of the fastest growing disease classifications within the United States; especially among the Latino population. Review of the literature reveals that up to 60% of individuals have micro and macro-vascular disease related complications present at the time of type 2 diabetes diagnosis. Additionally, it is estimated that for many the disease has been present for as long as seven years before formal diagnosis. Early diagnosis and effective treatment significantly reduces the long-term complications and costs often associated with diabetes. These numbers validate that aggressive, cost effective, and efficient screening methods are imperative for improved health outcomes.[br][bold]Methods:[/bold] A descriptive study was carried out with 225 Latino adults (female n=132; male n=93) ranging in age from 18-75 years (m=43). Demographic, health behavior (Health Promoting Lifestyle Profile II), anthropometric (height, mass, waist-hip ratio, Jackson-Pollock 3 site skinfolds), and fasting venous blood samples (lipid, insulin, glucose, HbA1c) were collected. Diabetes status was assessed utilizing traditional FPG and newly released HbA1c guidelines. Participants self-reporting a known diabetes diagnosis (n=27) were excluded from the analysis.[br][bold]Results:[/bold] Two and a half percent (n=5) met diabetes diagnosis criteria utilizing FPG compared to 7.6% (n=15) based on HbA1c criteria. Fifty-seven participants (28.9%) had HbA1cs between 5.6 and 6.4 of whom 56% (n=32) had normal FPG values. Of those with HbA1cs between 6.0 and 6.4 (n=22), 32% (n=7) had normal FPG levels. In comparison, of the 27% (n=53) who met [ldquo]impaired fasting glucose[rdquo] criteria, 10.2% (n=20) had normal (5.5 and lower) HbA1c values.[br][bold]Conclusions:[/bold] In this population, FPG had a 25% false negative and a 37.7% false positive screening rate when compared to HbA1c values. Fasting plasma glucose also failed to identify 66.7% of individuals who met HbA1c criteria for diabetes diagnosis. These results imply that utilizing HbA1c for type 2 diabetes screening is a more sensitive diagnostic tool and should become the new [ldquo]gold standard[rdquo] due to its increased accuracy, efficiency and lower overall cost (personal and financial) compared to traditional FPG methods.[br][br]Sources of Research Support: Boise State University School of Nursing Faculty Research Grant; Boise State University College of Health Sciences Mini-Research Grant; Boise State University Office of Sponsored Program Grant.[br][br]Nothing to Disclose: DMW, LS, SRS, JG 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 625 54 285 SAT-182 PO11-02 Saturday 224 2012


225 ENDO12L_SAT-183 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) The Well-Being and Treatment Satisfaction of Diabetic Patients in General Hospital in Korea Ie Byung Park, Sin Na Lee, Young Sil Eom, Sei-Hyun Kim, Ki Young Lee, Sihoon Lee, Yeun Sun Kim Gachon University Gil Hospital, Incheon, Republic of Korea [bold]Backround[/bold][br]The ambulatory diabetes care system in Korea is different from those in America. In Korea, the number of patients visited per day is about 80-160 diabetic patients, and the treatment time per patient is so short(3-5 minutes). We investigated whether the well-being and treatment satisfaction of diabetes in general hospital in Korea are different from those in westernized ambulatory care. We analysed relationship between well-being and treatment satisfaction and affecting factors on both.[br][bold]Methods[/bold][br]This study subjects were 382 diabetics, 20-80 years old, who visited the Department of Endocrinology at Gachon University Gil Hospital. We evaluated the well-being using 12-item well-being questionnaire(WBQ-12) consisting of three 4-item subscales(Negative Well-Being(NWB), Energy(ENE), and Positive Well-Being(PWB), and 12-item overall scale General Well-Being(GWB)). We evaluated the diabetes treatment satisfaction using Diabetes Treatment Satisfaction Questionnaire(DTSQ).[br][bold]Results[/bold][br]Cronbach[apos]s alpha for WBQ-12 and DTSQ were 0.819 and 0.819, respectively. The higher total WBQ-12(GWB) score was significantly associated with women(r=0.20), educational status(r=0.15), income(r=0.30), satisfaction for waiting time(r=0.20), exercise compliance(r=0.21), total pill number(r=-0.16) and HbA1c level(r=-0.15). The NWB, ENE, and GWB score in men is significantly different from those in women(2.16[plusmn]2.3 vs 3.89[plusmn]3.3; 6.69[plusmn]2.5 vs 5.90[plusmn]2.8; 23.25[plusmn]6.5 vs 20.56[plusmn]7.0, respectively). ENE score in men was lower than that in women(5.56[plusmn]2.8 vs 6.56[plusmn]2.6, p=0.01) and ENE score in insulin-user was lower than non-user(5.56[plusmn]2.8 vs 6.56[plusmn]2.6. p=0.01).[br]The higher DTSQ score was significantly associated with age(r=0.24), education(r=-0.12), waiting time for treatment(r=0.34), satisfaction of treatment time(r=0.60), diet compliance(r=0.42) and exercise compliance(r=0.35).[br]In multiple regression analysis, GWB score was significantly associated with income and satisfaction of treatment time and DTSQ score was significantly associated with diabetes duration, satisfaction of treatment time and diet compliance.[br][bold]Conclusion[/bold][br]Our results show that some affecting factors on the well-being and diabetes treatment satisfaction in ambulatory diabetic patients in Korea are similar to those in westernized outpatient clinics. But the environmental conditions such as treatment time and waiting time for treatment are likely to be important factors affecting on those scores compared with those in Europe and America.[br][br]Nothing to Disclose: IBP, SNL, YSE, S-HK, KYL, SL, YSK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1170 54 286 SAT-183 PO11-02 Saturday 225 2012


226 ENDO12L_SAT-184 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Gestational Diabetes in Assisted Reproductive Techniques and Spontaneous Pregnancy Radhika Jindal, Nitin Gupta, Mukul Gupta, Mohammad Asim Siddiqui, Subhash Kumar Wangnoo Indraprastha Apollo Hospital, New Delhi, India OBJECTIVE:[br]The aim of this study was to compare the maternal glycemia, weight gain and fetal outcomes in women diagnosed with Gestational Diabetes Mellitus (GDM) during pregnancy with Assisted Reproductive Techniques (ART) and spontaneous pregnancies.[br]METHODS:[br]Thirty-six women who had successful ART (irrespective of indication) and developed GDM during the course of their singleton pregnancy (ART group) and 37 non-IVF women with GDM (non-ART group) were included in the study after informed consent. They were matched according to age and pre-pregnancy BMI. None had any previous history of diabetes mellitus or GDM before their pregnancies. GDM was diagnosed as per the ADA criteria for diagnosis of GDM. The patients in both the groups received individualized care from the diabetes educators and the treating endocrinologists. We compared the maternal characteristics, course of pregnancy and neonatal outcome.[br]RESULTS:[br]The weight gain until the diagnosis of GDM in both non-ART and ART groups of women was not significantly different (9.51[plusmn]3.97 vs 10.0[plusmn]3.8kg, p=0.8 respectively). First trimester fasting glucose levels were found to be significantly higher in ART group (88[plusmn]15.2 vs 81[plusmn]10.3mg/dl, p=0.04). Second trimester oral glucose tolerance test (OGTT) results and glucose levels during GDM treatment did not differ between the groups. There were no significant changes in investigated fetal and neonatal parameters: 3rd trimester abdominal circumference, percentile and neonatal birth weight (3250[plusmn]641 vs 2900[plusmn]440g, p=0.22).[br]CONCLUSIONS:[br]GDM among women who underwent ART is associated with higher first trimester fasting glucose levels. We suggest that women undergoing ART be evaluated for dysglycemia at an earlier gestational age.[br][br]Nothing to Disclose: RJ, NG, MG, MAS, SKW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1460 54 287 SAT-184 PO11-02 Saturday 226 2012


227 ENDO12L_SAT-185 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Pre-IGT in Pregnancy: Comparison of HbA1c and Clinical Scoring in Patients with Gestational Diabetes (GDM) Mary Jane Yu Tanchee-Ngo, Leilani B Mercado-Asis University of Sto Tomas Hospital, Manila, Philippines Background: In the initial stage of Diabetes mellitus, the [beta]-cell compensates for the insulin resistance by increasing insulin secretion to maintain normoglycemia, which we termed pre-impaired glucose tolerance (pre-IGT state). The 2nd hour glucose is normal ([lt]140mg/dl) but with increased insulin levels ([gt] 30uIU/ml) after 75 grams oral glucose tolerance test (OGTT)1,2. Clinical scoring developed by us for non-pregnant subjects using risk factors of diabetes mellitus showed similar scores in normal and pre-IGT nonpregnant individuals implying that the latter is a subclinical condition2. Although our group demonstrated that a 2nd hr insulin level of 45 uIU after 75 grams OGTT done during the first trimester is predictive of development of GDM3, elucidation of pre-IGT among pregnant individuals with risk factors has yet to be done.[br]Objective: To identify pre-IGT among pregnant individuals with risk factors for GDM.[br]Methodology: This is an on-going retrospective cohort study of pregnant individuals 18-40 year-old seen at the Endocrine Clinic of our institution. Risk factors for diabetes were determined. All underwent Second-hour blood glucose and insulin after 75 g oral glucose tolerance test (OGTT) with glycosylated hemoglobin (HbA1c). They were then divided into 4 groups base on their glycemic status and insulin levels.: Group A (normoglycemia with normal insulin levels), Group B or the Pre-IGT group (normoglycemia with high insulin levels and Group C or the IGT and GDM group (hyperglycemia with high insulin levels). The clinical risk scoring of the normoglycemic group was then determined. The most frequent risk factor given the highest score and the less frequent risk factor given the lowest score. A p value equal or less than 0.05 was considered significant.[br]Results: A total of 11 out of 16 patients fulfilled the criteria. Thirty percent were found to have GDM with hyperinsulinemia whereas 70% were found to have pre-IGT. Although both insulin levels were increased, the value in the GDM group was not significantly different in the pre-IGT group (236 uIU vs. 121 uIU, p=0.053). The most frequent risk factors found in both groups is a family history of DM followed by an elevated BMI. There was no significant difference between the mean clinical risk scores of the two groups (p 0.62).[br]Conclusion: Preliminary observations showed pre-IGT exists among high-risk pregnant subjects whereas clinical risk scoring of GDM and pre-IGT patients did not vary.[br][br]1. Matawaran B, Mercado-Asis L. Comparison of Pancreatic Insulin Response to Hyperglycemia Among Filipino Subjects of Various Glycemic Status. Phil J Internal Medicine, 47: 25-30, Jan-Feb 2009. 2. Nisce I, Matawaran B, Mercado-Asis L. Clinical Risk Scoring is Not Useful in Screening Patients with Pre-IGT (Hyperinsulinemic State) : The Importance of 2nd-hour Insulin Measurement in OGTT Poster in PSEM 2011, EndoSoc USA 2011. 3. Quiambao D., Fonte, J., Mercado-Asis L., Matawaran B., AMORADO-SANTOS, F., Lejos, A., Cabatu, M., Decena, C., Dee, M., Gamilla, Z., Gutierrez, R., Hyperinsulinemia During the First Trimester as a Predictor in the Development of Gestational Diabetes Mellitus Among High Risk Pregnant Patients. 4. Standards of Medical Care in Diabetes-2011. January 2011 vol. 34 no. Supplement 1 S11-S61.[br][br]Nothing to Disclose: MJYT-N, LBM-A 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2252 54 288 SAT-185 PO11-02 Saturday 227 2012


228 ENDO12L_SAT-186 POSTER SESSION: Diabetes [amp] Clinical Care (1:30 PM-3:30 PM) Postpartum Women Have Less Insulin Resistance and Better Glucose Tolerance Compared with Pregnant and Non-Pregnant Women David Gustavo Garcia-Gutierrez, Ma Ludivina Robles-Osorio, Pablo Garcia-Solis, Juan Carlos Solis-S, Hebert Luis Hernandez-Montiel Facultad de Medicina, Universidad Aut[oacute]noma de Quer[eacute]taro, Quer[eacute]taro, Mexico INTRODUCTION:[br]Pregnancy is a period during which there are changes such as insulin resistance and more frequent glucose intolerance and gestational diabetes. The study of this conditions and its changes during postpartum are important due to the group of patients who develop important changes during pregnancy, such as glucose intolerance and gestational diabetes than are prognostic factors to develop glucose intolerance and type 2 diabetes in the future.[br]METHODS:[br]Cross sectional study, we studied 304 women, mean age was 26.8[plusmn]5.3 years old; 192 pregnant, 47 women were in postpartum (between 8 to 12 weeks after delivery), age 26.3[plusmn]5.3 years old; 65 non-pregnant women, mean age 28.7[plusmn]5-8 years old. Glucose and insulin were measured on fasting and 1, 2 and 3 hrs. after an oral glucose load.[br]RESULTS:[br]We investigated the weight before pregnancy and current weight in no pregnant women, we found 30.6% overweight women and 15.1% obesity, both diagnosis were present in 45.7%. According to the Institute of Medicine 21.9% of pregnant women had increased more weight per trimester than recommended.[br]We found that fasting glucose was different between first [italic]vs[/italic] second (p[lt]0.001) and third trimesters (p=0.012), a significant difference was found only in the first hour between first and third trimester (p=0.05), but not difference was found within the other groups regarding to glucose and insulin levels. When we compared the glucose area under the curve (AUC) we found a significant difference between the groups (p[lt]0.001); being the higher glucose and insulin AUC within the third trimester pregnant women, and the lowest AUC for glucose and insulin in the postpartum group, even compared with the non-pregnant women.[br]Between the pregnant women there was no difference within the different trimesters on insulin and glucose levels.[br]CONCLUSIONS:[br]We found in a population with a high prevalence of diabetes and metabolic syndrome, a significant difference between the pregnant women and the postpartum (8 to 12 weeks after delivery), with glucose and insulin lower levels even comparing with non-pregnant women, this finding can be related to metabolic changes given after pregnancy, after a state of significant insulin resistance and a increase in glucose levels during the OGTT. The study of this changes can be of physiological interest and also in order to identify which factors also can contribute to a good recovery after pregnancy and contribute to develop type 2 diabetes in the future.[br][br]Sources of Research Support: Programa para el mejoramiento del profesorado, secretaria de educacion publica. (PROMEP).[br][br]Nothing to Disclose: DGG-G, MLR-O, PG-S, JCS-S, HLH-M 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2347 54 289 SAT-186 PO11-02 Saturday 228 2012


229 ENDO12L_SAT-187 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Hsp60, a Leptin-Induced Mitochondrial Chaperone, Impacts on Central Insulin Signaling Andre Kleinridders, Siegfried Ussar, Jane Hvarregaard Christensen, Marcelo Mori, Peter Bross, C Ronald Kahn Joslin Diabetes Center, Boston, MA; Aarhus University Hospital and Aarhus University, Aarhus, Denmark; Aarhus University, Aarhus, Denmark The adipocyte-derived hormone leptin is secreted after food consumption and acts on the hypothalamus to control energy metabolism by increasing energy expenditure and promoting satiety. Heat shock protein (Hsp) 60, a nuclear encoded mitochondrial chaperone, is crucial for proper folding of mitochondrial proteins and plays a pivotal role in maintaining mitochondrial function and cell homeostasis. Here we show a 50% decrease in the level of Hsp60 in the hypothalamus at both the mRNA and protein level of leptin-resistant db/db mice and leptin-deficient ob/ob mice. This is due to direct regulation of Hsp60 expression by leptin via activation of STAT3 as overexpression of a dominant negative STAT3 abolishes leptin[apos]s ability to induce Hsp60 expression. Consistent with this, intraperitoneal injection of leptin in normal mice led to an upregulation of Hsp60 in the hypothalamus [italic]in vivo[/italic].[br]As Hsp60 is crucial for the integrity of the mitochondrial matrix, we assessed mitochondrial respiration using isolated mitochondria from hypothalamic samples of db/db mice in a Seahorse Flux Analyzer. When compared to control mitochondria, mitochondria from db/db mice exhibited a defect in the electron transport chain activity. This defect was preserved [italic]in vitro[/italic] by lentiviral knockdown of Hsp60 in the hypothalamic cell line N25/2. This was associated with decreased levels of multiple protein subunits of complexes I, II, III and V. As a consequence of this, reactive oxygen species production was increased as assessed by measuring lipid peroxidation. In addition, the mitochondrial dysfunction produced by knockdown of Hsp60 [italic]in vitro[/italic] led to activation of c-Jun kinase pathway, increased IRS1 Ser307 phosphorylation, and resistance to normal insulin signaling, which could be restored by inhibiting reactive oxygen species using N-acetyl-L-cysteine. Consistent with the insulin resistance observed [italic]in vitro[/italic], mice with a haploinsufficiency of Hsp60 showed insulin resistance in the hypothalamus due to mitochondrial dysfunction specifically in the hypothalamus.[br]Together these data show that leptin plays an important role in regulating mitochondrial function in the hypothalamus by regulating the mitochondrial chaperone Hsp60 which influences insulin sensitivity in the hypothalamus. These data provide a novel pathway of leptin/insulin crosstalk in the central nervous system which may impact on hypothalamic control of energy homeostasis in obesity and insulin resistant states.[br][br]Nothing to Disclose: AK, SU, JHC, MM, PB, CRK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 930 55 290 SAT-187 PO14-02 Saturday 229 2012


230 ENDO12L_SAT-188 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Insulin Action on Brain Affects Serotonergic and Dopaminergic Neurotransmission and Mediates Behavioral Changes with Aging Andre Kleinridders, Laura Cappellucci, Sara Vienberg, Emmanuel N Pothos, C Ronald Kahn Joslin Diabetes Center, Boston, MA; Tufts University School of Medicine, Boston, MA Insulin is secreted from the pancreas in response to rise in blood glucose and acts on peripheral tissues to control glucose and lipid metabolism. Insulin also acts on neurons in the brain in control of energy homeostasis and feeding behavior. We have previously shown that mice with a disruption of the insulin receptor in the brain (NIRKO mice) have increased food intake leading to some features of metabolic syndrome. In addition, these mice develop hypothalamic hypogonadism, but have no detectable behavioral abnormalities at a young age. Here we show that NIRKO mice subjected to aging develop significant alterations in the dopaminergic and serotonergic systems leading to increased anxiety and decreased exploratory drive, as well as impaired motor coordination.[br]To test anxiety and explorative behavior, aging NIRKO mice were subjected to mouse tail suspension, forced swimming and adapted open field tests. By 17 months of age, all of these tests showed significantly increased periods of immobility indicative of increased anxiety, decreased explorative drive and depressive-like behavior. These changes were not apparent in the 11 month old mice. Assessment of motor skills in the older cohort using the balance beam also revealed impairment of NIRKO mice in motor balance and coordination.[br]To understand the cause of these phenomena, we examined neurotransmitter receptor expression and neurotransmitter release in control and NIRKO mice with aging. Dopamine release in the mesolimbic system assessed using carbon fiber amperometry displayed a significant decrease in amplitude and width of the dopamine peak, as well as a decreased half-life of dopamine in the nucleus accumbens, which was independent of neurotransmitter reuptake and not altered by the reuptake inhibitor nomifensine. This phenotype was already observed at age of 8 month where no behavioural abnormalities were detected. Furthermore, even in young (3 month old) NIRKO mice, we could observe decreased expression of some serotonin receptors (Htr1B, Htr2A and Htr7) in the hippocampus of NIRKO mice. Insulin treatment of cultured neuronal cells in vitro showed that these serotonin receptors were directly induced by insulin treatment.[br]Thus, taken together these data show that insulin signaling can act directly on neurons to alter both serotonergic and dopaminergic signaling. While initially these alterations are clinically silent, with aging, these alterations results in changes in behavior and motor skills.[br][br]Nothing to Disclose: AK, LC, SV, ENP, CRK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1998 55 291 SAT-188 PO14-02 Saturday 230 2012


231 ENDO12L_SAT-189 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Novel Role of Insulin and IGF-1 Signaling in Regulation of DNA Imprinting Jeremie Boucher, Marika Charalambous, Kim Zarse, Marcelo Mori, Michael Ristow, Anne Ferguson-Smith, C Ronald Kahn Joslin Diabetes Center, Boston, MA; University of Cambridge, Cambridge, UK; University of Jena, Jena, Germany Insulin and insulin like growth factor 1 (IGF-1) control many biological processes including cellular metabolism, proliferation, differentiation and apoptosis. Insulin and IGF-1 receptors (IR and IGF1R) also create novel signals in the unoccupied state, such that preadipocytes lacking both insulin and IGF-1 receptors are resistant to apoptosis. Double knockout cells (DKO) also exhibit major changes in miRNA regulation. Among the 371 miRNAs detectable in preadipocytes, 134 were differentially expressed between control and DKO cells (p[lt]0.05) and 89 of them were down-regulated in DKO cells compared to controls, in many cases by 10- to [gt]1000-fold. This occurred without any changes in expression of proteins involved in miRNA maturation and processing such as Dicer, Drosha, DGCR8, TRBP, Ago2 and Exportin 5. Most of these down-regulated miRNAs reside in two miRNA clusters on mouse chromosomes 2 and 12 previously shown to be regulated by imprinting. Inactivation of the IR alone in mouse embryonic fibroblasts also resulted in a down-regulation of many of these imprinted miRNAs. In DKO cells, the imprinted genes in these loci, Dlk1, Gtl2, Rtl1, Dio3 and Sfmbt2, whether normally maternally or paternally expressed, along with several imprinted genes on other chromosomes, such as IGF2, H19 or Phlda2 were highly down-regulated or undetectable, indicating generalized epigenetic misregulation of imprinted genes. Pyrosequencing of bisulfite-treated DNA from the DKO preadipocytes revealed increased methylation of several differentially methylated regions, such as the Dlk1-DMR, IG-DMR, Gtl2 promoter or the H19 DMD, which occurred without changes in expression in DNA methyl transferases 1, 3a or 3b. Treatment of DKO cells with a methyltransferase inhibitor partially reversed the changes in miRNA and gene expression. Increased methylation correlated with a [gt]70% decrease in expression of the transcription factor Zac1, which has been shown to regulate a large imprinted network. Thus, disruption of insulin and IGF-1 signaling in somatic cells can induce changes in the epigenome, causing a down-regulation in expression of imprinted genes, at least in part, due to down-regulation of Zac1 and increased DNA methylation.[br][br]Nothing to Disclose: JB, MC, KZ, MM, MR, AF-S, CRK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2189 55 292 SAT-189 PO14-02 Saturday 231 2012


232 ENDO12L_SAT-190 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Attenuation of High Glucose-Induced Oxidative Stress in Brain Pericytes by Ethoxyzolamide, a Mitochondrial Carbonic Anhydrase Inhibitor Tulin O Price, Nader Sheibani, Ping Patrick, Nuran Ercal, Gul N Shah Saint Louis University, St Louis, MO; Missouri University Science[amp]Technology, Rolla, MO; University of Wisconsin, Madison, WI Diabetes mellitus has strongly been associated with the imbalance between production of reactive oxygen species (ROS) and the antioxidant enzyme system. Overproduction of mitochondrial ROS results in oxidative stress which is augmented during diabetes in insulin-insensitive tissues such as brain. Pericytes (PC) in the microvasculature of brain are especially susceptible to oxidative stress. Mitochondrial carbonic anhydrases CA VA and CA VB (mCAs) regulate oxidative metabolism of glucose, and thus, may play an important role in the ROS generation and oxidative stress. We have recently shown that inhibition of mCAs reduces diabetes-induced oxidative stress in the mouse brain and rescues cerebral PC loss. Here we tested the hypothesis that pharmacological inhibition of mCAs attenuates high glucose-induced intracellular oxidative stress in brain PC.[br]We have isolated conditionally immortalized cerebral PC from transgenic Immorto mouse. These cells were analyzed for primary PC markers by fluorescence[ndash]activating cell sorting (FACS) and for the presence of mCAs by RT-PCR and Western blotting. These cells were grown in low glucose (LG, 5.7 mM) and high glucose (HG, 40.7 mM) for 6 days. The measures of oxidative stress were reduced glutathione (GSH), and activities of superoxide dismutase (SOD), glutathione peroxidase (GPx), and glutathione reductase (GR). For pharmacological inhibition of mCAs, ethoxyzolamide (ETZ) was added to the culture medium from day one of the experiment.[br]The PC expressed PDGFR-[beta], NG2, CD13, and [alpha]-SMA markers. PECAM-1, the endothelial cell marker was not expressed. These cells were successfully passaged and maintained in culture for several months without loss of these markers. Six days exposure to high glucose resulted in significant decrease in GSH (100[plusmn]5.4 LG, 39.95[plusmn]3.18 HG, p[lt]0.0007), SOD (100[plusmn]17.1 LG, 41.58[plusmn]7.44 HG, p[lt]0.03), and GR (100[plusmn]9.86 LG, 82.64[plusmn]1.49 HG, p[lt]0.05) and an increase in GPx (100[plusmn]13.02 LG, 130.3[plusmn]6.65 HG, p[lt]0.05) activities compared to low glucose indicating increased intracellular oxidative stress. Treatment of PC with ETZ significantly reduced high glucose-induced oxidative stress.[br]These results provide the first evidence that high glucose induces oxidative stress in cultured mouse brain PC, and pharmacologic inhibition of mCAs attenuates high glucose-induced oxidative stress.[br][br]Nothing to Disclose: TOP, NS, PP, NE, GNS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1804 55 293 SAT-190 PO14-02 Saturday 232 2012


233 ENDO12L_SAT-191 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Mitochondrial Carbonic Anhydrase Inhibition Protects Kidneys from Hyperglycemia-Induced Oxidative Damage in Diabetic Mice Uma Muthyala, Tulin Price, Ping Patrick, Gul Shah Saint Louis University, St Louis, MO [bold]Introduction[/bold]: Diabetes mellitus (DM) is the leading cause of end-stage renal disease.Increased mitochondrial oxidative stress (OxSt) mediated by reactive oxygen species (ROS) is implicated in hyperglycemia-induced kidney dysfunction.[br][bold]Objective[/bold]: We hypothesize that inhibition of mitochondrial carbonic anhydrases (mCA), regulators of oxidative metabolism of glucose, reduces OxSt in the kidneys and helps slow down the onset and progression of diabetic nephropathy (DN).[br][bold]Methods[/bold]: We used mCA double knockout (DKO) and diabetic mice to test our hypothesis. Diabetes was induced by streptozotocin and pharmacological inhibition of mCA was accomplished by CA inhibitor, topiramate(TOP), administered daily for 12 weeks. Urine samples were analyzed for albumin and creatinine. Kidneys were extracted for determination of OxSt.[br][bold]Results[/bold]: Diabetes-induced OxSt indicated by decreased reduced glutathione (GSH, 100 [plusmn] 9.22 control, 71.89 [plusmn] 3.82 DM, 107.1 [plusmn] 9.06 DM+TOP, p [lt] 0.02) and increased 4-hydroxy-2-trans-nonenal (HNE, 100 [plusmn] 17.66 control, 607.1 [plusmn] 176.6 DM, 290.8 [plusmn] 52.02 DM+TOP, p [lt]0.02) was reversed with TOP treatment. An elevated albumin/creatinine ratio in DM (100.93 [plusmn] 9.43 g/mg) was reduced upon TOP treatment (73.91 [plusmn] 5.43 g/mg, p[lt]0.05) as well. The superoxide dismutase(SOD) activity remained unchanged in DM and DM+TOP mice. The above results were supported by DKO data.[br][bold]Discussion[/bold]: The unchanged SOD activity in DM and DM+TOP mice suggests that mCA inhibition reduces OxSt and albuminuria by decreasing the production of ROS rather than elimination of ROS by increasing SOD activity.[br][bold]Conclusions[/bold]: Mitochondrial CA may provide a novel therapeutic target for slowing down the onset and progression of DN.[br][br](1)Brownlee M. The pathobiology of diabetic complications: a unifying mechanism. Diabetes 2005;54(6). (2)Saito Y, Kida H, Takeda S, Yoshimura M, Yokoyama H, Koshino Y, Hattori N. Mesangiolysis in diabetic glomeruli: its role in the formation of nodular lesions. Kidney Int. 1988 Sep;34(3):389-96. (3)Kriz W, Lemley KV. The role of the podocyte in glomerulosclerosis. Curr.Opin.Nephrol.Hypertens. 1999 Jul;8(4):489-97.[br][br]Nothing to Disclose: UM, TP, PP, GS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 160 55 294 SAT-191 PO14-02 Saturday 233 2012


234 ENDO12L_SAT-192 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) [D-Leu-4]-OB3, a Synthetic Peptide Amide with Leptin-Like Activity, Enhances the Effects of Orally Delivered Exenitide (Byetta[reg]) and Pramlinitide Acetate (Symlin[reg]) on Body Weight Gain and Glycemic Control in Leptin-Resistant Male C57BLK/6-m [italic]db/db[/italic] Mice Matthew C Leinung, Patricia Grasso Albany Medical College, Albany, NY In the present study, exenitide (Byetta[reg]) or pramlinitide acetate (Symlin[reg]), currently administered by subcutaneous injection, were reconstituted in Intravail[reg], a patented transmucosal absorption enhancing agent, in the absence or presence of [D-Leu-4]-OB3, and delivered orally by gavage to leptin-resistant male C57BLK/6-m [italic]db/db[/italic] mice twice daily for 14 days. Body weight gain, food and water intake, blood glucose, and serum insulin levels were measured. Mice receiving Intravail[reg] alone for 14 days were 21.1% heavier at the end of the test period, whereas oral delivery of Byetta[reg] or [D-Leu-4]-OB3 in Intravail[reg] resulted in significantly less body weight gain, 13.9% and 11.5%, respectively. Most worthy of note, mice receiving Byetta[reg] and [D-Leu-4]-OB3 were 2.2% lighter than they were at the beginning of the study. Similar results were seen with Symlin[reg]. In another study, mice receiving Symlin[reg] or [D-Leu-4]-OB3 in Intravail[reg] were 26.8% and 25.4% heavier, respectively, after 14 days, while Intravail[reg] treated control mice were 38.2% heavier than their initial body weight. Co-administration of Symlin[reg] and [D-Leu-4]-OB3 did not significantly enhance the effect of Symlin[reg] on body weight gain. Food intake was significantly reduced by Byetta[reg], Symlin[reg] and [D-leu-4]-OB3 alone, and co-delivery of Byetta[reg] or Symlin[reg] with [D-Leu-4]-OB3 did not induce any further reduction. Oral delivery of Byetta[reg] or Symlin[reg] in Intravail[reg] significantly lowered blood glucose levels by 20.4% and 22.0%, respectively, and co-delivery of Byetta[reg] or Symlin[reg] with [D-Leu-4]-OB3 further reduced blood glucose by 32.3% and 40.0% respectively, of initial levels. A dose-related increase in serum insulin was observed in response to increasing concentrations of Byetta[reg], and [D-Leu-4]-OB3 enhanced the insulin response to Byetta[reg] at all concentrations tested. No similar increases in serum insulin occurred in response to increasing concentrations of Symlin[reg]. The results of these studies indicate that (1) oral delivery of Byetta[reg] and Symlin[reg] is feasible; (2) the bioactivity of Byetta[reg] and Symlin[reg] is retained following oral delivery in Intravail[reg]; and (3) the effects of Byetta[reg] and Symlin[reg] on energy balance and glycemic control can be enhanced by co-administration with [D-Leu-4]-OB3, a synthetic peptide amide with leptin-like activity.[br][br]Sources of Research Support: Willard B. Warring Memorial Fund. Aegis Therapeutics, San Diego, CA.[br][br]Nothing to Disclose: MCL, PG 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 133 55 295 SAT-192 PO14-02 Saturday 234 2012


235 ENDO12L_SAT-193 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) CNS Vitamin D Improves Glucose but Not Insulin Tolerance in Diet-Induced Obesity Stephanie R Sisley, Randy J Seeley, Darleen A Sandoval Cincinnati Children[apos]s Hospital Medical Center, Cincinnati, OH; University of Cincinnati, Cincinnati, OH Vitamin D deficiency is associated with obesity and type 2 diabetes, although the mechanism is unknown. Multiple lines of evidence support the CNS activity of the NF[kappa]B pathway as a link in the glucose dysregulation accompanying diet-induced obesity. Given that vitamin D receptors are present in the hypothalamus, the key CNS region for weight and glucose regulation, we sought to determine if acute downregulation of this pathway in the hypothalamus with 1,25-hydroxyvitamin D, an IKK[beta] inhibitor in the NF[kappa]B pathway, decreases food intake or improves glucose tolerance. No effects on food intake were observed after direct administration of 100[mu]g 1,25-hydroxyvitamin D or vehicle (1[mu]l) into the third cerebral ventricle (i3vt) of diet-induced obese Long-Evans rats 60 minutes prior to the dark cycle. We then performed an i3vt injection of 1,25-hydroxyvitamin D 60 minutes prior to an intraperitoneal glucose tolerance test (ipGTT). I3vt 1,25-hydroxyvitamin D significantly decreased the glucose excursion 15 and 30 minutes after the glucose load compared to vehicle. Interestingly, no significant differences in glucose levels were seen during an intraperitoneal insulin tolerance test after i3vt injection of 1,25-hydroxyvitamin D compared to vehicle. We speculate that the positive ipGTT results are secondary to an improved first phase insulin response. Therefore, acute inhibition of NF[kappa]B by vitamin D, specifically within the CNS, improves glucose tolerance. However, the mechanism of improvement is not secondary to improved insulin sensitivity.[br][br]Sources of Research Support: NIH Grant 5T32ES010957-1 awarded to SRS; Fellows Development Research Grant Program in Diabetes, Obesity and Fat Cell Biology awarded to SRS.[br][br]Disclosures: DAS: Research Funding, Johnson & Johnson, Novo Nordisk, Pfizer, Inc; Speaker, Novo Nordisk. Nothing to Disclose: SRS, RJS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1953 55 296 SAT-193 PO14-02 Saturday 235 2012


236 ENDO12L_SAT-194 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Physiological Effect of Peripheral Serotonin on Glucose and Lipid Metabolism in Mice and Sheep Hitoshi Watanabe, Ryo Saito, Tatsuya Nakano, Hideyuki Takahashi, Yu Takahashi, Keisuke Sumiyoshi, Yuya Nagasawa, Shunsuke Terada, Natsumi Okada, Dian W Harjanti, Natsumi Sekiguchi, Hiroaki Sano, Kouichi Watanabe, Hisashi Aso Tohoku University, Sendai, Japan; Iwate University, Morioka, Japan [Objective][br]Serotonin is a monoaminergic neurotransmitter with activities that modulate central and peripheral functions. Serotonin affects food intake, sleep, anxiety, sexual behavior and mood in the central nervous system. On the other hand, the functions of serotonin in peripheral tissue have not yet been fully elucidated. Further, serotonin is thought not to be able to pass the blood-brain barrier. Thus, there are two independent serotonin systems: one in the central nervous system and the other in the periphery. Recently, peripheral serotonin is known to be associated with glucose metabolism mainly because of its regulation of the secretion of insulin in pancreatic [beta] cells. Additionally, the concentration of serotonin in the blood of mice fed a high fat diet was much higher than that of lean control mice. These studies suggest that serotonin may play important roles with regard to glucose and lipid metabolism. In this study, we have investigated the physiological effect of peripheral serotonin on the plasma levels of metabolites in both mice and sheep.[br][Results][br]Mice (C57BL/6, male, 7 weeks old) were fasted for 12 h before the experiment. After the intraperitoneal injection of 1 mg serotonin, the plasma glucose, insulin, triglyceride, NEFA, cholesterol and bile acids concentrations were measured. Plasma glucose and insulin significantly elevated after the injection. Plasma triglyceride, cholesterol and NEFA concentrations decreased. The concentration of bile acids in plasma increased after injection. These changes in plasma levels of metabolites and hormone were induced by several kinds of serotonin receptors using experiments of serotonin receptor antagonists. Wethers (crossbred, averaging 47.2 [plusmn] 7.8 kg of body weight) were fasted for 24 h before the experiment, and then intravenous injected with serotonin (40 [mu]g/kg body weight). Serotonin also induced the elevations of plasma glucose and insulin concentrations through the same serotonin receptor in wethers as well as in mice. On the other hand, the concentrations of plasma triglyceride and NEFA increased and the level of plasma bile acids decreased by serotonin injection. Additionally, serotonin did not affect the plasma cholesterol concentration. These results reveal that there are quit differences in the physiological functions of peripheral serotonin against lipid metabolism between mice and sheep.[br][br](1) Watanabe et al., Endocrinology 2010; 151:4776-4786.[br][br]Nothing to Disclose: HW, RS, TN, HT, YT, KS, YN, ST, NO, DWH, NS, HS, KW, HA 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 598 55 297 SAT-194 PO14-02 Saturday 236 2012


237 ENDO12L_SAT-195 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Insulin and Leptin Action on Kiss1 Neurons and the Regulation of Puberty and Reproduction Xiaoliang Qiu, Abigail Dowling, Joseph Marino, Latrice Faulkner, Laura Nedorezov, Jennifer Hill University of Toledo, Toledo, OH The neuropeptide kisspeptin, encoded by the Kiss1 gene, is necessary for puberty and fertility. Humans and mice with loss-of-function mutation of Kiss1 or its receptor are infertile due to hypogonadotropic hypogonadism. Insulin and leptin receptors are widely distributed throughout the hypothalamus and play a pivotal role in the regulation of energy balance, metabolism, and reproduction. Recent data show that leptin[apos]s effect on puberty in mice (KiSS1-Cre, ObR[sup]flox/flox[/sup]) does not require KiSS1 neurons. In order to study whether insulin -sensing plays an important role in Kiss1 neurons, we generated Kiss1 neuron specific insulin receptor knockout mice (Kiss1-Cre, IR [sup]flox/flox[/sup] mice). In addition, we produced Kiss1 neuron-specific insulin receptor and leptin receptor knockout mice (Kiss1-Cre, IR [sup]flox/flox[/sup] ObR[sup] flox/flox[/sup] mice) to look at the potential synergic role of the two receptors in the regulation of Kiss1 neurons. Here, we describe effects on vaginal opening, first estrus cycle, and fertility. Kiss1-Cre, IR [sup]flox/flox[/sup] female mice delayed vaginal opening (31.4[plusmn]0.5 VS 34.0[plusmn]0.9 days) and first estrus cycle (39.6[plusmn]1.2 VS 44.1[plusmn]1.6 days). Luteinizing hormone (LH) in Kiss1-Cre, IR [sup]flox/flox[/sup] mice were lower compared with control on early puberty 31 days old, but not adult. Body weight, food intake, and glucose regulation were comparable. Kiss1-Cre, IR [sup]flox/flox[/sup] ObR[sup] flox/flox[/sup] mice were examined for changes in pubertal timing, reproduction and metabolism as well. These results have important implications for understanding how insulin and leptin signaling in Kiss1 neurons may be involved in puberty and reproduction.[br][br]Sources of Research Support: NIH grant R00HD056491 to JWH.[br][br]Nothing to Disclose: XQ, AD, JM, LF, LN, JH 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2272 55 298 SAT-195 PO14-02 Saturday 237 2012


238 ENDO12L_SAT-196 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Reduction of Glucose-6-Phosphate Transporter Induces Suppression of 11[beta]-Hydroxysteroid Dehydrogenase Type 1 Expression and Improvement of Glucose Homeostasis in Diabetic Mouse Yanjun Liu, Ying Wang, Yuichi Nakagawa, Wei Wang, Alexei Lyzlov, Kabirullah Lutfy, Theodore C Friedman Charles Drew University of Medicine and Science, Los Angeles, CA; Hamamatsu University School of Medicine, Hamamatsu, Japan Pre-receptor activation of glucocorticoids via 11[beta]-hydroxysteroid dehydrogenase type 1 (11[beta]-HSD1) has been identified as an important mediator of metabolic syndrome. Hexose-6-phosphate dehydrogenase (H6PDH) mediates 11[beta]-HSD1 amplifying tissue glucocorticoid production by driving intracellular NADPH exposure to 11[beta]-HSD1 and requires glucose-6-phosphate transporter (G6PT) to maintain its activity. To test the efficiency of reducing G6PT expression to treat hyperglycemia and insulin resistance, we targeted the G6PT gene in type 2 diabetic mice using G6PT ASO specific to G6PT and explored the impact of G6PT on the pre-receptor metabolism of glucocorticoids in type 2 diabetes and obesity. We observed that G6PT ASO treatment of db/db mice markedly reduced hepatic G6PT mRNA and protein levels and improved glucose homeostasis and insulin resistance. The reduction of hepatic G6PT expression paralleled the suppression of 11[beta]-HSD1 and H6PDH production in the liver. These findings suggest that G6PT plays an important role in the modulation of pre-receptor activation of glucocorticoids and provide new insights into the role of G6PT in the development of type 2 diabetes.[br][br]Sources of Research Support: Y. Liu is supported by NIH grants SC1DK087655.[br][br]Nothing to Disclose: YL, YW, YN, WW, AL, KL, TCF 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1060 55 299 SAT-196 PO14-02 Saturday 238 2012


239 ENDO12L_SAT-197 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Glucagon-Induced Phosphorylation of the Salt-Inducible Kinase-2, a Key Mediator That Controls Hormone-Regulated Hepatic Gluconeogenesis Kashyap Patel, Olga Goransson, Calum Sutherland, Kei Sakamoto College of Life Sciences, University of Dundee, Dundee, UK; Lund University, Lund, Sweden; Ninewells Hospital and Medical School, University of Dundee, Dundee, UK The hormones insulin and glucagon are central to regulating glucose homeostasis via controlling hepatic gluconeogesis and peripheral glucose uptake. When the regulation of glucose homeostasis by these hormones dysfunctions, metabolic disorders develop, such as type 2 diabetes. Thus deciphering the details of molecular and cellular signaling network controlled by insulin and glucagon promises to provide new strategies for treating type 2 diabetes. Recent work has proposed that the salt inducible kinase-2 (SIK2), a closely related member of AMP-activated protein kinase (AMPK), plays a key role as a signaling mediator in the control of hormone-regulated hepatic gluconeogenesis. The aim of current study was to identify novel and key phosphorylation sites on SIK2 regulated by glucagon and insulin and elucidate the molecular mechanism by which SIK2 is regulated by these hormones in the liver. We have performed a phospho-peptide mapping analysis by a mass-spectrometry using recombinant SIK2 proteins isolated from mouse primary hepatocytes with or without glucagon or insulin treatment. This revealed that glucagon caused an increase in several phosphorylation sites including Ser358 and Ser587, but not Thr175 site in the T-loop of kinase domain, and these results were validated using phospho-specific antibodies to each site. Interestingly, but contrary to the previous work, we failed to see an increase in Ser358 phosphorylation in response to insulin at multiple time points measured (5-40 min) in mouse primary hepatocytes. To investigate if the phosphorylation of SIK2 by glucagon has impact on SIK2 kinase activity, endogenous SIK2 was immunoprecipitated from glucagon-treated mouse primary hepatocytes and in vitro kinase assay was performed. We observed no significant change in SIK2 activity between non-treated and glucagon-stimulated hepatocytes. We also performed mutagenesis analysis and observed that activity of Ser358Ala or Ser587Ala mutant SIK2 proteins isolated from HEK293 cells was similar compared to that of wild-type SIK2. Taken together, we have identified glucagon-regulated phosphorylation sites on SIK2, which do not appear to be critical for directly controlling SIK2 enzymatic activity, but may be important for localization or protein-protein interaction to regulate target proteins and glucose metabolism in hepatocytes.[br][br]Sources of Research Support: Wellcome Trust Grant awarded to KA.[br][br]Nothing to Disclose: KP, OG, CS, KS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1673 55 300 SAT-197 PO14-02 Saturday 239 2012


240 ENDO12L_SAT-198 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Hyperinsulinemia Promotes Metastasis to the Lung in a Mouse Model of Her2-Mediated Breast Cancer Rosalyn D Ferguson, Dara Cohen, Nyosha Alikhani, Aviva Tobin-Hess, Ruslan Novosyadlyy, Nadine Haddad, Derek Leroith Mount Sinai School of Medicine, New York, NY The Her2 oncogene is expressed in around 30% of human breast cancers and is associated with frequent metastatic progression and poor outcome.Epidemiological studies show that breast cancer incidence and mortality rates are higher in women with type 2 diabetes. Here we use a mouse model of Her2-mediated breast cancer on a background of hyperinsulinemia to determine how elevated circulating insulin levels affect Her2-mediated primary tumor growth and subsequent progression to lung metastasis. Hyperinsulinemic (MKR) mice were crossed with doxycycline-inducible Neu (rtTANeu) mice to produce the rtTANeuMKR mouse model. Both rtTANeu controls and rtTANeuMKR mice were administered doxycycline in drinking water to induce mammary tumor formation. At time of sacrifice total mammary tumor weight from both groups was measured and lungs were removed so that metastases could be quantified. After 90 days on doxycycline, although no significant increase in tumor weight was observed in rtTANeuMKR compared to rtTANeu mice, the number of lung metastases was significantly higher in rtTANeuMKR compared to rtTANeu mice (rtTANeuMKR 16.41 [plusmn] 4.18 vs. rtTANeu 5.36 [plusmn] 2.72). We analyzed signaling pathways downstream of the insulin receptor (IR)/insulin-like growth factor receptor 1 (IGF-1R) in tumor tissues. As well as finding increased activation of S6 kinase, an important mediator of cell proliferation, we also observed an increase in Snail, a transcription factor which is known to be important for the epithelial to mesenchymal transition (EMT) in breast cancer metastasis. We conclude that while primary tumors were similar in size, the hyperinsulinemia in the rtTANeuMKR mice resulted in increased EMT changes and therefore more aggressive primary tumors and a higher numbers of metastatic events.[br][br]Nothing to Disclose: RDF, DC, NA, AT-H, RN, NH, DL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 537 55 301 SAT-198 PO14-02 Saturday 240 2012


241 ENDO12L_SAT-199 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Hypoglycemic Effect of Aqueous Extract of [italic]Carica papaya[/italic] Leaf in Streptzotocin-Induced Diabetic Rats Isela Juarez Rojop, P H Miranda-Osorio, Teresa Ramon-Frias, Hidemi Aguilar-Mariscal, Jorge Ble-Castillo, A Rodriguez-Hernandez, Maria A Jimenez-Santos, Juan C Diaz-Zagoya Universidad Ju[agrave]rez Aut[oacute]noma de Tabasco, Villahermosa, Mexico; Universidad Ju[agrave]rez Aut[oacute]noma de Tabasco, Comalcalco, Mexico Diabetes is one of the major global public health problems and rapidly getting worse particularly in the developing nations (Unwin et al., 2009). In addition, the use of plants herbal remedies against diseases that constitutes economic problems such as diabetes (Adisa et al., 2011). [italic]Carica papaya[/italic] belongs to the family of [italic]Caricaceae[/italic] and several species of [italic]Caricaceae[/italic] have been used as remedy against a variety of diseases (Perez et al., 2003). The present study was to determine the effect of aqueous extract of [italic]C. papaya[/italic] leaf in streptozotocin (STZ)-induced diabetes in rats. Diabetes was induced in male Wistar rats (250-300g) using a single intraperitoneal injection of STZ (60mg/Kg). An experimental groups of non diabetic (ND) and diabetic (D) rats received an aqueous extract of [italic]C. papaya[/italic] a doses (3.5, 7.5 and 15 g/500 ml) for 30 days while other group ND and D received water for the same length of time. At the end of treatment, the animals were sacrificed by decapitation and blood sample was collected. Level of serum glucose, cholesterol, triglyceride and HDL-cholesterol concentrations were determined by using spectrometric methods. Level of insulin was assayed by an ELISA kit. Treatment with extract of [italic]C. papaya[/italic] reduced a dose-dependent manner level of glucose in D rats. Plasma insulin levels were increased by treatment in ND but not modify in D rats. Additionally,[italic] C. papaya[/italic] diminished the triglyceride level in D rats. The results indicated that aqueous extract of [italic]C. papaya[/italic] showed a clear hypoglycaemic effect in diabetic rats. Such an effect cannot be mediated by an enhanced insulin secretion.[br][br]Sources of Research Support: PFICA.UJAT-2010-C06-10.[br][br]Nothing to Disclose: IJR, PHM-O, TR-F, HA-M, JB-C, AR-H, MAJ-S, JCD-Z 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1064 55 302 SAT-199 PO14-02 Saturday 241 2012


242 ENDO12L_SAT-200 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Characterization of Amylin Receptor and Amylin-Regulated Insulin Secretory Pathways in Rat Insulinoma and Human Pancreatic Islet Cells Yasemin Cagil, Saurabh Trikha, Aleksandar Jeremic The George Washington University, Washington, DC Human amylin is a 37-amino acid peptide hormone that is produced and co-secreted with insulin by pancreatic islet beta-cells. In contrast to insulin, the physiological role of amylin is less clear. Studies indicate that amylin may, together with insulin, regulate secretion of insulin and other hormones from islets through autocrine/paracrine signaling mechanisms. However, the exact mechanism and receptors involved in regulatory effect of amylin on insulin secretion needs to be established. Hence in this study, the modulatory effect of amylin on basal and glucose-evoked insulin release in cultured rat insulinoma (RINm5F) cells and human islet cells were investigated using ELISA immunoassay. Low concentrations of amylin (1-10 nM) inhibited both basal and glucose-evoked insulin secretion in rat and human islets. This inhibitory effect of amylin on insulin secretion was also observed in cultures containing high (20 mM) potassium chloride. This result suggests that amylin acts downstream or at the level of glucose-evoked depolarization of beta-cells. This finding is consistent with previous studies showing the hyper-polarizing effect of human amylin on electrical activity of islet beta-cells. The physiological range (10[sup]-10 [/sup]- 10[sup]-8 [/sup]M) in which amylin modulates insulin release suggests involvement of amylin receptor in this secretory process. To test this hypothesis, we examined expression of amylin receptor (AM-R) constituents, calcitonin receptor (CT-R) and receptor activity modifying protein (RAMP), in two cell types by western blot and immunocytochemistry. Western blot analysis demonstrated expressions of type-1 and type-2 AM-R isoforms in human and rat pancreatic cells, respectively. Immunoconfocal microscopy further showed increased co-localization of CT-R and RAMP-2 on plasma membrane of RINm5F cells upon exposure to low doses of human amylin (1-10 nM) for 24 hours, indicating cells[apos] capacity for amylin sensing, which may play a role in regulation of insulin secretion. Further studies are needed to clarify causal connection between amylin receptor expression and amylin biological function including regulation of insulin release in pancreatic islets.[br][br]Nothing to Disclose: YC, ST, AJ 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1665 55 303 SAT-200 PO14-02 Saturday 242 2012


243 ENDO12L_SAT-201 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Uncoupling Protein 3 Inhibits Glucose Oxidation in Muscle Cells Romain Harmancey, Truong Nguyen Lam, Heinrich Taegtmeyer University of Texas Medical School at Houston, Houston, TX Glucose and fatty acids are competing substrates for energy provision in muscle. Fatty acid-mediated inhibition of glucose oxidation (Randle cycle) results from the inhibition of phosphofructokinase and pyruvate dehydrogenase (PDH). However, additional control sites may exist. Because fatty acids are potent activators of uncoupling protein (UCP) 3 expression, we proposed that the mitochondrial protein contributes to the inhibition of glucose oxidation in myocytes. L6 rat myoblasts were differentiated into myocytes, and changes in UCP3 expression levels were correlated to PDH activity by determination of its inhibitory phosphorylation status on Ser232, Ser293, and Ser300. Cellular rates of glucose oxidation were determined by quantitative collection of [[sup]14[/sup]C]O[sub]2 [/sub]following incubation with [U-[sup]14[/sup]C]glucose. Glycolytic flux was determined indirectly by quantification of lactate released in the incubation medium. To further examine the effect of UCP3 on glucose metabolism, we then transfected L6 myocytes with two siRNAs targeting the mitochondrial protein. UCP3 was not expressed in proliferating L6 myoblasts, but there was a 16-fold increase in UCP3 expression, both at the mRNA and protein levels, seven to eight days after the initiation of myogenic differentiation. The rise in UCP3 expression was tightly correlated to a gradual increase in PDH phosphorylation on Ser232 (r=0.9753; [italic]p[/italic][lt]0.001), Ser293 (r=0.9657; [italic]p[/italic][lt]0.01), and Ser300 (r=0.9461; [italic]p[/italic][lt]0.01). Compared to myoblasts, the rates of glucose oxidation were consequently reduced by more than 50% in the differentiated myocytes. In the meantime, lactate release was increased, suggesting that glycolysis becomes gradually uncoupled from glucose oxidation. The knockdown of the UCP3 transcript approached 70% with both siRNA sequences, resulting in a more than 60% decrease of UCP3 protein levels. This reduction in the protein level of UCP3 was sufficient to increase the rates of glucose oxidation in myocytes. We conclude that reduced UCP3 expression favors glucose oxidation and improves the coupling of glucose metabolism in cultured myocytes. Fatty acid-mediated expression of UCP3 may represent an additional control point for fatty acid inhibition of glucose oxidation.[br][br]Sources of Research Support: NHLBI Grant 5R01HL061483; Postdoctoral fellowship 09POST2060155 from the American Heart Association awarded to RH.[br][br]Nothing to Disclose: RH, TNL, HT 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1650 55 304 SAT-201 PO14-02 Saturday 243 2012


244 ENDO12L_SAT-202 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Testosterone Deficiency Is Associated with Decreased Expression of Muscle GLUT4 and Hexokinase 2 as Key Regulators of Insulin Sensitivity in the Testicular Feminized Mouse David S McLaren, Daniel M Kelly, Samia Akhtar, Kevin S Channer, Thomas H Jones University of Sheffield, Sheffield, UK; Royal Hallamshire Hospital, Sheffield, UK; Barnsley Hospital NHS Foundation Trust, Barnsley, UK Testosterone deficiency is common in men with type 2 diabetes (T2D). We have shown testosterone replacement therapy (TRT) improves insulin resistance and glycaemic control. The mechanisms by which testosterone mediates this action are unknown but may be due to a combination of effects on muscle, liver and adipose tissues. This study investigates the expression of Glut4 and HK2, (two key proteins involved in insulin sensitivity) in muscle tissue of the testicular feminised (Tfm) mouse which exhibit non-functional androgen receptors and low circulating levels of testosterone.[br]Tfm mice were fed a high-cholesterol diet ad libitum for 28 weeks and received either physiological testosterone replacement (mixed testosterone esters, Sustanon100) or placebo (saline) and were compared to wild-type littermates (WT). Striated muscles tissue was collected from the thigh. Expression of Glut 4 and HK2 mRNA and protein expression was analysed by qPCR and Western blotting.[br]There was a significant decrease in the relative mRNA expression of Glut-4 (0.59[plusmn]0.14, p=0.02) and HK2 (0.5[plusmn]0.16,p=0.01) in Tfm mice compared to WT. TRT did not significantly alter mRNA expression of Glut4 or HK2. Western blotting confirmed reduced protein expression of HK2 (0.33[plusmn]0.09vs0.04[plusmn]0.01,p=0.005) and Glut4 (1.05[plusmn]0.22vs0.57[plusmn]0.07,p=0.037) in Tfm mice compared to WT. TRT showed no change in HK2 protein expression compared to Tfm placebo (0.05[plusmn]0.02vs0.04[plusmn]0.01,p=0.627), but an increase in Glut4 was observed (1.09[plusmn]0.16vs0.57[plusmn]0.07,p=0.005).[br]Testosterone deficiency is associated with decreased expression of Glut4 and HK2. TRT did not have any effect on HK2 in the Tfm mouse suggesting that testosterone acts via the androgen receptor to stimulate HK2 expression. Although testosterone failed to increase Glut4 mRNA in the Tfm an increase in protein was observed indicating that post-translational mechanisms are involved. This study shows that testosterone has important actions on modulating glucose metabolism in muscle. This may explain part of the mechanism by which testosterone improves insulin sensitivity in T2D.[br][br]Nothing to Disclose: DSM, DMK, SA, KSC, THJ 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1453 55 305 SAT-202 PO14-02 Saturday 244 2012


245 ENDO12L_SAT-203 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Potential Mechanism of Type 2 CRF Receptor-Mediated Skeletal Muscle Insulin Resistance Hongxia Chao, Haochen Li, Peilin Chen, Chien Li University of Virginia Health System, Charlottesville, VA Insulin resistance for glucose disposal by skeletal muscle (SM) is an essential and perhaps primary defect for Type 2 diabetes and metabolic syndrome. The underlying mechanism of muscle insulin resistance has not been fully delineated and is likely due to an interaction between multiple extrinsic and intrinsic factors. Urocortin 2 (Ucn 2), a member of the corticotropin-releasing factor (CRF) family, and its selective receptor, the type 2 CRF receptor (CRFR2), is highly expressed in SM. It has been suggested that CRFR2 and its ligand may be involved in regulating glucose homeostasis in muscle.[br]In the present study, we found that CRFR2 expression was greatly increased in the muscle of obese diabetic Ob/Ob mice, but was significantly reduced in the muscle of exercised mice compared to that of sedentary control mice. Consistent with our in vivo observation, CRFR2 expression in insulin resistant C2C12 myotubes induced by either chronic insulin or palmitic acid treatment was greatly elevated compared to that of control cells. Functional study showed overexpression of CRFR2 in SM cells blocked insulin-induced glucose uptake. Conversely, reduced CRFR2 expression improved insulin sensitivity in insulin resistant myotubes. Moreover, we found that cAMP is required for the inhibitory effect of Ucn 2 on insulin-induced glucose uptake and blocking Epac and protein kinase A, two prominent cAMP downstream effectors, failed to modulate the effect of Ucn 2. Interestingly, pretreating cells with rapamycin prevented the inhibitory effect of Ucn 2 on insulin-induced glucose uptake. Furthermore, Ucn 2 attenuated insulin-induced tyrosine608 phosphorylation of insulin receptor substrate 1 (IRS1) and increased phosphorylation of IRS1 serine 636/639 residues, whereas treatment with rapamycin reversed the phosphorylation of these residues. Finally, Ucn 2 failed to inhibit glucose uptake induced by platelet-derived growth factor, which stimulates PI3-kinase signaling in an IRS1 independent manner. This result further supports the notion that CRFR2 signaling modulates insulin[apos]s action at the level of IRS1.[br]In summary, we show CRFR2 expression is closely correlated with the status of muscle insulin sensitivity and CRFR2 signaling plays a critical role in modulating insulin-induced glucose uptake. Furthermore, we found that IRS1 is the key nodal point where CRFR2 signaling modulates insulin action and the effect of CRFR2 may be mediated by a novel cAMP-mTOR signaling pathway.[br][br]Sources of Research Support: NIH Grant DK078049 awarded to CL.[br][br]Nothing to Disclose: HC, HL, PC, CL 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2026 55 306 SAT-203 PO14-02 Saturday 245 2012


246 ENDO12L_SAT-204 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Adiponectin Restores Insulin Responsiveness in Human Neuroblastoma Cell Line SH-SY5Y Emna Chaabouni, Gael Lemerle, Yacir Benomar, Michel Parquet, Mohammed Taouis University Paris-Sud, Orsay, France; CNRS, Orsay, France Adiponectin regulates energy homeostasis through the modulation of glucose and fatty acid metabolism in peripheral tissues. However, its central effect on energy balance remains unclear and controversial. Recent advances in our understanding of the signal transduction mechanisms used by adiponectin in the periphery and in the hypothalamus suggest that intracellular cross-talk between adiponectin, leptin and insulin may occur. In the present study we aimed to study the impact of adiponectin on insulin responsiveness in human neuroblastoma cell line SH-SY5Y. In order to study the potential cross talk between adiponectin and insulin signaling pathways, it is necessary to demonstrate that SH-SY5Y cells express insulin and adiponectin receptors. We have previously shown that these cells express insulin receptor, and here using immunohistochemical approach we show that SH-SY5Y cells express both AdipoR-1 and Adipo-R2. Then we studied the functionality of these receptors by measuring the phosphorylation of AMPkinase in response to adiponectin. We show that adiponectin increased AMP-K phosphorylation in a dose dependent manner. Since in literature adiponectin is considered as an anti-diabetic hormone, we studied the plausible impact of adiponectin in delaying or abolishing insulin resistance associated to an overexposure of cells to insulin. Firstly, we show that a pretreatment with insulin significantly down-regulated insulin receptor expression at the protein level which is accompanied by an impairment of insulin-dependent Akt phosphorylation. To investigate the potential anti-insulin resistance effect of adiponectin, we studied the impact of adiponectin on Akt phosphorylation after insulin overexposure. As expected, in the absence of insulin pretreatment, acute insulin stimulation increased significantly Akt phosphorylation. This effect was completely abolished when cells were overexposed to insulin. Interestingly, when cells were overexposed to both insulin and adiponectin the insulin-dependent Akt phosphorylation was completely restablished protecting then cells from insulin resistance. In addition, we show that adiponectin exerted, most likely, its effect by reducing the insulin-dependent down-regulation of insulin receptors.[br]In conclusion, we show that adiponectin in a neuronal cells is able to maintain insulin responsiveness in an environment favoring insulin resistance through probably the inhibition of insulin receptor down-regulation.[br][br]Nothing to Disclose: EC, GL, YB, MP, MT 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1182 55 307 SAT-204 PO14-02 Saturday 246 2012


247 ENDO12L_SAT-205 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Low-Carb/High-Fat Diets Reduce Fasting Glucose and Insulin Levels, but Are Associated with Insulin Resistance and Reduced Pancreatic [beta]-Cell Mass in Rats Maximilian Bielohuby, Stephanie Sisley, Nadja Herbach, Darleen Sandoval, Juliane Ramisch, Joyce Sorrell, Dominik Menhofer, Rudiger Wanke, Kerstin Stemmer, Matthias H Tschop, Randy J Seeley, Martin Bidlingmaier Medizinische Klinik und Poliklinik IV, Klinikum der LMU, Munich, Germany; University of Cincinnati, Cincinnati, OH; College of Medicine, University of Cincinnati, Cincinnati, OH; LMU, Munich, Germany; Helmholtz Centre for Health and Environment and Technical University, Munich, Germany Low-carbohydrate/high fat (LCHF) diets are claimed to induce weight loss in overweight subjects and to ameliorate glucose metabolism. To investigate the effects of LCHF diets on glucose and insulin metabolism independent of the energetic intake, we pair-fed male rats isoenergetic amounts of standard rodent chow (CH) or one of two different LCHF diets (% of metabolizable energy, fat/protein/CHO: LCHF-1 (78.7/19.1/2.2), LCHF-2 (92.8/5.5/1.7) and CH (16.7/19./64.3)). After 3 weeks of feeding the respective diets, oral glucose tolerance tests (OGTT), insulin tolerance tests (ITT) and hyperinsulinemic-euglycemic clamps were performed (n=4-5/challenge test and diet group). After 4 weeks on the diets, rats were fasted for 6h and sacrificed for collection of trunk blood and pancreas. Serial pancreas sections from rats which have not been previously challenged were used to determine total [beta]-cell volume by quantitative stereological techniques (n=5/diet group). Results: Basal fasting glucose and insulin levels were lower in rats fed LCHF vs. CH diets (p[lt]0.05). Total pancreas volumes in rats fed LCHF diets were 20% (LCHF-1; p[lt]0.01) and 30% (LCHF-2; p[lt]0.001) lower compared to CH. Quantitative stereological analyses revealed that both LCHF diets led to significantly (p[lt]0.001) lower total [beta]-cell volumes, both absolute and relative to pancreas volume and body weight. During OGTT, the area under the glucose curve was significantly higher in rats fed both LCHF diets, but insulin secretion was not affected. The decline in glucose during ITT was significantly delayed in LCHF vs. CH fed animals. Clamp studies showed that the glucose infusion rate was about 3-fold higher (p[lt]0.05) and endogenous glucose production (EGP) over than 50% lower in rats fed CH. In contrast, EGP was not inhibited and clearance of glucose was significantly reduced by both LCHF diets. In summary, feeding LCHF diets led to lower fasting glucose and insulin levels. At first sight these data support a beneficial role of LCHF diets for glucose metabolism. However, dynamic challenge tests revealed that rats fed LCHF diets developed insulin resistance. Moreover, lower basal insulin levels of rats fed LCHF diets are most likely explained by a loss of pancreatic [beta]-cell mass. This occurred independent of the relative abundance of fat and protein in LCHF diets and in the absence of increased energetic intake. Our data clearly argue against a beneficial effect of LCHF diets on glucose and insulin metabolism.[br][br]Disclosures: DS: Research Funding, Johnson & Johnson, Novo Nordisk, Pfizer, Inc.; Speaker, Novo Nordisk; MB: Study Investigator, Chiasma. Nothing to Disclose: MB, SS, NH, JR, JS, DM, RW, KS, MHT, RJS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 951 55 308 SAT-205 PO14-02 Saturday 247 2012


248 ENDO12L_SAT-206 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) A New Syntaxin4 Binding Partner Tctex1d2 Is Specifically Expressed in Adipocytes and Inhibits GLUT4 Translocation through Interference of Doc2b-Syntaxin4 Interaction Atsushi Nohara, Shuichi Okada, Tsugumichi Saito, Kihachi Ohshima, Jeffrey E Pessin, Masatomo Mori Gunma University Graduate School of Medicine, Maebashi, Japan; Gunma University, Maebashi, Japan; Albert Einstein College of Medicine, New York, NY ctex1d2 is a novel gene that encodes for a functionally unknown protein that contains a Tctex1 domain found in dynein light chain family members. Examination of gene expression during adipogenesis demonstrated a marked increased in Tctex1d2 gene expression that was essentially undetectable in pre-adipocytes and markedly induced during 3T3-L1 adipocyte differentiation. Acute insulin stimulation (up to 30 min) in 3T3-L1 adipocytes over expressing Tctex1d2 had no effect on the phosphorylation of the insulin receptor [beta] subunit, IRS-1, Erk, or Akt. However Tctex1d2 overexpression significantly inhibited insulin-stimulated GLUT4 translocation. Syntaxin4 is a binding partner for Doc2b and the v-SNARE protein VAMP2 that are required for insulin-stimulated GLUT4 translocation. Over expression of Tctex1d2 was found to associate with Syntaxin4 and prevent the insulin-stimulated association of Doc2b. Based upon these data, we hypothesize that Tctex1d2 and Doc2b share binding sites on Syntaxin4 thereby inhibiting insulin induced GLUT4 translocation through interference of Doc2b-Syntaxin4 interaction.[br][br]Nothing to Disclose: AN, SO, TS, KO, JEP, MM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 32 55 309 SAT-206 PO14-02 Saturday 248 2012


249 ENDO12L_SAT-207 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Estrogen Facilitates Insulin Signaling in a Leptin-Dependent Manner Shiva PD Senthil Kumar, Xian Liu, Haifei Shi Miami University, Oxford, OH Estrogens regulate glucose homeostasis. Postmenopausal women with decreased levels of endogenous estrogen have increased risk of insulin resistance, and treatment with estradiol (E2; the primary form of estrogen) improves insulin sensitivity. Leptin and insulin, two hormones that regulate energy homeostasis, have overlapping intracellular PI3/Akt signaling pathway and elicit similar metabolic actions. We hypothesize that the insulin sensitizing action of estrogen on insulin-targeted tissues is influenced by leptin. Three male mouse models were used in this study, standard chow-fed lean mice with low circulating leptin level and normal leptin sensitivity, high-fat diet-induced obese mice with high circulating leptin level and leptin resistance, and leptin-deficient [italic]ob/ob[/italic] mice. First the role of E2 in insulin sensitivity was assessed using intraperitoneal insulin tolerance test in combination with injections of E2 or its vehicle. Injection of a sub-threshold dose of insulin (0.5 U/kg) did not significantly change glucose levels of lean mice; however, when the lean mice were co-injected with 200 mg/kg E2 and 0.5U/kg insulin, their glucose levels declined more rapidly, with significantly greater glucose disappearance rate and lower glucose levels, than the lean mice injected with 0.5U/kg insulin and vehicle for E2. The obese mice co-injected with 1U/kg insulin and E2 cleared glucose efficiently, with their glucose disappearance rates being faster than the obese mice injected with insulin alone. In contrast, [italic]ob/ob[/italic] mice co-injected with E2 and insulin had similar glucose levels as the [italic]ob/ob[/italic] mice treated with insulin alone. Thus, administrations of insulin and E2 had greater effects in lowering glucose levels than insulin alone in wildtype, but not in leptin deficient mice. Second E2[apos]s effects on insulin and leptin signaling pathways were investigated. Phosphorylation of Akt (common markers for insulin and leptin signaling), MAPK (marker for insulin signaling), and STAT3 (marker for leptin signaling) in metabolic tissues was measured by Western blot analysis. The ratios of pAkt/total Akt, pMAPK/total MAPK, and pSTAT3/total STAT3 in skeletal muscle, white adipose tissue and the hypothalamus of wildtype mice increased after co-injection of E2 and insulin comparing to insulin injection alone. Our data indicate that acute E2 improves insulin sensitivity and suggest that this effect is related to an activated insulin and leptin signaling in the periphery and the CNS.[br][br]Sources of Research Support: National Institutes of Health National Institute of Diabetes and Digestive and Kidney Diseases [Grant 1R15DK090823-01 to HS] and American Heart Association [Grant 10SDG4520028 to HS].[br][br]Nothing to Disclose: SPDSK, XL, HS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1233 55 310 SAT-207 PO14-02 Saturday 249 2012


250 ENDO12L_SAT-208 POSTER SESSION: Insulin [amp] Nutrient Metabolic Action in Animal Models (1:30 PM-3:30 PM) Adult Rats Exposed to Ethanol In Utero Have Increased Hepatic Gluconeogenesis Due to Oxidative Stress-Induced Histone Deacetylases Xing-Hai Yao, Hoa Khanh Nguyen, Jonghan Lee, Gregoire Nyomba University of Manitoba, Winnipeg, Canada [bold]Introduction:[/bold] Prenatal ethanol (EtOH) increases hepatic gluconeogenesis in adult rat offspring by unknown mechanism[bold]. [/bold][br][bold]Methods:[/bold] Four month-old male rat offspring exposed to EtOH with or without tauroursodeoxycholic acid (TUDCA) during pregnancy days 1-7 (early), 8-14 (mid) and 15-21 (late) were compared with those from pair-fed (PF) dams. We performed pyruvate challenge test and measured liver reactive oxygen species (ROS), phosphoenolpyruvate carboxykinase (PEPCK), glucose 6-phosphatase (G6Pase), nuclear histone deacetylases (HDACs), and acetylated Foxo1 (AcFoxo).[br][bold]Results:[/bold] EtOH vs PF vs TUDCA (Mean[plusmn]SE, *P[lt]0.05, **P[lt]0.01):[br]-pyruvate induced glycemia (AUC): early: 1485[plusmn]93 vs 1159[plusmn]133* vs 1188[plusmn]76*; mid: 1565[plusmn]55 vs 1229[plusmn]67** vs 1259[plusmn]83*; late: 1734[plusmn]70 vs 1242[plusmn]84** vs 1375[plusmn]115*;[br]-ROS: early: 118.84[plusmn]13.07 vs 72.61[plusmn]6.65** vs 81.83[plusmn]9.56*; mid: 124.25[plusmn]12.51 vs 65.46[plusmn]9.02** vs 79.91[plusmn]8.58*; late: 119.68[plusmn]14.71 vs 73.52[plusmn]9.49* vs 79.62[plusmn]7.90*;[br]-PEPCK: early: 1.60[plusmn]0.15 vs 1.07[plusmn]0.13* vs 1.30[plusmn]0.13; mid: 2.03[plusmn]0.21 vs 1.14[plusmn]0.10** vs 1.20[plusmn]0.15*; late: 1.90[plusmn]0.29 vs 0.91[plusmn]0.12** vs 1.06[plusmn]0.09**;[br]-G6Pase: early: 1.63[plusmn]0.24 vs 0.95[plusmn]0.13* vs 1.33[plusmn]0.13; mid: 1.47[plusmn]0.12 vs 0.97[plusmn]0.12* vs 1.21[plusmn]0.12; late: 1.42[plusmn]0.08 vs 1.05[plusmn]0.10* vs 1.25[plusmn]0.12;[br]-HDAC activity: early: 0.92[plusmn]0.07 vs 0.43[plusmn]0.05** vs 0.63[plusmn]0.09*; mid: 0.75[plusmn]0.05 vs 0.45[plusmn]0.05** vs 0.52[plusmn]0.05*; late: 0.72[plusmn]0.07 vs 0.38[plusmn]0.05** vs 0.52[plusmn]0.05*;[br]-HDAC3: early: 1.73[plusmn]0.12 vs 0.94[plusmn]0.06** vs 1.07[plusmn]0.11**; mid: 1.60[plusmn]0.13 vs 1.05[plusmn]0.08** vs 1.16[plusmn]0.09*; late: 2.73[plusmn]0.25 vs.98[plusmn]0.13** vs 1.25[plusmn]0.09**;[br]-HDAC4: early: 2.04[plusmn]0.27 vs 0.76[plusmn]0.14** vs 1.21[plusmn]0.27*; mid: 1.87[plusmn]0.27 vs 0.96[plusmn]0.10** vs 1.26[plusmn]0.09*; late: 2.37[plusmn]0.47 vs 0.93[plusmn]0.11**vs 1.31[plusmn]0.45*;[br]-HDAC5: early: 1.95[plusmn]0.22 vs 1.09[plusmn]0.12** vs 1.04[plusmn]0.14**; mid: 1.85[plusmn]0.21 vs 1.09[plusmn]0.09* vs 1.17[plusmn]0.21*; late: 2.63[plusmn]0.55 vs 0.72[plusmn]0.17** vs 1.34[plusmn]0.20*;[br]-HDAC7: early: 3.32[plusmn]0.79 vs 0.76[plusmn]0.11** vs 1.47[plusmn]0.33*; mid: 2.56[plusmn]0.55 vs 0.87[plusmn]0.27** vs 1.33[plusmn]0.23*; late: 2.18[plusmn]0.52 vs 0.96[plusmn]0.13* vs 1.05[plusmn]0.14*;[br]-AcFoxo: early: 0.17[plusmn]0.03 vs 0.98[plusmn]0.07** vs 0.46[plusmn]0.05*; mid: 0.22[plusmn]0.06 vs 1.01[plusmn]0.06** vs 0.43[plusmn]0.04*; late: 0.16[plusmn]0.02 vs 1.07[plusmn]0.15** vs 0.48[plusmn]0.05*.[br][bold]Conclusion:[/bold] Adult rats exposed to EtOH in utero, have increased gluconeogenesis, oxidative stress, and HDAC expression, but reduced Foxo acetylation. Oxidative stress likely increases HDACs, which deacetylate Foxo, leading to gluconeogenic genes transcription. These anomalies occur even in early pregnancy and can be prevented by TUDCA administration to the EtOH dams.[br][br]Sources of Research Support: Canadian Diabetes Association grant OG-3-11-3419-BN.[br][br]Nothing to Disclose: X-HY, HKN, JL, GN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 124 55 311 SAT-208 PO14-02 Saturday 250 2012


251 ENDO12L_SAT-209 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Hallucinations Associated with Miglitol Use in a Patient with Chronic Kidney Disease and Hypothyroidism Thet Hlaing, Shilpa Chaudhari, Alan Sacerdote, Gul Bahtiyar Woodhull Medical and Mental Center, Brooklyn, NY; State University of New York (SUNY) Downstate Medical Center, Brooklyn, NY; New York University School of Medicine, New York, NY; St George[apos]s University School of Medicine, Grenada, Grenada A 71 year old woman with Type 2 diabetes mellitus (Type 2DM), chronic kidney disease stage IV (CKD stage IV), primary hypothyroidism, and osteoarthritis whose prescribed treatment included miglitol 50 mg thrice daily with the first bite of meals reported that she suffered visual hallucinations while taking miglitol, which resolved within a few days of stopping the drug. When she resumed miglitol hallucinations recurred within a few days and again resolved within a few days of stopping the drug. At no point were hallucinations associated with any symptom of hypoglycemia or a low finger stick glucose reading. Glycemic control was suboptimal with HbA1c by HPLC = 11.2%. She was still hypothyroid on thyroxine replacement with TSH by chemiluminescence = 39.7 mIU/ml.[br]Although, in general, miglitol is minimally absorbed following oral administration (2-3%) it is mostly eliminated unchanged via the renal route and thus, may accumulate, in the setting of renal failure. Likewise, incompletely compensated hypothyroidism may result in drug accumulation. Once absorbed, miglitol is able to cross the blood/brain barrier and potentially inhibit the alpha-glucosidase enzymes of the the central nervous system (CNS), eg isomaltase, resulting in CNS glycogen accumulation mimicking genetic glycogen storage diseases, eg Pompe disease, which may also present with hallucinations. Inhibition of CNS alpha-glucosidases may also result in formation of aneurysms. Although one series has reported a 2% incidence of hallucinations in patients taking the related drug, acarbose, the risk is not widely appreciated and it is not a reported adverse effect in the package insert for either medication.[br]We conclude that alpha-glucosidase inhibitors should be used with caution in patients with coexisting renal insufficiency and incompletely treated hypothyroidism. CNS changes, in the absence of hypoglycemia should dictate drug discontinuation.[br][br]Ritesh Patil, MD, MPH and Jack L. DePriest, MD, FCCP, FACP. J Gen Intern Med. 2009 May; 24(5): 683[ndash]686.[br][br]Nothing to Disclose: TH, SC, AS, GB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 136 56 312 SAT-209 PO47-02 Saturday 251 2012


252 ENDO12L_SAT-210 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Severe Hyperglycemic Hyperosmolar State and Acute Mental Status Changes in a Child with New-Onset Type 1 Diabetes Mellitus Nohemie Brevil, Priya Khanna Mount Sinai Hospital, Chicago, IL [italic][bold]Introduction:[/bold][/italic] Hyperglycemic hyperosmolar state (HHS) is a life-threatening diabetic emergency that is thought to occur most commonly in adults patients with type 2 diabetes mellitus. However HHS is increasingly being seen in pediatric patients presenting with new onset type 1 and type 2 diabtes mellitus, and is associated with a high mortality rate.[br][italic][bold]Clinical Case:[/bold][/italic] An 8.9-year old, previously healthy, African American female with a low BMI (14.3 kg/m2) presented to our emergency department with history of weakness and altered mental status for two days. After initial evaluation, the patient was admitted to our pediatric intensive care unit (PICU) for mild diabetic ketoacidosis (DKA), hypernatremia, severe dehydration, renal insufficiency, and hyperosmolarity. Her initial lab findings were: Serum glucose 1258 mg/dL (65-99), serum sodium 155 mEq/L (138-145), corrected sodium 173 mEq/L, serum osmolality 412 mOsm/kg (278-305), serum bicarbonate 11 mmol/L (22-29), blood urea nitrogen 90 mg/dL (5-18), creatinine 4.69 mg/dL (0.3-0.7), venous pH 7.11, beta-hydroxybutyrate 15.3 mmol/L (0.1-0.3), urine ketone 10mg/dL. Her hemoglobin A1c was found to be 10.1%. Islet cell antibodies were negative while those to glutamic acid decarboxylase (GAD) were positive at more than 30 U/ml. These results are consistent with HHS in a new onset type 1 diabetic. The patient[apos]s acidosis corrected within six hours of presentation. She continued to have altered mental status and decreased cognition for three days. Her hyperosmolarity and hypernatremia resolved slowly over four days with careful IV hydration. Once her neurologic status returned to baseline and she was ready for oral feeds, she was started on subcutaneous insulin therapy. She was discharged home after one week of hospitalization.[br][italic][bold]Conclusion:[/bold][/italic] This case highlights the fact that pediatric patients with new onset type 1 diabetes mellitus can present with both DKA and HHS. Management strategies are undefined, but we do know that these patients require careful IV hydration and appropriate fluid replacement to improve their clinical outcome and decrease their mortality rate.[br][br]Nothing to Disclose: NB, PK 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 957 56 313 SAT-210 PO47-02 Saturday 252 2012


253 ENDO12L_SAT-211 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) A Case of Steroid Diabetes with Severe Acute Pancreatitis after Liraglutide and Bezafibrate Administration Naoki Hiroi, Mariko Sue, Kenzaburo Oda, Yoshie Oka, Rika Shigemitsu, Ryo Iga, Aya Yoshihara, Shuki Usui, Koji Kuboki, Gen Yoshino Toho University, Tokyo, Japan [Introduction] It is reported that incidence of drug induced acute pancreatitis is only 0.2[ndash]2%, however the true incidence is not clear. The mechanism of drug induced damage of the pancreas is not still completely understood.[br][Case] A 58-year-old, nonalcoholic Japanese male with steroid induced diabetes, who had taken prednisolone 30 mg daily for 3 years for rheumatoid arthritis and interstitial pneumonia, presented with a 3 days history of mid-epigastric pain 3 months after treatment of liraglutide 0.6 mg daily and 1 month after treatment of bezafibrate 200mg daily. Levels of fasting plasma glucose and glycosylated hemoglobin were 172 mg/dL and 6.5% (NGSP), respectively. Liver function test results and serum creatinine concentration were almost normal. Level of triglyceride was 155 mg/dL. Serum lipase concentration was 79 U/L (reference range, 11-53 U/L), and serum amylase concentration was 513 U/L (reference range, 24-137 U/L) at admission. Abdominal computed tomography (CT) showed changes consistent with pancreatitis, and the intestinal thickness and ascites. Liraglutide and bezafibrate were discontinued. Conservative therapy for acute pancreatitis resulted in rapid resolution of symptoms, normal lipase concentration (37 U/L).[br][Discussion] In this case, diabetes and hypertriglyceridemia were stable, and the past history which caused pancreatitis was not seen. He took many drugs for treatment of rheumatoid arthritis and interstitial pneumonia, however, administration of liraglutide and/or bezafibrate might cause pancreatitis, because there were no drugs introduced other than liraglutide and bezafibrate newly.[br][Conclusions] Liraglutide and/or bezafibrate should be used cautiously in patients with diabetic patients.This case suggests that liraglutide- and/or bezafibrate-induces pancreatitis is rare but potentially serious adverse effects.[br][br]Nothing to Disclose: NH, MS, KO, YO, RS, RI, AY, SU, KK, GY 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1522 56 314 SAT-211 PO47-02 Saturday 253 2012


254 ENDO12L_SAT-212 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Effect of Vildagliptin on Central Aortic Blood Pressure in Diabetic and Hypertensive Individual Luciana Neves Cosenso-Martin, Jose Fernando Vilela-Martin, Luiz Tadeu Giollo-Junior State Medical School of Sao Jose do Rio Preto (FAMERP), Sao Jose do Rio Preto, Brazil Background: Vildagliptin is a incretin-based therapy for the treatment of the type 2 diabetes (DM). Incretins are gastrointestinal hormones, predominantly glucagon-like peptide 1 (GLP-1), which are inactived by dipeptidyl peptidase-4 (DPP-4), and they stimulate insulin secretion. Vildagliptin is a DPP-4 inhibitor, improving [beta]-cell function, and decreasing glucagon secretion. GLP-1 has vasodilatory actions, and improves endothelium function in animals and humans. Thus, the objective of this case report was to evaluate the effect of the vildagliptin in the endothelium using the applanation tonometry of the radial artery (AT). [br]Clinical Case: Woman, 51 years-old, caucasian. She related DM for 4 years and high blood pressure for 6 years. Medicines in use: metformin 1700 mg/day, pioglitazone 30 mg/day, sinvastatin 10 mg/day, enalapril 5 mg/day and amlodipine 10 mg/day. No smoking. No DM in the family. At examination: Blood Pressure (BP): 129X81 mmHg; Heart rate: 78 bpm; BMI: 30 kg/m2; waist circumference: 91 cm[br]Glycated hemoglobina (HbA1c): 11,11%; Microalbuminuria: normal.[br]Evaluation of the central BP and Augmentation Index (AI) by AT were made before and after treatment with vildagliptin 100 mg/day. After 3 months, the patient returned taking vildagliptin and the medicines above described. No adverse effects were related. BP: 100X70 mmHg. HbA1c: 7,2%. AI and systolic central aortic BP reduced, 96% to 72% and 127 to 101 mmHg, respectively.[br]Clinical Lessions: AT is a noninvasive technique used to examine arterial elastic properties. An increased AI is associated with higher cardiovascular risk, and it was reported in patients with DM or impaired glucose tolerance. Higher values of AI indicate increased arterial stiffness. In health young individual, the systolic central aortic BP is around 20 mmHg below the systolic peripheric BP. Without vildagliptin the systolic central BP was only 2 mmHg lower than peripheric BP. This fact could be considered as an endothelium dysfunction, which was reversed by the vildagliptin, demonstrated by AI and systolic central BP reduction. Lower values for AI indicate a good elasticity of the arterial wall. We think this effect is not exclusive of the vildagliptin, because glucose control also can improve endothelium function. Others studies are necessary to confirm our results.[br]Conclusion: This case report is interesting because evaluate the central BP, demonstrating the effect of a DPP-4 inhibitor in endothelium dysfunction.[br][br]Nothing to Disclose: LNC-M, JFV-M, LTG-J 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1317 56 315 SAT-212 PO47-02 Saturday 254 2012


255 ENDO12L_SAT-213 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Puberty Onset of Diabetes Mellitus with 49, XXXXY Syndrome Takamasa Ichijo, Fumitaka Saida, Eiko Yoshida, Kaoru Yamashita, Hiromi Ouchi, Mariko Higa Saiseikai Yokohamashi Tobu Hospital, Yokohama, Japan [bold]Backbround: [/bold]The 49, XXXXY syndrome is a rare X-chromosome polysomy characterized by mental retardation, skeletal defects, craniofacial anomalies and hypogonadism (1). This syndrome is considered the most severe variant of Klinefelter syndrome. Although the incidence of diabetes among patients with Klinefelter syndrome or other types of X-chromosome polysomy is higher (2), the literature reported about puberty development of diabetes with 49, XXXXY syndrome are rare and only 2 cases have been reported in our knowledge. One possibility of this rare is most patients with 49, XXXXY syndrome are diagnosed in the newborn period rather than in adulthood as usually seen in 47, XXY Klinefelter syndrome.[br][bold]Clinical Case: [/bold]A 24-year-old man was admitted to our hospital for the treatment of hyperglycemia. He was diagnosed as 49, XXXXY syndrome at his birth and diabetes at 18-year-old. He was first treated by 1mg of glimepiride and increased the dose, but his plasma glucose level was not well controlled, and insulin injection therapy was started at 20-year-old. When he admitted to our hospital, he was treated by 30% insulin aspart premix twice a day, 24U at breakfast and 22U at supper, and his fasting glucose level was 143mg/dl and HbA1c was 7.9%. His height, weight and body mass index were 173.6 cm, 64.6 kg, and 21.4, respectively. His sexual development was Tanner stage I and small testes were palpable in his scrotum. His endogenous insulin profile, the fasting plasma c-peptide (CPR), 24 H urine CPR, and the glucagon test, showed 3.9 ng/ml (normal: [gt]0.5 ng/ml), 62.2 [mu]g/day (normal: [gt]20 [mu]g/day), and DCPR 1.2 ng/ml (normal: [gt]1.0 ng/ml), and his insulin secretion was conserved. His sex hormonal tests showed LH 13.0 mIU/ml (normal: 0.8-5.7 mIU/ml), FSH 40.8 mIU/ml (normal: 2-8.3 mIU/ml), and serum free testosterone [lt]0.5 pg/ml (normal: 2.55-7.53 pg/ml). We adjusted his administrating insulin and discharged with 22U of NPH at breakfast and 20U of 30% insulin aspart premix at supper, and his fasting glucose level and HbA1c were 112 mg/dl and 6.4% after a month. Because of his mother[apos]s refusal, the testosterone replacement therapy has not been started.[br][bold]Conclusion:[/bold] This is the third case report of 49, XXXXY syndrome with diabetes. We rather insist a periodical follow up is important to detect the development of diabetes and prevent the complications. We must refocus on his/her quality of life including prevention of diabetic complications in patients with 49,XXXXY syndrome.[br][br](1) Horm Res 2006;65;14-17. (2) J Clin Endocrinol Metab 1969;29;1062-1073.[br][br]Nothing to Disclose: TI, FS, EY, KY, HO, MH 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 182 56 316 SAT-213 PO47-02 Saturday 255 2012


256 ENDO12L_SAT-214 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Successful Treatment of Acute Esophageal Necrosis Associated with Diabetic Ketoacidosis in a Patient with Type 1 Diabetes Maiko Mouri, Tsuguka Shiwa, Masayasu Yoneda, Tomokazu Awaya, Shuhei Nakanishi Chugoku Rosai Hospital, Kure, Japan; Graduate School of Biomedical Sciences, Hiroshima University, Hiroshima, Japan A 64-year-old Japanese man was admitted to our hospital due to frequent vomiting and poor appetite for 2 days. He was diagnosed as diabetes 9 years ago and had been treated with biphasic insulin twice a day. He used to drink a lot and had drunk too much before his vomiting began. On admission, laboratory findings showed hyperglycemia (1,108 mg/dl) and metabolic acidosis, pH 7.215 with low bicarbonate levels. Ketone bodies in urine were elevated. These clinical findings indicated the diabetic ketoacidosis. Further, he presented melena. Upper gastrointestinal endoscopic examination showed black esophageal mucosa of the distal esophagus and erosion from just below the pharyngoesophageal constriction to the esophagogastric junction because of the presence of melena. Thus, he was diagnosed as diabetic ketoacidosis accompanying acute esophageal necrosis (AEN). He was immediately treated with continuous insulin administration to properly control blood glucose level as well as with total parenteral nutrition, oral intake restriction and intravenous administration of proton pump inhibitor. AEN was successfully treated without any complications. He was diagnosed as type 1 diabetes based on the following data. Anti-GAD antibody was positive (3.1 U/ml, normal range 0-1.4 U/ml), fasting serum C-peptide level was 0.1 ng/ml (normal range 0.8-2.5 ng/ml), and urinary C-peptide level was 0.4 [mu]g/day (normal range 22.8-155.2 [mu]g/day). After his recovery from AEN, his previous insulin therapy was replaced to basal-bolus insulin therapy to control the blood glucose level. AEN is a rare and life-threatening disorder. AEN usually develops to the perforation of esophagus, esophageal stricture, and superinfection of mucosa. The prognosis of AEN is largely dependent on the underlying disease, and the mortality rate approximately approaches to more than 30%. The immediate identification of AEN and the accurate management of blood glucose level with continuous insulin administration may lead to successful treatment of AEN without any complications. Taken together, AEN should be considered as one of the differential diagnoses of upper gastrointestinal bleeding in especially patients with diabetic ketoacidosis.[br][br]Nothing to Disclose: MM, TS, MY, TA, SN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 161 56 317 SAT-214 PO47-02 Saturday 256 2012


257 ENDO12L_SAT-215 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Use of Calcium-Stimulated Venous Sampling in Nesidioblastosis So-Young Kim, Ashutosh Pareek, Michael Via Beth Israel Medical Center, New York, NY; Beth Israel Medical Center, New York, NY Objective: To describe the utility of calcium stimulated venous sampling in suspected nesidioblastosis.[br]Case: A 36-year-old obese woman with a history of type 2 diabetes (DM2) and Roux-en-Y gastric bypass (RYGB) surgery five years earlier presented with syncope episodes she described as [ldquo]blackouts[rdquo] for 3 months, occurring at least three times a week. The episodes lasted less than one minute without any warning symptoms or signs of seizure-like activity. During one episode, the patient measured a finger-stick glucose (FSG) of less than 30 mg/dL. All DM2 medications were stopped, however, her episodic symptoms continued.[br]She was admitted to our institution for further evaluation and was given a 5% dextrose infusion. The following day, she again had symptomatic hypoglycemia with FSG of 43 mg/dL that improved with a 25 g dextrose infusion. At this time, patient had elevated serum insulin (6.8 uIU/mL, n[lt]3 uIU/ml), low pro-insulin ([lt]5pmol/L, n[lt]5pmol/L), low c-peptide (0.29 ng/mL, n[lt]0.69 ng/mL), non-detected sulfonylurea, insulin antibody [lt]0.4 U/mL, and low beta-hydroxybutyrate (38 mcg/mL, n[gt]280.8 mcg/mL). Computed tomography of abdomen/pelvis and magnetic resonance imaging of the brain did not demonstrate any masses. She produced a normal response to cosyntropin stimulation. A calcium localization test demonstrated a 2- to 3-fold increase of the insulin levels in the celiac, superior mesenteric, gastroduodenal, splenic, and hepatic artery. The patient was started on acarbose with meals and did not experience any further hypoglycemic episodes. She was subsequently given a 2-month course of parenteral nutrition for suspected malabsorption. She remained euglycemic after the parenteral nutrition was discontinued.[br]Discussion: Nesidioblastosis is an uncommon cause of hypoglycemia that is increasingly recognized following RYGB surgery. The diagnosis is often made clinically and confirmed histologically. In most cases, however, pancreatic surgery is not required. Arterial infusion of calcium followed by venous sampling can aid in the diagnosis of this condition.[br]Conclusion: In patients with prior RYGB surgery, the diagnosis of nesidioblastosis as a cause for episodic hypoglycemia may be established through calcium localization testing.[br][br]Nothing to Disclose: S-YK, AP, MV 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 38 56 318 SAT-215 PO47-02 Saturday 257 2012


258 ENDO12L_SAT-216 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Use of Insulin Pump in Patients with Cystic Fibrosis-Related Diabetes: A Case Series Saket Gupta, Ronan Canavan St Vincent[apos]s University Hospital, Dublin, Ireland Cystic fibrosis related diabetes (CFRD); the most common complication of cystic fibrosis (CF) is associated with 6-fold increase in morbidity [amp] mortality. At present insulin is the only recommended medical treatment for CFRD (1) and multiple insulin injections (MDI) is the most effective regimen (2). Many CF patients don[apos]t adhere to regular insulin injections. A number of case reports and a small non [ndash] randomised study has reported improvement of glycaemic control and quality of life using continuous subcutaneous insulin infusion (CSII) via an insulin pump in the CFRD patients (3). However, there are no guidelines regarding the use of insulin pump therapy as a cost-effective treatment for CFRD.[br]We report our experience of CSII use in a series of CFRD patients at our institute.[br]The first patient, 18 year female, diagnosed with CFRD at age 15, was commenced on MDI in view of catabolic state (BMI 19 kg/m2) and poor glycaemic control (HbA1c 9.3%). She was poorly compliant to MDI and her nutritional status continued to decline despite PEG feed and subsequently CSII was commenced. During a 2-year follow on CSII, BMI increased to 24 kg/m2 however HbA1c failed to improve while FEV1 (69% of predicted) remained stable.[br]The second patient, 18 years male, diagnosed with CFRD at early age required insulin pump therapy because of declining lung function (FEV1, 50% of predicted) and malnutrition (BMI 18.2 kg/m2). The management of CFRD was complicated by his underlying decompensating liver disease. Intermittently he required hospital admissions for the management of complications related to liver disease. Despite serious illness, his BMI remained stable (19 kg/m2) and HbA1c fell (7.4% to 5.5 %). He subsequently underwent successful liver transplantation.[br]Another patient passed away within two years of commencing CSII of end-stage respiratory disease.[br]In summary, the use of CSII in our patients with CFRD resulted in improvement in the metabolic control nutritional status, and stabilisation of lung function. All three patients tolerated CSII with no pump related problems. Use of CSII in patients with CFRD has potential benefits in this complicated group of patients. Previous reports and our experience show that CSII may be a safe [amp] effective therapeutic alternative to MDI treatment in CFRD patients. However indications of CSII in CFRD should be based on individual patient requirement.[br][br]1. Moran A, et al., Diabetes Res Clin Pract 1999; 45: 61[ndash]73. 2. Borowitz D et al., J Pediatr Gastroenterol Nutr. 2002 Sep;35(3):246-59. 3. Hardin DS et al., J Cyst Fibros 2009 May;8(3):174-8. Epub 2009 Jan 21.[br][br]Nothing to Disclose: SG, RC 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2081 56 319 SAT-216 PO47-02 Saturday 258 2012


259 ENDO12L_SAT-217 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Insulin Autoimmune Syndrome Mimicking Insulinoma: A Challenging Diagnosis Francesca Lugli, Donato Iacovazzo, Alessandra Fusco, Antonio Bianchi, Domenico Milardi, Sabrina Chiloiro, Marilda Mormando, Linda Tartaglione, Serena Piacentini, Andrea Giaccari, Alfredo Pontecorvi, Laura De Marinis Policlinico [quot]A Gemelli[quot] - Catholic University, Rome, Italy Insulin autoimmune syndrome (IAS) is a rare condition characterized by hypoglycemia due to autoantibodies against endogenous insulin. It is correlated, in approximately 50% of cases, to specific drugs intake. Hypoglycemia occurs typically during the late post-prandial period.[br]We report 2 cases of IAS patients who came to our attention because of recurrent and severe hypoglycemia.[br]Case 1: a 29 years old female, affected by Graves disease treated with methimazole, was evaluated in 2006 because of recurrent hypoglycemia. Insulinemia was [gt]1000 [micro]UI/ml. Fasting test was negative as well abdomen MRI. After total thyroidectomy glycemic and insulin levels normalized.[br]Case 2: a 45 years old female came to our attention in 2011 because of recurrent post-prandial hypoglycemia. Insulinemia was [gt]1000 [micro]UI/ml. An OGTT revealed late reactive hypoglycemia, while fasting test was normal as well EUS. Dosing of insulin antibodies was 658 U/ml ([lt]0.4). With an adequate diet and acarbose treatment the symptoms improved.[br]In these 2 cases, inappropriately high levels of insulin were associated with late post-prandial hypoglycemia with no evidence of pancreatic lesions and negative fasting tests.[br]In the evaluation of patients with hypoglycemia and inappropriate secretion of insulin, IAS should be considered in the differential diagnosis, especially in patients with late post-prandial hypoglycemia with history of specific drugs intake and negative imaging.[br][br]Nothing to Disclose: FL, DI, AF, AB, DM, SC, MM, LT, SP, AG, AP, LDM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1514 56 320 SAT-217 PO47-02 Saturday 259 2012


260 ENDO12L_SAT-218 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Clinical Characteristics of Fifteen Cases of Fulminant Type 1 Diabetes Mellitus in Our Hospitals Mariko Sue, Aya Yoshihara, Ryo Iga, Shuki Usui, Masahiko Miyagi, Mayumi Ishikawa, Yasuyo Ando, Hiromi Ouchi, Naoki Hiroi, Mariko Higa, Gen Yoshino Toho University School of Medicine, Ota-ku, Japan; Saiseikai Yokohama-shi Tobu Hospital, Tsurumi-ku, Japan [lt]Introduction[gt][br]Fulminant type 1 diabetes mellitus (FT1DM) is characterized by the remarkable acute onset and rapid development of ketoacidosis with nearly total destruction of pancreas b cells, whose etiology is hypothesized to be related to viral infection. We present clinical characteristics of 15 cases of FT1DM we experienced in our hospitals.[br][lt]Cases[gt][br]5 were women and 10 were men. The average age was 42.3 [plusmn] 15.5 year old.[br][lt]Results[gt][br]14 cases revealed acute onset with flu-like and/or abdominal symptoms. The average duration of onset was 4.5 [plusmn] 2.8 days. There was a discrepancy between extremely high plasma glucose concentration (1259 [plusmn] 574 mg/dL) and low levels of HbA1c (6.3 [plusmn] 0.5%). In addition, the serum and urine CPR levels were 0.1 [plusmn] 0.1 ng/mL and 2.4 [plusmn] 3.6 [mu]g/day, respectively, reflecting the severely damaged insulin secretion.[br]Even the relationship between FT1DM and viral infection is frequently suggested, the reports of specific viral infection are few. Though 2 cases revealed elevated herpes simplex virus or measles virus IgM, they did not represent the symptoms related to these viral infections, nor were the DNA levels of viruses negative.[br]In most patients with FT1DM, auto-antibodies are undetectable. However, there also are a few reports of cases with anti GAD Ab positive, or the cases that revealed changes from Ab negative to positive. In our cases, 3 cases were anti-GAD Ab positive, including 2 cases in low titer, and 1 case whose anti-GAD Ab changed from negative to positive, and subsequently changed to negative.[br][lt]Discussion[gt][br]In the first report, the negative status of auto-antibody was one of the sub criteria of FT1DM with the hypothesis that viral infection dominantly plays role in the pathogenesis. However, as many cases with anti-GAD Ab positive are reported, there remains the possibility that some part of autoimmune process contributes to FT1DM.[br]The absence of abnormality in imaging is one of the characters in FT1DM. Only 1 case described the swelling of pancreas in abdominal CT scan, which led to the misdiagnosis of acute pancreatitis. Including this, there were 2 cases misdiagnosed as gastritis or pancreatitis, whose plasma glucose concentration elevated to 1009 and 2230 mg/dL by the administration of inappropriate treatment, including intravenous infusion containing glucose. Though the concept of FT1DM has been accepted more recently, the mistake or delay in diagnosis are not rare even in Japan, where this disease was first reported.[br][br]Nothing to Disclose: MS, AY, RI, SU, MM, MI, YA, HO, NH, MH, GY 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 145 56 321 SAT-218 PO47-02 Saturday 260 2012


261 ENDO12L_SAT-219 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Dunnigan-Type Familial Partial Lipodystrophy [mdash] Case Report Sandra Belo, Angela Magalhaes, Paula Freitas, Cristina Gamboa, Davide Carvalho Centro Hospitalar de S[atilde]o Jo[atilde]o, Porto, Portugal; Centro Hospitalar de S[atilde]o Jo[atilde]o, Porto, Portugal; Universidade do Porto, Porto, Portugal [bold]INTRODUCTION:[/bold] Familial partial lipodystrophy, Dunnigan variety (FPLD), is a well recognized autosomal dominant disorder due to heterozygous missense mutations in lamin A/C (LMNA) gene. The Dunnigan-type familial partial lipodystrophy is characterized by a variable loss of fat in the extremities and trunk and excess subcutaneous fat in the chin and supraclavicular area. Associated metabolic abnormalities include low leptin, insulin resistance and dyslipidemia. [bold]CASE:[/bold] Female, 24 years old, past medical history of acute pancreatitis (1st episode October 2007, complicated with thrombosis of the superior mesenteric vein), combined dyslipidemia, diagnosed in this context, and diabetes, diagnosed in 2009, at first classified as secondary to pancreatitis. She had been referred to Endocrinology appointments but with irregular complacence. On August 2011 she was referred to the Pregnancy Endocrine pathology appointments in the context of 7 weeks pregnancy in a patient with diabetes. At physical examination she presented loss of fat in the extremities and trunk and excess subcutaneous fat in the chin. Weight 58 kilograms. At the first visit the therapy was insulin (initial total dose 106U), heparin and salmon oil. She presented good glycaemic control and triglycerides 1566 mg/dL ([lt]150mg/dL). Anti-GAD antibodies had been already requested and were positive. Dunnigan lipodystrophy was suspected and genetic study was requested. In September 2011 the patient was admitted to the emergency department for acute pancreatitis. Triglycerides were 9975mg/dL. As an inpatient, after improvement of TG with alimentary pause, therapy with fenofibrate was begun. She was then discharged. In October she presented TG of 3000mg/mL. She was then admitted for metabolic control. Therapy was then begun also with omega-3 polyunsaturated fatty acids and concentrated protein supplement after which TG were always around 1000-2000mg/dL. Genetic study revealed the presence of c.1444C[gt]T (p.Arg482Trp) heterozygote mutation in LMNA gene. The patient is now 34 weeks pregnant. [bold]CONCLUSION:[/bold] It is important to recognize the clinical features of this rare lipodystrophic syndrome due to its potentially severe consequences. This patient may benefit, after delivery, from leptin therapy.[br][br]Nothing to Disclose: SB, AM, PF, CG, DC 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2200 56 322 SAT-219 PO47-02 Saturday 261 2012


262 ENDO12L_SAT-220 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Deep Neck Infection in a Diabetic Patient Joana Menezes, Elisabete Rodrigues, Jose Filipe Ramos, Hugo Guimaraes, Margarida Santos, Davide Carvalho Centro Hospitalar S[atilde]o Jo[atilde]o, Porto, Portugal; Centro Hospitalar S[atilde]o Jo[atilde]o, Porto, Portugal Introduction: Deep neck infections may be lethal if life-threatening complications occur, such as descending mediastinitis, pleural empyema, pericardial effusion, epidural abscess, adult respiratory distress syndrome and septic shock, especially in immunocompromised hosts such as diabetic patients(1). The occurrence of deep neck infections has been declining since the advent of antibiotic therapy; however, they do still occur and represent challenging diagnostic and treatment problems(2).[br]Case-Report: A 63-year-old woman with type 2 diabetes mellitus diagnosed 10 years ago and treated with oral antidiabetic drugs (metformin 2000mg/day, acarbose 50mg/day and gliclazide MR 30mg/day), hypertension and dyslipidemia was admitted in Otolaryngology department with neck swelling and redness of the submentonian region, with one week of evolution. She complained of odynophagia, dysphagia and neck pain, without fever, chills, trismus or voices changes. Laboratory results revealed white blood cell count over 12000cells/mm[sup]3[/sup] CPR=430.7mg/L and HbA1c=9.2%. The infection source of deep neck infection was unclear. The patient underwent a neck CT scan that showed a right parapharyngeal abscess with 13mm diameter with slight compression of upper air way, extending up to the lower edge of the right lobe of the thyroid with a longitudinal extent of about 85mm. The abscess involved the right half of the hyoid bone, showing a maximum diameter at this level of 30mm. CT scan also demonstrated densification of soft tissues under the chin region, compatible with inflammatory changes and cervical bilateral lymph nodes. The patient started on insulin therapy with good metabolic control, on intravenous antibiotic therapy (gentamicin and clindamycin) and underwent surgical drainage of the abscess. The results of microbiologic culture of purulent collection from surgery were negative as well as Koch Bacillus test. She presented a good clinical course, with discharge 8 days after admission.[br]Conclusion: Complications of deep neck infections are often life threatening. Clinical evidence and early radiologic diagnosis provide valuable information in defining the origin, location and extension of neck infections. In patients with diabetes mellitus, the clinical course is more severe and there is a poorer prognosis. The combination of appropriate intravenous antibiotic therapy, surgical drainage, securing of airway and good metabolic control are recognized cornerstones of treatment(3).[br][br](1)Mu-Kuan Chen et al, Am J Otolaryngol 2000; 21:169. (2) Silvio Marra et al, Am J Otolaryngol 1996; 17(5): 287. (3) Salih Bakir et al, Am J Otolaryngol 2010; 33: 63.[br][br]Nothing to Disclose: JM, ER, JFR, HG, MS, DC 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1415 56 323 SAT-220 PO47-02 Saturday 262 2012


263 ENDO12L_SAT-221 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Case of New-Onset Type 2 Diabetes Mellitus in a Young Bodybuilder with No Known Family History of the Disease Niyaz Gosmanov, Donny Wynn OUHSC, Harold Hamm Diabetes Center, Oklahoma City, OK; Oklahoma City VA Medical Center, Oklahoma City, OK We present a 41 yo male of Puerto-Rican descent with past history of hypertension (HTN) and hypertriglyceridemia (HTG) has initially presented with diabetes in January 2011 when he was admitted to the hospital for nausea, abdominal pain, weakness and was found to have blood glucose values (BG) in 400[apos]s mg/dl range, triglycerides (TG) of 3200 mg/dl range (with no biochemical evidence of pancreatitis), HgA1c of 11.1% and diagnosed with hyperosmolar hyperglycemic state. He was promptly started on NPH 5 units BID and metformin 500 mg tid and discharged from the hospital with BG readings in 200 mg/dl range. Follow up with PCP resulted in addition of Glyburide 10 mg po bid and titration of NPH up to 10 units twice. Five months after initial presentation, his BGs were ranging 100-150 mg/dl; he had additional testing showing negative anti-GAD or islet cell antibodies and c-peptide of 2.7, insulin of 13 while glucose was 199 mg/dl. He has resumed his bodybuilding routine, though has made substantial dietary modifications with significant reduction of his carbohydrate (CHO) intake resulting in 20 lb weight loss. Twelve months after initial presentation, A1c is 6.1% on Acarbose 100 mg pot tid, Metformin 1000mg bid and occasional regular insulin (for large protein-enriched powder mix routinely used for work-out).[br]It remains unclear which factor dominated in the disease development in the physically active individual with no known family history of diabetes. It might be critically important that the patient[apos]s ethnic background has predisposed him to limited ability to produce insulin on high CHO diet. Of interest, normalization of TG levels has paralleled improved of diabetes control. Patient[apos]s life style and possible use of steroids or growth hormone have been investigated and shown that his Testosterone levels (total and free) as well as IGF-1, LH and FSH were normal and ratios have not indicated signs of exogenous injections; we have incidentally found that patient[apos]s prolactin level was slightly elevated. Patient[apos]s report about four other peer bodybuilders developing diabetes on similar exercise and meal regimens indicated that additional educational effort is needed to increase awareness about possibility of Type 2 diabetes development even among individuals on high diet/high exercise regimen with advice to periodically have lab tests. We stress need for supervision of dietary supplementation content abundantly used by bodybuilders for recreation purposes.[br][br]1. Matthew J Geraci, Mario Cole and Peter Davis., Hum Exp Toxicol 2011 30: 2007.[br][br]Nothing to Disclose: NG, DW 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1670 56 324 SAT-221 PO47-02 Saturday 263 2012


264 ENDO12L_SAT-222 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Long-Term Stabilization of Diabetic Nephropathy and CAD with Triple Therapy of T2 Diabetes with TZD, Metformin and Ramipril Lee Alice Goscin Largo Medical Center, Largo, FL Both ACE Inhibitors and ARBS have shown reduction in microalbuminuria in both type 1 DM and type 2 DM. The Lewis study (1) showed captopril reduced death, dialysis and transplantation in type1 Dm. ARBS had similar benefits in subjects with type2 DM. The Hope trial (2) showed 3577 people with type2 DM over 55 years old with a history of a previous cardiovascular event or at least one other risk factor, 10 mg of ramipril reduced cardiovascular events and mortality by [ndash] 25%. In patients with type 2 DM, hypertension, microalbuminuria and renal insufficiency (Creatinine [gt]1.5 mg/dl), ARBS have been shown to delay the progression of nephropathy. This 77 year old man with triple coronary bypass, 750 mg of Proteinuria and previous Creatinine of 1.5 complicated by hyperkalemia has remained with creatinine of 1.1 and the same proteinuria for 14 years by using triple therapy of rezulin and then rosiglitazone, metformin and ramipril.[br]A second patient, a 58 year old woman with metabolic syndrome, experienced improvement of her Creatinine of 1.5 to 1.1 when started cautiously on metformin 500mg per day. The postulated mechanism may be reduction of endothelial activation and coagulation such as sVCAM, sICAM, and v WF. Twenty years of clinical experience with metformin as a first line medication in spite of creatinine levels slightly higher than1.6 suggests more consideration of higher creatinine clearance is warranted. A third patient remained with the same level of macroproteinuria for 5 years by remaining on her TZD. Indeed the PROACTIVE study (3) showed decrease in death, nonsilent MIs and strokes were reduced by 16%. In addition TZD[apos]s are beneficial in treating NASH and NAFL. These specific three patients suggest more studies are needed with emphasis on starting population and timing as most experienced clinicians practice. Removing existing patients in the United States such as this man from life-saving rosiglitizone is not warranted.[br][br](1) Lewis, N. Engl J. Med.,1963;. (2) Lancet, 2000;. (3) Dormandy, JA et al 2005, Lancet;366: 1279-1289.[br][br]Nothing to Disclose: LAG 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2377 56 325 SAT-222 PO47-02 Saturday 264 2012


265 ENDO12L_SAT-223 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) The Relation between Diabetes Children and Insulin-like Growth Factor 1 (IGF-1) Araz Omer Rasheed University/55, Sulaimani, Iraq Introduction:[br]Insulin-like growth factor 1 (IGF-1) also known as somatomedin C or mechano growth factor is a protein that in humans is encoded by the IGF1 gene.It plays an important role in childhood growth and continues to have anabolic effects in adults.[br]Background:[br]IGF-1 consists of 70 amino acids. IGF-1 is produced primarily by the liver as an endocrine hormone as well as in target tissues in a paracrine/autocrine fashion. Production is stimulated by growth hormone (GH) and can be retarded by under nutrition, growth hormone insensitivity.[br]There is evidence of a metabolic role for IGF-I in type 1 diabetes.[br]In conclusion, IGF-I had a direct effect on glucose and protein metabolism, which was maintained during the hyperinsulinemic euglycemic clamp. This suggests that IGF-I acts in concert with insulin and may have an important role in maintaining glucose homeostasis and protein metabolism in type 1 diabetes.[br]As its name indicates IGF-1, has properties similar to insulin, and it has been shown to improve blood sugar profiles in type 2 diabetics.IGF-1 administration has actually normalized the insulin resistance in a group of healthy volunteers. While in these short term studies IGF-1 therapy did not completely do away with the need for insulin, it did improve the control of blood sugar and so may help prevent the complications of diabetes.[br]Clinical Case:[br]In the present study 40 children were randomly selected at Sulaimani pediatrics hospital aged (1-15) years old and subjected to be tested. The children were divided into two three groups according to their ages [amp]sex [amp]sugar level and IGF-1 and regiments to be teasing.[br]Result:[br]In our result we observed that IGF-1 has no any relation with age[apos]s[amp]sex. The correlation between (duration diabetes was (0.096) and IGF-1) is not significant.[br]And the correlation between (Low level sugar and IGF-1 was (0.14)) and for (high sugar level was (0.191)) is not significant.[br]For the sex the correlation between (males and IGF-1 was (0.837)) and (female [amp]IGF-1 was (0.096)) is not significant.[br]and the Correlation between (Age and IGF-1) is not significant.[br]Conclusion:[br]The obtained result showed that there were no significant effects (age[apos]s[amp]sex) of IGF-1 while it has a powerful rule with abnormality of sugar level.[br][br]Nothing to Disclose: AOR 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 118 56 326 SAT-223 PO47-02 Saturday 265 2012


266 ENDO12L_SAT-224 POSTER SESSION: Diabetes [amp] Insulin Resistance: Uncommon Case Reports (1:30 PM-3:30 PM) Tropical Diabetic Hand Syndrome [mdash] Case Report Paula PB Silva, Gabriella D Arcari, Lenira C Stella, Fadlo F Filho, Isabela TI Carvalho, Renata P Bacchin Beneficencia Portuguesa, S[atilde]o Paulo, Brazil Diabetes Mellitus classically correlates to injuries of the extremeties, including debilitating injuries of hands.Typical diabetic complications related to the duration of the ilness and glycemic control, include infections, Dupuytren[apos]s Contracture syndrome,carpal tunnel syndrome and finger on trigger, more prevalent if diabetics than the general population.Tropical diabetic hand syndrome (TDHS) is a complication that affects patients with DM around the topics. Despite the prevalence of TDHS is still unclear in the literature, is defined as any diabetic adult patient with cellulitis, infection and gangrene. A 36-year-old woman, with DM1 for 35 years, went to the hospital with a necrotic and ulcero-infected lesion in the second chirodactyl of the right hand. She had already an antecedent of amputation of distal phalanx of the third finger on the right hand, after an insect bite. Physical examination revealed blood pressure of 90x50 mmHg, heart beat frequency was of 110/min. A small ulcer on the right second and third metacarpal bones was very warm and red with a purulent secretion. Laboratory test revealed leukocytosis, blood glucose level was 783 mg/dl and her HbA1c was of 12,5%. Ultrasonography of right hand diagnosed osteomyelitis and abscess on the those affected fingers. After glycemic control, anthibiotic therapy and a surgery for excision of the distal phalanx of the finger, she went home. However, after some days, the patient complained of pain, edema on the site of surgery and had the amputation of her second finger metacarpal. TDHS a potentially serious, because it can progress rapidly to gangrene. The predisposition to TDHS is multifactorial and includes uncontrolled diabetes, neropathy and insulin therapy. Antimicrobial tratment should be instituted immediatly based on the culture of biopsy and normally is necessary incision and drainage of the area. Without rapid and agressive treatment can occur permanent incapacity, amputation or death. The TDHS ends up to be neglected, causing delay in forwarding to the hospital. Prevention strategies include education of patients nad professionals for the examination of the hands, and the importance of establishing quick treatment regardless of severity of the injury as soon as the onset of the symptoms.[br][br]F Shiavon et al, Reumatismo 2004;56 (3), 139-142. Papanna et al, J Diabetes and its complications; 2010, 154-162.[br][br]Nothing to Disclose: PPBS, GDA, LCS, FFF, ITIC, RPB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2128 56 327 SAT-224 PO47-02 Saturday 266 2012


267 ENDO12L_SAT-225 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Measurement of Aldosterone in Serum by Isotope Dilution Liquid Chromatography Tandem Mass Spectrometry Yuesong Wang, Gabrielle D Gay, Matthew Gatcombe, Julianne Cook Botelho, Hubert W Vesper Centers for Disease Control and Prevention, Atlanta, GA Aldosterone is a mineralocorticoid hormone secreted from the adrenal zona glomerulosa. Aldosterone secretion has been linked to the pathogenesis of hypertension and related cardiovascular disease. It is a major regulator of body fluid and ion transport, such as sodium reabsorption and potassium excretion, at very low circulating levels.[br]The measurement of aldosterone in serum is used clinically to investigate primary and secondary aldosteronism, and other disorders. Interpretation of the aldosterone level is typically conducted simultaneously with the plasma renin, salt intake, and in the context of upright or supine position. Immunoassay-based methods for measuring aldosterone are widely used in research and clinical diagnosis. However, antibodies can have varying selectivity and poor inter-laboratory reproducibility when measuring aldosterone in serum which limits their use in patient care and public health activities. Liquid chromatography tandem mass spectrometry (LC-MS/MS) can provide highly specific measurements of aldosterone. Herein, a highly specific, high-throughput procedure for measuring aldosterone in human serum by high performance liquid chromatography coupled tandem mass spectrometry was developed.[br]Aldosterone in serum is isolated from the sample matrix using liquid-liquid extraction. Interfering compounds are separated from the analyte using high performance liquid chromatography. Aldosterone is then measured by mass spectrometry. Sample processing is performed in an automated manner using well plates. Sample IDs are scanned automatically and linked to the LC-MS files. The HPLC separation was performed on a C18 column with a gradient and the mobile phases of water and methanol. Aldosterone and its stable isotope labeled internal standard were measured by selected reaction monitoring (SRM) in negative-ion mode. Aldosterone is quantified using the ion transition m/z 359 to 189 and confirmed by the transition m/z 359 to 331. The method performance such as sensitivity, recovery, robustness, and accuracy was well validated.[br][br]Nothing to Disclose: YW, GDG, MG, JCB, HWV 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2181 57 328 SAT-225 PO09-01 Saturday 267 2012


268 ENDO12L_SAT-226 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Ratio of Plasma Aldosterone to Plasma Renin (ARR) Using Two Fully Automated Chemiluminescence Immunoassays Jenny Manolopoulou, Alexandra Bennett, Philipp Grimminger, Evelyn Fischer, Anja Pallauf, Sven Diederich, Martin Reincke, Martin Bidlingmaier Immunodiagnostic Systems Ltd, Boldon, UK; Medizinische Klinik und Poliklinik IV, Munich, Germany; Endokrinologikm Berlin am Gerdanmenmarkt, Berlin, Germany Background: Primary aldosteronism (PA) is reported to occur in 5-10% of the hypertensive population with increased incidence observed in recent years with the introduction of screening using the aldosterone to renin ratio (ARR). Variability in results from commercially available assays means there is a need for method specific cut-off values. We present here preliminary clinical data of two novel chemiluminescence immunoassays for plasma aldosterone (PAC) and plasma direct renin (DR) concentrations on an automated analyzer (IDS-iSYS), comparing results to commonly used assays.[br]Methods: PAC and DR were measured on the IDS-iSYS (Boldon, UK) and compared to the Siemens Aldosterone radioimmunoassay (RIA) and Diasorin Liaison Renin (LSN). 20 healthy controls, 26 essential hypertensives (EH) and 36 PA patients were included. PA was defined by lack of PAC salt-load suppression to [lt]50pg/mL using the Siemens RIA. Mineralocorticoid receptor antagonists and b-blockers were excluded. Hypokalemia was controlled in all subjects. Our laboratory cut-off during screening for PA is 12 (Siemens PAC (ng/L)/Liaison DR (mU/L)).[br]Results: Inter-assay imprecision on the aldosterone iSYS at 68pg/ml - 310pg/ml was at 15% - 4.3%, intra-assay CVs were 12-4%. For the renin iSYS assay, at 10.9mU/L - 309mU/L inter-assay imprecision was at 15% - 3.5%. PAC in controls, EHs and PAs were (mean; 25-75%ile): Siemens: 157(84.2-196), 123(73.3-137), and 274(190-320)pg/ml; iSYS: 115 (50.5-182), 114(68.7-131), and 279(172-348) pg/ml. In the same groups for DR: LSN assay: 33(21.2-43.1), 62(8.9-82.2), and 7.9(2.0-9.5) mU/L; iSYS: 40.1(27.4-47.8), 77.9(13.6-110), and 14.1(4.9-13.7) mU/L. Good correlation was observed for the aldosterone, iSYS=0.947RIA + 0.71, R2=0.71 and renin assays, iSYS=1.057LSN [plusmn] 0.067, R2=0.77. Using our current cut-off for the RIA and LSN at 12, ROC analysis of this cohort gives a sensitivity of 96.7% and specificity of 85.1%. At this cut-off the iSYS assays provided 93.6% sensitivity, so that a lower cut-off at 6.3 would provide corresponding 96.7%/84.8% sensitivity/specificity.[br]Conclusions: Our data demonstrate that the new assays present a convenient practical alternative for the simultaneous measurement of PAC and DR on a single automated analyzer. Comparable discrimination between the three groups and the same diagnostic power could be provided with a lower method specific ARR cut-off. The availability of these automated assays will facilitate screening and diagnosis of PA.[br][br]Disclosures: MB: Study Investigator, Chiasma. Nothing to Disclose: JM, AB, PG, EF, AP, SD, MR 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1052 57 329 SAT-226 PO09-01 Saturday 268 2012


269 ENDO12L_SAT-227 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Role of Adrenal Vein Sampling in Primary Aldosteronism. Impact of Different Diagnostic Criteria on Subtype Diagnosis Maria-Verena Cicala, Anna Patalano, Monica Salva, Diego Miotto, Beatrice Rubin, Raffaele Pezzani, Barbara Mariniello, Franco Mantero Endocrinology Unit, University of Padua, Padua, Italy; Radiology Unit, University of Padua, Padua, Italy In patients with primary aldosteronism (PA), adrenal vein sampling (AVS) is considered the gold standard to distinguish between unilateral and bilateral autonomous production of aldosterone, while diagnostic imaging tests by CT scan or MRI are useful but often inconclusive for the diagnosis of PA. To date agreement is lacking on the best criteria indicating successful cannulation and lateralization.[br]The aim of the study was to evaluate the impact of different diagnostic criteria for the successful cannulation and lateralization on subtype diagnosis and to compare the difference of the findings between adrenal CT scan and AVS.[br]Sixty-seven patients with confirmed PA underwent AVS. The different diagnosis of PA subtype reached using AVS data assessed by more permissive (type 1) and strict (type 2) criteria were compared. Al patients performed CT scan before AVS and imaging results were compared with results of AVSs (using both criteria).[br]Using Type 1 criteria AVSs were successful in 86,3% of patients, while they were successful in only 66,6% using type 2 criteria. Type 1 criteria also led to a higher rate of diagnosis of unilateral PA (85% of successful procedures) than type 2 (75%). There was considerable disparity in the diagnosis reached using the 2 different criteria, with a concordance in only 50% of patients. In conclusion more permissive criteria for successful cannulation and lateralization on AVS can lead to incorrect diagnosis and accordingly to inappropriate treatment options. In the selected group of patients with successful AVS, CT findings correlated with AVSs findings and correctly identified unilateral or bilateral disease in 57,9% of patients using type 1 criteria and in 47,7% using type 2 criteria. Finale diagnosis was based on histological results in 29 patients (43,9%) which underwent adrenalectomy based on AVSs findings. On the basis of CT findings alone 17,5% of patients from the first group and 31,8% of patients of the second group probably would have been incorrectly bypassed as candidates for adrenalectomy. CT scanning lacks sensitivity and specificity and should, therefore, be followed by AVS, which is the only reliable means of differentiating unilateral from bilateral PA and lateralizing APAs preoperatively. However, there are still controversies to be solved by large prospective studies on the criteria to adopt for defining the most appropriate cut off for both correct cannulation and lateralization.[br][br]Nothing to Disclose: M-CC, AP, MS, DM, BR, RP, BM, FM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1252 57 330 SAT-227 PO09-01 Saturday 269 2012


270 ENDO12L_SAT-228 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Evaluation of Reproducibility of Captopril Loading Test in Primary Aldosteronism Masataka Kudo, Ryo Morimoto, Yoshitsugu Iwakura, Yoshikiyo Ono, Ken Matsuda, Akira Sugawara, Sadayoshi Ito, Fumitoshi Satoh Tohoku University Graduate School of Medicine, Sendai, Japan; Tohoku University Graduate School of Medicine, Sendai, Japan [italic]Introduction[/italic]: The plasma aldosterone concentration: renin ratio (ARR) is widely used for the screening of primary aldosteronism (PA), and then the captopril loading test (CAPT) is widely adopted as a confirmatory test of PA. There are many reports concerning about the accuracy of CAPT in PA diagnosis but little is known about its reproducibility. We, therefore, investigated the reproducibility of the ARR in CAPT among PA patients. [italic]Method:[/italic] A total of 48 PA subjects (male: 20, female: 28) who had CAPT 2 times were extracted from our PA patients data base in this study. The first CAPT (50 mg PO) was performed as an outpatient procedure and diagnosed as PA (basal ARR[gt]20 and the ARR in CAPT[gt]20). Successively, adrenal vein sampling (AVS) was performed to ascertain whether unilateral or bilateral excessive aldosterone secretion. The Second CAPT was performed after AVS during hospital stay. Blood samples were obtained before captopril administration, after that at 60min, 90min and 120min. The reproducibility of the ARR in CAPT at each time was assessed with Passing and Bablok regression, coefficient of reproducibility, and Bland-Altman plots. [italic]Results:[/italic] 12/48 patients presented with unilateral excessive aldosterone secretion and were underwent adrenalectomy as aldosterone producing adenoma (APA), whereas 36/48 patients presented with bilateral. The ARR in CAPT at 60min and 90min showed a highly significant within-patient correlation (ARR 60min: r=0.85 P[lt]0.01, ARR 90min: r=0.71 P[lt]0.01) and reproducibility. The ARR at basal and 120min showed a slightly lower correlation (basal ARR:r=0.45, ARR 90min:r=0.52). Bland-Altman plot showed no proportional, magnitude-related, or absolute systematic error between the ARR. [italic]Conclusion:[/italic] It is already reported that the basal ARR was affected by many factors, e.g. posture, time of blood sampling, sodium intake, kinds of drugs, gender and some genetic factors. Nevertheless, the ARR in CAPT showed a good reproducibility. From the points of view of reproducibility, the ARR 60 min was best. These data support that CAPT is a useful confirmatory test of PA.[br][br](1)Gian Paolo Rossi et al.,Hypertension 2010; 55:83. (2)Mohsen Agharazii et al.,Hypertension 2001; 37:1440.[br][br]Nothing to Disclose: MK, RM, YI, YO, KM, AS, SI, FS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1535 57 331 SAT-228 PO09-01 Saturday 270 2012


271 ENDO12L_SAT-229 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Cosyntropin Stimulation Test for the Subtype Classification in Primary Aldosteronism Kazutaka Nanba, Tetsuya Tagami, Tamiko Tamanaha, Kanako Nakao, Yohei Ueda, Maiko Kakita, Mika Tsuiki, Takeshi Usui, Akira Shimatsu, Mitsuhide Naruse National Hospital Organization Kyoto Medical Center, Kyoto, Japan Background: Primary aldosteronism (PA) is the common cause of endocrine hypertension with the prevalence up to 3 to 10% of hypertensive patients. Although adrenal venous sampling (AVS) is the gold standard for subtype classification in PA, it is available only in limited specialized centers. The aim of the study was to investigate the clinical significance of cosyntropin (ACTH) stimulation test for subtype classification in PA. Patients and Methods: Sixty patients with PA who underwent ACTH stimulation test were studied. The subjects were diagnosed as having either unilateral (n=41) or bilateral PA (n=19) based upon AVS, adrenal scintigraphy, and/or adrenal surgery. We evaluated the diagnostic significance of ACTH stimulation test in differentiating unilateral PA from bilateral PA. Results: Peak PAC (P[lt]0.01) and peak PAC/cortisol (P[lt]0.05) after ACTH stimulation were significantly higher in patients with unilateral PA than those with bilateral PA. Receiver operating characteristic curve analysis showed a peak PAC value of 403 pg/ml had a sensitivity of 70.7% and specificity of 79.0%, and a value of 596 pg/ml had a sensitivity of 46.3% and specificity of 100%. A peak PAC/cortisol value of 19.7 (PAC, pg/ml; cortisol, mcg/dl) had a sensitivity of 58.5% and specificity of 89.5% and a value of 30.5 had a sensitivity of 26.8% and specificity of 100%. Conclusions: ACTH stimulation test could discriminate between unilateral and bilateral PA and is useful in selecting the patients who should undergo AVS for localization before surgery.[br][br]Nothing to Disclose: KN, TT, TT, KN, YU, MK, MT, TU, AS, MN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1449 57 332 SAT-229 PO09-01 Saturday 271 2012


272 ENDO12L_SAT-230 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Aldosterone Effect on Erythrocyte Membranes: Comparison with Patients Affected by Hyperaldosteronism and Controls Chiara Sabbadin, Luciana Bordin, Gabriella Dona, Leila Bakdounes, Cristina Fiore, Giulio Clari, Decio Armanini University of Padua, Padua, Italy; University of Padua, Padua, Italy Aldosterone action has been associated with both genomic and non-genomic mechanisms. Aldosterone induces rapid non genomic effects by rising intracellular calcium and cAMP contents, triggering Na[sup]+[/sup]/K[sup]+[/sup] channel exchangers and increasing the activities of protein kinases, such as MAPK and PKC, and NADPH-oxidase. NADPH-oxidase system is the main superoxide generator and is probably involved in the aldosterone induced pro-inflammatory action, maybe in parallel with genomic effects.[br]Taking into account the particular sensitiveness of erythrocytes to the oxidative assault generated by reactive oxygen species (ROS), and their peculiarity as a-nucleated cells, erythrocytes can be particularly useful in the study of the non-genomic effect of aldosterone. In addition, previous studies evidenced that both the association in high molecular weight aggregates (HMWA) and the Tyr-phosphorylation (Tyr-P) level of erythrocyte membrane band 3 can be used as parameters to monitor the redox status of these cells.[br]We studied 12 patients affected by hyperaldosteronism (HA) and 6 healthy controls (HC), evaluating band 3 HMWA formation and Tyr-P levels, both in baseline conditions and after diamide treatment. Diamide is a bland oxidant which is efficaciously utilized to evidence pre-existent membrane alterations induced by oxidative stress.[br]In affected patients, diamide-induced band 3 Tyr-P was higher than controls, with a average increase of 132[plusmn]33%. In addition, also formation of band 3 HMWA induced by diamide was much higher in patients compared to healthy subjects (321.9[plusmn]8.7 compared to 130[plusmn]2%). This was expected due to diamide-induced disulfide bond formation between cysteine residues of band 3. More interestingly, in basal conditions, i.e. in absence of diamide, HA patients showed a net increase in the HMWA content (183[plusmn]8%) compared to HC, thus confirming that membranes from patients were altered.[br]In conclusion, our study demonstrates that aldosterone induces oxidative-like stress in a-nucleated cells, such as erythrocytes, by altering membrane structure. It would be of great interest to investigate this mechanism, through which this compound can regulate this non-genomic effect.[br][br]Nothing to Disclose: CS, LB, GD, LB, CF, GC, DA 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1274 57 333 SAT-230 PO09-01 Saturday 272 2012


273 ENDO12L_SAT-231 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Predicting Factors of the Postsurgical Renal Function in Primary Aldosteronism Kanako Nakao, Kazutaka Nanba, Tamiko Tamanaha, Mika Tsuiki, Tetsuya Tagami, Takashi Usui, Akira Shimatsu, Mitsuhide Naruse National Hospital Organization Kyoto Medical Center, Kyoto, Japan Primary aldosteronism (PA) is the most common cause of secondary hypertension. Although decline in renal function especially that experienced after adrenalectomy (ADX) has been demonstrated, details of the mechanism remain to be elucidated. Aim of the study was to investigate the factors predicting renal outcome after ADX in PA. Fourteen patients with PA and 4 patients with non-functioning adrenal tumor (NFT) as control were studied. Estimated glomerular filtration rate (eGFR), serum potassium, plasma aldosterone concentration (PAC), and plasma renin activity (PRA) before and 1 month after ADX were analyzed. The study was approved by the institutional ethical committee. Baseline characteristics (M[plusmn]SD, PA vs. NFT) were as follows: age; 48[plusmn]9 vs. 65[plusmn]9 (yrs.), systemic blood pressure; 139[plusmn]10 (estimated duration of hypertension: 11.1[plusmn]10.2 yrs.) vs. 114[plusmn]10 (mmHg), eGFR; 92.8[plusmn]24.8 vs. 75.3[plusmn]19.2 (ml/min/1.73 m2), serum potassium; 3.1[plusmn]0.9 vs. 4.4[plusmn]0.3(mEq/L), PRA; 0.3[plusmn]0.3 vs. 1.0[plusmn]0.6 (ng/ml/h), PAC; 342[plusmn]235 vs. 87[plusmn]35 (pg/mL), and ARR; 2216[plusmn]2712 vs. 131[plusmn]102. Twelve PA patients (85.7%) showed decline in eGFR after ADX (92.8[plusmn]24.8 to 71.9[plusmn]24.1, p=0.04). Post-ADX eGFR were correlated negatively to estimated duration of hypertension (p=0.02) and preoperative PAC (p=0.04). There was no significant correlation between the post-ADX eGFR and the patient[apos]s age, blood pressure, serum potassium, and PRA. Chronological change in post-ADX eGFR could be further investigated in 13 PA patients. It was found that eGFR was already declined 1 day after ADX but did not show significant change over the period of 1 year. By contrast to the change in PA patients, there was no change in eGFR in NFT patients after ADX. The present study clearly demonstrated that the post-ADX decline in eGFR is a common and specific complication in PA patients. Duration of hypertension and preoperative PAC are the most important factors predicting the post-ADX eGFR, while there was no further aggravation of the renal function over a period of 1 year. It is therefore concluded that early diagnosis with correction of hypertension and hyperaldosteronism is essential to prevent decline of renal function after ADX in PA.[br][br]Sources of Research Support: Research Grant from National Hospital Organization in Japan for the Clinical Collaborative Study on Primary Aldosteronism.[br][br]Nothing to Disclose: KN, KN, TT, MT, TT, TU, AS, MN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1531 57 334 SAT-231 PO09-01 Saturday 273 2012


274 ENDO12L_SAT-232 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Prospective Multi-Center Study of National Hospital Organization in Japan on the Verification of the Guideline for Primary Hyperaldosteronism (PHAS-J2) Mitsuhide Naruse, Yasuishi Miyazaki, Tsuyoshi Tanaka, Masatoshi Shimizu, Sumire Otani, Shirou Hata, Atsushi Ogo, Kazuya Yonezawa, Kazuo Yoshida, Mamoru Sawamura, Rika Araki, Makito Tanabe, Mika Tsuiki, Tomoko Suzuki, Takurou Shimbo, Study PHASJ2 NHO Kyoto Medical Center, Kyoto, Japan; Misato Kenwa Hospital, Misato, Japan; NHO Mie Chuou Medical Center, Tsu, Japan; NHO Kobe Medical Center, Kobe, Japan; Saitama National Hospital, Wako, Japan; Nagasaki Goto Chuoh Hospital, Goto, Japan; NHO Kyushu Medical Center, Fukuoka, Japan; NHO Hakodate Hospital, Hakodate, Japan; NHO Nagasaki Kawatana Medical Center, Kawatana, Japan; Nishigunma National Hospital, Shibukawa, Japan; National Mie Hospital, Tsu, Japan; NHO Kokura Medical Center, Kokura, Japan; National Center for Global Health and Medicine, Tokyo, Japan Primary aldosteronism (PA) is the major cause of secondary hypertension. The guideline for PA established by the Endocrine Society recommends case detection, confirmatory testing, and subtype classification as the essential process. Given that PA is very common in hypertension, the guideline should be verified in the general practice of hypertension in the city hospital. Aim of the study was to investigate the implementation of each step by the multi-center collaborative study of National Hospital Organization (NHO) in Japan (PHAS-J2). Total 25 NHO and collaborated city hospitals with beds from 280 to 700 participated into the study. The study was approved by the institutional ethical committee. Patients with hypertension (20 to 75 yrs.) were enrolled. Plasma aldosterone to renin activity ratio (ARR) with a cutoff of 20 was used for case detection and captopril test was used as a confirmatory testing. Captopril-positive patients were subjected to subtype classification with adrenal CT, 131I-adosterol scintigraphy, and adrenal venous sampling (AVS). Total 1360 patients were enrolled. Screening test was positive in 21% and captopril test was positive in 58% of the screening-positive patients. Collectively, prevalence of PA was 12.2% of the patients. Adrenal tumor was visualized only in 32% on CT. Implementation of each step was 85% in captopril test, 93% in adrenal CT, 14.9% in adrenal scintigraphy, and 39% in AVS. In Kyoto Medical Center, total 242 patients with hypertension were enrolled. Screening test was positive in 46.7% and captopril test was positive in 65.1% of the screening-positive patients, providing the prevalence of PA as 30.3%. Adrenal tumor was detected in 44.4% on CT. While captopril test and adrenal CT was implemented in 76.1% and 96.4%, respectively, AVS was done only in 25.0% by various reasons. The present study of the NHO in Japan demonstrated that the prevalence of PA with positive screening and confirmatory testing is as low as 10% of the hypertension. In contrast to the screening and confirmatory testing, however, implementation of the subtype classification with AVS was still not high enough. Whatever the reasons underlying the results of the verification, the process of subtype classification requires alternatives as well as further advance in AVS for the standardization of clinical practice of PA in hypertension.[br][br]Sources of Research Support: Research Grant from the National Hospital Organization Japan for the Clinical Collaborative Study of Primary Aldosteronism.[br][br]Nothing to Disclose: MN, YM, TT, MS, SO, SH, AO, KY, KY, MS, RA, MT, MT, TS, TS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1385 57 335 SAT-232 PO09-01 Saturday 274 2012


275 ENDO12L_SAT-233 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Subtype Classification by Computed Tomography and Adrenal Venous Sampling in Primary Aldosteronism Sachiko Tsukamoto Kawashima, Kazutaka Nanba, Kanako Nakao, Yohei Ueda, Maiko Kakita, Tamiko Tamanaha, Mika Tsuiki, Takeshi Usui, Tetsuya Tagami, Akira Shimatsu, Mitsuhide Naruse National Hospital Organization, Kyoto Medical Center, Kyoto, Japan [Background] Subtype classification is the key procedure in determining the therapeutic strategy of primary aldosteronism (PA). Although CT has spread to many hospitals, it doesn[apos]t indicate the function of tumor nor detect microadenoma. By contrast, although adrenal venous sampling (AVS) is the gold standard for subtype classification, it is invasive and available only in limited centers. [Objective] Aim of this study was to compare the subtype classification by CT and AVS and to investigate the influence of different criteria of AVS for lateralization. [Subjects and methods] Forty eight patients with PA proved by confirmatory tests were studied. CT was decided to be positive if the tumor size was [ge]7 mm in its diameter. AVS was performed with ACTH stimulation and 4 different criteria were used for laterality: 1) adrenal PAC[ge]14000pg/ml, 2) PAC ratio of both sides(A/A)[ge]4, 3) lateralized ratio (LR)[ge]4 or 2.6, and 4) contralateral ratio (CR) [lt]1, respectively. [Results] On adrenal CT, 26 patients showed a unilateral adenoma and 22 patients showed normal or equivocal findings. In the CT-positive group, subtype classification by AVS was unilateral in 16 (62%), bilateral in 7 (27%), and contralateral in 3 (11%) with PAC criteria, unilateral in 14 (54%) and bilateral in 12 (46%) with A/A criteria, unilateral in 11 (42%) and bilateral in 15 (58%) with LR criteria[ge]4, unilateral in 13 (50%) and bilateral in 13 (50%) with LR criteria[ge]2.6, and unilateral in 10 (38%) and bilateral in 16 (62%) with CR criteria. In the CT-negative group, subtype classification by AVS was bilateral in 15 (68%) and unilateral in 7 (32%) with PAC criteria, bilateral in 18 (82%) and unilateral in 4 (18%) with A/A criteria, bilateral in 19 (86%) and unilateral in 3 (14%) with LR criteria[ge]4, bilateral in 15 (68%) and unilateral in 7 (32%) with LR criteria[ge]2.6, and bilateral in 20 (91%) and unilateral in 2 (9%) with CR criteria. Numbers of the criteria (PAC, A/A, LR[ge]4, and CR) full-filled in each patient of CT-positive group varied in each patient: 1 (n=16, 61%), 2 (n=14, 54%), 3 (n=10, 39%), and 4 (n=10,39%). The concordant rate of adrenal CT with AVS differed depending on the AVS criteria used. [Conclusions] Concordance of the subtype classification by adrenal CT and AVS depends on its criteria, cut-off, and the number of criteria full-filled, leading to a reversal of the subtype. Subtype classification even by AVS needs further standardization in terms of its diagnostic criteria.[br][br]Sources of Research Support: Research Grant from National Hospital Organization for the collaborative clinical study on primary aldosteronism.[br][br]Nothing to Disclose: STK, KN, KN, YU, MK, TT, MT, TU, TT, AS, MN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 1089 57 336 SAT-233 PO09-01 Saturday 275 2012


276 ENDO12L_SAT-234 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Reevaluation of ACTH Loading in Adrenal Venous Sampling for the Diagnosis of Primary Aldosteronism Akiyo Tanabe, Aya Tsumagari, Mika Tsuiki, Tsuyoshi Tajima, Shuji Sakai, Atsuhiro Ichihara Tokyo Women[apos]s Medical University, Tokyo, Japan; Tokyo Women[apos]s Medical University, Tokyo, Japan; Kyoto Medical Center, Kyoto, Japan [bold]Objectives: [/bold]Corticotropin (ACTH) loading is commonly used in adrenal venous sampling (AVS) for lateralization of primary aldosteronism (PA). The aim of the present study was to reevaluate the diagnostic utility of ACTH loading during AVS. [bold]Subjects and methods: [/bold]AVS data from 127 patients with PA were analyzed. ACTH was administrated by bolus injection (250 [mu]g) in 20 patients and by continuous infusion (125 [mu]g/h) in 107 patients. The adrenal vein (AV) to inferior vena cava (IVC) cortisol (C) ratio [gt]2 before ACTH and [gt]5 after ACTH were used to document successful cannulation of each AV. Unilateral aldosterone excess was confirmed with an aldosterone lateralization ratio (LR) [gt]3; where the affected adrenal AV plasma aldosterone concentration (PAC)/C ratio is divided by the contralateral adrenal AV PAC/C ratio. In addition, the absence of aldosterone secretory autonomy was documented when the contralateral PAC/C ratio in the AV was less than that in IVC (contralateral ratio).[br][bold]Results: [/bold]1) Success rates of AVS. The success rate in the right AV was 60% before and 83% after ACTH loading. The success rate in left AV was 87% before and 97% after ACTH. 2) The concordant rate of lateralization before and after ACTH loading. In the patients with successful bilateral AVS both before and after ACTH (n=68), concordance in lateralization was seen in 69% of the patients. 3) The aldosterone LR. In the patients with proven unilateral aldosterone-producing adenoma (APA) by surgery (n=51), the accuracy rate of AVS lateralization was not different before (82.4%) or after (82.4%) ACTH. A change from negative to positive aldosterone LR was seen in 7.8% of the patients and an increase of this ratio [gt]100% in 23.5% of the patients after ACTH. By contrast, a change from positive to negative aldosterone LR was seen in 7.8% of the patients and a decrease in this ratio by [gt]50% was found in 29.4% of the patients after ACTH. 4) The contralateral ratio. In patients with APA, the accuracy rate of the absence of aldosterone excess from the contralateral AV was increased from 72.5% before to 90.2% after ACTH. The change from negative to positive result was seen in 17.6% of the patients after ACTH; there were no changes from positive to negative results.[br][bold]Conclusions: [/bold]ACTH loading increases the cortisol concentration gradient from the AV to IVC; as a result, it becomes easy to confirm successful AVS. However ACTH loading does not always improve the diagnostic value of AVS in lateralization of PA.[br][br]Nothing to Disclose: AT, AT, MT, TT, SS, AI 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 718 57 337 SAT-234 PO09-01 Saturday 276 2012


277 ENDO12L_SAT-235 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Usefulness of Super-Selective Adrenal Venin Sampling for Detecting Tiny Adrenal Lesions in Primary Aldosteronism Masao Omura, Kohzoh Makita, Seishi Matsui, Jun Inoue, Maki Nagata, Kunio Yamaguchi, Yukio Kakuta, Yoko Matsuzawa, Jun Saito, Tetsuo Nishikawa Yokohama Rosai Hospital, Yokohama, Japan; Tokyo-kita Social Insurance Hospital, Tokyo, Japan; Yokohama Rosai Hospital, Yokohama, Japan; Yokohama Rosai Hospital, Yokohama, Japan; Yokohama Rosai Hospital, Yokohama, Japan Introduction: Primary aldosteronism (PA) is one of the common diseases among hypertensives, and aldosterone-producing adenoma (APA) is usually treated by unilateral adrenalectomy. We had recently developed a new method of supper-selective ACTH-stimulated adrenal venous sampling (SS-ACTH-AVS) for exactly detecting the main lesion(s) of excess aldosterone or cortisol production. Then we attempted to partially remove the unilateral lesion of hyperaldosteronism detected by SS-ACTH-AVS.[br]Methods:SS-ACTH-AVS is done by using a strip-tip type 2.2 Fr micro-catheter (Omura-Makita Catheter; Koshin Medical Inc. Japan), of which the tip possesses three-quarters hall for obtaining easily adrenal effluent at intra-adrenal tributary veins. SS-ACTH-AVS was performed in 50 patients with PA, demonstrating unilateral CT-detectable adrenal nodules (Ult-N). Adrenal effluents were sampled at more than 2 intra-adrenal first-degree tributary veins in each adrenal gland after ACTH stimulation. When concentration of aldosterone in effluent sampled at one of the tributary veins connecting to any nodule was [gt]1400ng/dl and the others were [lt] 1400ng/dl, Ult-N was diagnosed as APA. Then, the patients with APA were treated by laparoscopic unilateral partial resection of the Ult-N without removing adherent normal tissues. When aldosterone was [gt]1400ng/dl in all effluents sampled at tributaries of the unilateral adrenal, showing the presence of Ult-N and was [lt]1400ng/dl in all effluents sampled at each tributary of the contralateral adrenal gland, the patients were also diagnosed as unilateral APA and treated by unilateral total adrenalectomy.[br]Results: Thirty patients, showing Ult-N were exactly diagnosed as APA after functionally detecting the localization by SS-ACTH-AVS, and were treated by unilateral partial adrenalectomy. Pathological examination of resected adrenal glands demonstrated that APA was completely removed without destruction of the capsules nor removing normal tissues. One year after adrenalectomy, concentrations of aldosterone in peripheral blood samples were normalized in 30 patients treated by partial adrenalectomy as well as in 20 treated by total one.[br]Discussion: SS-ACTH-AVS is a quite new method for detecting and localizing tiny adrenal lesions, and is also promising for choosing how to remove the adrenal lesion inducing hyperaldosteroneima, such as unilateral partial and total adrenalectomy.[br][br]Nothing to Disclose: MO, KM, SM, JI, MN, KY, YK, YM, JS, TN 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 881 57 338 SAT-235 PO09-01 Saturday 277 2012


278 ENDO12L_SAT-236 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Validation of the Confirmatory Tests for the Diagnosis of Primary Aldosteronism; an Institutional Experience Takeshi Nigawara, Takako Moriyama, Aya Sugiyama, Shingo Murasawa, Satoshi Yamagata, Yutaka Watanuki, Yuko Tsushima, Shinobu Takayasu, Ken Terui, Satoru Sakihara, Kazunori Kageyama, Toshihiro Suda Hirosaki University Graduate School of Medicine, Hirosaki, Japan The high incidence of primary aldosteronism (PA) within the hypertensive population is widely acknowledged, and a need for efficacious diagnostic procedures has become a topic of discussion. Among various confirmatory tests, upright-furosemide test (Up-Fur), captopril test (Cap) and saline test (Sal) are officially recommended by Japanese Ministry of Health, Welfare and Labor, and positivity in any two of these tests is considered to corroborate the diagnosis of PA. We validated the usefulness of these examinations in our series of patients, together with cosyntropin test ([mu]g iv) without dexamethasone suppression, which is different from the method advocated by Sonoyama et al. (JCEM 2011). Clinical data of one hundred and ninety patients examined in suspicion of PA (plasma renin activity [lt] 2.0 ng/ml/min and plasma aldosterone concentration (Aldo) [ge] 8.0 ng/dl) in our department from January 2008 to June 2011 was collected, and subjected to statistical analysis. For patients taking angiotensin-converting enzyme inhibitors or angiotensin II receptor blockers, calcium channel blockers or alpha-adrenergic inhibitors were prescribed in place of the former antihypertensive agents, prior to endocrinological examinations. A post-injection (30 or 60 min) plasma aldosterone concentration (Aldo) [gt] 20 ng/dl or peak Aldo/cortisol ([mu]g/dl) ratio [gt] 0.85 was tentatively considered as positive for PA in Cos. The basal Aldo in PA group (n=146) was 17.0 [plusmn] 9.1 ng/dl, whereas that in non-PA group (n=28) was 13.0 [plusmn] 5.2 ng/dl. The sensitivity of Cos, Up-Fur, Cap and Sal was 98.6, 92.4, 79.0 and 61.0%, respectively; while the specificity was 25.0, 91.0, 100 and 100%, respectively. This demonstrated that the above criteria in Cos rendered good sensitivity but very poor specificity. Alteration of the criterion to peak Aldo [gt] 30 ng/dl gave specificity of 92.0%, but resulted in a significant decline of sensitivity. Although Cos has an advantage in high sensitivity and the low risk of adverse effects, it may have a limited diagnostic value as a single test. The most useful single examination proved to be Up-Fur, but it is not feasible in elderly patients who have locomotor disabilities. In conclusion, the choice of confirmatory tests for PA should be made according to the patient[apos]s status, in combination with Up-Fur, when possible.[br][br]Nothing to Disclose: TN, TM, AS, SM, SY, YW, YT, ST, KT, SS, KK, TS 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 2018 57 339 SAT-236 PO09-01 Saturday 278 2012


279 ENDO12L_SAT-237 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) The Analysis of 87 Cases with Primary Aldosteronism [mdash] Evaluation of the Confirmatory Tests and Therapeutic Effect Against Atherosclerosis Takamasa Ichijo, Eiko Yoshida, Ayumi Yoshifuji, Kaoru Yamashita, Hiromi Ouchi, Mariko Higa Saiseikai Yokohamashi Tobu Hospital, Yokohama, Japan It is known primary aldosteronism (PA) quickens the atherosclerosis even if the blood pressure is well controlled. Since the guideline was established, we analyzed the usefulness of the confirmatory tests and therapeutic effect.[br]We screened the PA suspicious patients and 87 of those showed ARR[ge]20, which we considered positive. We, then, performed the confirmatory tests, such as rapid ACTH stimulating, captopril challenge, upright plus furosemide, and saline infusion tests, to those patients to diagnose PA. When any of those confirmatory tests were positive, and when surgical treatment is practicable, the diagnosis unilateral hyperaldosteronism (UHA) or bilateral hyperaldosteronism (BHA) was made by ACTH loading-adrenal venous sampling (AVS) after CT scans. We, then, finally perform unilateral laparoscopic adrenalectomy upon AVS documented unilateral hyperaldosteronism. Otherwise, we administrated mineralcorticoid receptor blockers (MRBs), spinorolactone or eplerenone.[br]The mean age was 55.3[plusmn]13.6 years old including 35 males and 52 females, and the mean ARR was 69.1[plusmn]82.0. Twenty-seven and 31 cases out of 65 AVS performed cases showed plasma aldosterone concentration[ge]1,400 ng/dl in either or both side, respectively. We then calculated the lateral ratio by aldosterone/cortisol in higher side to the other lower side, and 18 cases showed the ratio [gt]2.6, which we consider the laterality positive. Our AVS results revealed the mean ARR of UHA was significantly higher than one of BHA, 83.5[plusmn]110.1 vs. 51.7[plusmn]31.2, respectively, as we reported.[br]We next analyzed the anti-atherosclerotic effect of adrenalectomy and MRBs administration by the intima media thickness (IMT) of cervical artery, the brachial-ankle pulse wave velocity (baPWV) and ankle brachial index (ABI) of legs among 20 treated patients followed more than 1 year. Five cases were performed adrenalectomy and 15 cases were administrated MRBs among the AVS performed patients. The annual change of these surrogate makers revealed the adrenalectomy was more effective than MRBs, -0.04 vs. +0.01 mm, -57.33 vs. -36.11 cm/s, -0.03 vs. -0.02, respectively.[br]In conclusion, our results confirmed the usefulness of the multiple measurement of ARR to distinct between UHA and BHA before proceeding to the confirmatory tests, and both adrenalectomy and MRBs administration were effective, not only for anti-hypertensive but anti-atherosclerotic effect, though the former showed better outcome.[br][br]Nothing to Disclose: TI, EY, AY, KY, HO, MH 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 158 57 340 SAT-237 PO09-01 Saturday 279 2012


280 ENDO12L_SAT-238 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Long-Term Outcome Following Adrenalectomy of Primary Hyperaldosteronism Caused by Unilateral Adrenal Hyperplasia Shalini Dabbadi, Jason C Jacob, Faripour Forouhar, Beatriz Tendler, Steven Shichman, Dougald MacGillivray, David Miner, Carl Malchoff University of Connecticut Health Center, Farmington, CT; Hartford Hospital, Hartford, CT; University of Connecticut Health Center, Farmington, CT; Maine Medical Center, Portland, ME; Middlesex Hospital, Middletown, CT Introduction. Primary hyperaldosteronism (PA) is caused by aldosteronomas, bilateral adrenal hyperplasia of the adrenal glomerulosa (BAH), and less frequently by unilateral hyperplasia of the adrenal glomerulosa (UAH). It is not known if UAH is a distinct entity amenable to long-term surgical cure or if it is an asymmetric variant of BAH with a propensity for PA recurrence over time following adrenalectomy. Few studies have carefully examined the long-term outcome of UAH following adrenalectomy. Hypothesis. We test the hypothesis that adrenalectomy produces a long-term cure in patients with PA caused by UAH. Methods. At the University of Connecticut Health Center we performed an IRB approved retrospective analysis of PA patients who underwent simultaneous bilateral adrenal vein sampling and subsequent unilateral adrenalectomy (1996-2010). The pathology was reviewed to identify all patients with UAH. Cure was defined as a post-adrenalectomy serum potassium concentration [gt]4.0 mEq/L without supplementation plus either serum aldosterone concentration (SAC) [lt]7 ng/dl or SAC (ng/dl) to plasma renin activity (PRA) (ng/ml/h) ratio [lt]15. SAC and PRA were measured with the patient sitting, and dynamic testing was not required to confirm normalization of the renin-aldosterone system (RAS). Results. We identified 8 patients with UAH; 1 refused post-adrenalectomy evaluation. Of the 7 available patients, 6 were cured by adrenalectomy. Hypokalemia resolved in the single patient who was not cured by adrenalectomy, but other medical problems precluded safe dynamic testing of the RAS. One patient died from unrelated causes 2 y following adrenalectomy. Pathologically, the adrenal glands of all UAH patients demonstrated triangular glomerulosa hyperplasia with nodular architecture, and 4 had dominant nodules from 0.5 cm to 1.2 cm in diameter. The 6 cured patients were followed for a mean of 8.0 y (range = 2.0 y to 15.2 y) after adrenalectomy, and each remained cured of PA. Discussion. There are limited reports concerning the long-term outcome of UAH patients following adrenalectomy. The complete long-term resolution of all RAS abnormalities in 6 of 7 patients with UAH is most consistent with the hypothesis that UAH is usually a distinct entity and not an asymmetric variant of BAH. Conclusion. Approximately 85 % of UAH patients experience a long-term cure following adrenalectomy. UAH is likely a distinct entity and not an asymmetric variant of BAH.[br][br]Nothing to Disclose: SD, JCJ, FF, BT, SS, DM, DM, CM 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 373 57 341 SAT-238 PO09-01 Saturday 280 2012


281 ENDO12L_SAT-239 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Serum Selenium Concentrations and Hypertension in the German Population Ioanna Gouni-Berthold, Bernhard Michalke, Wilhelm Krone, Eliseo Guallar, Heiner Berthold University of Cologne, Cologne, Germany; German Research Center for Environmental Health, Neuherberg, Germany; Johns Hopkins Bloomberg School of Public Health, Baltimore, MD; Research Groupon Geriatrics, Berlin, Germany Selenium is an essential element with antioxidant properties mediated through various selenoenzymes. Because oxidative stress is involved in hypertension development, it has been suggested that selenium may be involved in blood pressure control and hypertension prevention. The few studies that have investigated the association between serum selenium concentrations and blood pressure have been inconsistent, reporting inverse, null, or positive associations. We explored the relationship between serum selenium concentrations and blood pressure levels and hypertension in the German population. We undertook a cross-sectional analysis of 792 Caucasian subjects (46% male; mean [plusmn] SD age, 53 [plusmn]14 years) who participated in the Lipid Analytic Cologne (LIANCO) cohort. LIANCO was a cohort study designed to assess the relationships between genetic mutations, serum lipoproteins, other biochemical parameters, and clinical data in hypertension, diabetes and atherosclerotic disease. Hypertension was defined as having a systolic blood pressure [ge]140 and/or a diastolic blood pressure [ge]90 mmHg. About half of the cohort was diagnosed as hypertensive. Selenium was measured by inductively coupled plasma-dynamic reaction cell-mass spectrometry (ICP-DRC-MS). Mean [plusmn] SD serum selenium concentration was 68 [plusmn] 32 [mu]g/L, i.e about half of the concentrations seen in the US population. The multivariable adjusted (for sex, age, and body mass index) differences (95% confidence intervals) in blood pressure levels comparing the highest ([gt] 91.9 [mu]g/L) to the lowest ([lt]42.8 [mu]g/L) quartile of serum selenium were 5.2 (1.4 to 8.9), 2.8 (0.7 to 4.8), and 2.4 ([ndash]0.4 to 5.2) mmHg for systolic, diastolic, and pulse pressure, respectively ([italic]P[/italic] for trend for all [lt]0.003). The corresponding multivariable adjusted odds ratio for the presence of hypertension was 1.52 (0.98 to 2.36; [italic]P [/italic]for trend=0.004).[br]The present data suggest that even in a population with very low serum selenium concentrations higher serum selenium concentrations are associated with higher blood pressure levels and a higher prevalence of hypertension. These findings call for careful evaluation of the risks and benefits associated with high selenium concentrations, and may question the rational of the current widespread use of selenium supplements.[br][br]Nothing to Disclose: IG-B, BM, WK, EG, HB 2012-06-23T13:30:00 Expo 2012-06-23T00:00:00 1899-12-30T13:30:00 43 57 342 SAT-239 PO09-01 Saturday 281 2012


282 ENDO12L_SAT-240 POSTER SESSION: Endocrine Hypertension: Renin-Angiotensin-Aldosterone System (1:30 PM-3:30 PM) Low-Sodium Diet Is a Significant Predictor of Increased Catecholamine Levels in Normotensive and Hypertensive Men and Women Aditi R Saxena, Bindu Chamarthi, Ellen W Seely Brigham and Women[apos]s Hospital, Boston, MA Context: Many factors are proposed to affect catecholamine levels, including: age, gender, hypertensive disease and dietary sodium. However, no studies have examined relative effects of these factors within the same study population.[br]Objective: To determine what factors significantly predict both plasma and urinary catecholamine levels.[br]Methods: Hypertensive and normotensive subjects (n=308) were studied in high (250 mEq sodium per day) and low sodium balance (10 mEq of sodium per day). Antihypertensive medications were discontinued in all subjects two weeks before the study. Twenty-four hour urine collections were used to confirm sodium balance and to measure urinary norepinephrine, epinephrine, and creatinine. Plasma norepinephrine and epinephrine were measured in the morning after an overnight fast. Gender, race, age, body mass index (BMI), dietary salt exposure (high salt vs. low salt), hypertension status, and mean arterial pressure were demographic variables of interest. Repeated measures multivariate linear regression models were used to examine the effect of these covariates on plasma and urinary epinephrine and norepinephrine levels. Best-fit lines were obtained using the least-squares method.[br]Results: Of the variables listed above, only the following factors significantly predicted plasma norepinephrine levels: age ([beta]= 2.1, SE=0.8, p=0.005), Black race ([beta]=50.2, SE 23.0, p=0.03), and low salt diet ([beta]=64.9, SE=11.5, p[lt]0.0001). With respect to 24-hour urinary NE, there were a greater number of significant predictors: age ([beta]=0.5, SE=0.2, p=0.005), female gender ([beta]= -10.7, SE=3.5, p=0.002), BMI ([beta]=1.5, SE=0.4, p=0.0009), low salt diet ([beta]=16.4, SE=2.0, p[lt]0.0001). Race did not significantly predict urinary NE levels, controlling for other variables. In contrast, only female gender and low salt diet significantly predicted plasma epinephrine levels, with the following effects: female gender ([beta]= -6.5, SE=2.9, p=0.02) and low salt diet ([beta]=4.8, SE=2.4, p=0.05). With respect to 24-hour urinary epinephrine, only female gender was a significant predictor ([beta]= -4.1, SE=0.9, p[lt]0.0001).[br]Conclusion: In relation to other variables known to affect catecholamine levels, low dietary sodium is the most significant predictor of increased catecholamine levels. These results suggest that multiple factors affect plasma and urinary catecholamine l