Cytokines Increase Global DNA Methylation While Altering Insulin and Amylin Expression  in Human Beta Cells

Presentation Number: LB MON 77
Date of Presentation: April 3rd, 2017

Jacqueline Pena Velasco*, Michael G Spelios, Lauren A Kenna, Siham D Accacha, Regine Tipon and Eitan M Akirav
Winthrop University Hospital, Mineola, NY

Abstract

Both Type 1 and Type 2 diabetes are characterized by progressive beta cell failure. The actual mechanism leading to beta cell destruction is unclear. Cytokines are inflammatory mediators implicated in the loss of functional beta cell mass during development of Type 1 and Type 2 diabetes. Epigenetic modification such as DNA methylation, regulates gene expression without altering the DNA sequence. DNA methylation is crucial to beta cell differentiation and function. To investigate the effect of long-term exposure of human beta cells to cytokines, EndoC-Bh1 cells, a human beta cell line, were cultured in the presence or absence of a cytokine cocktail (TNF-α 0.005 µg/ml, IFN-Ƴ 0.03 µg/ml, and IL-1β 0.0006 µg/ml) for 14 days. Cells were harvested at day 5-, 10- and 14-day time points. Gene expression was measured using real-time PCR. Global DNA methylation was investigated using Epigentek Methylflash Methylated DNA Quantification kit. Our results showed reduced insulin gene expression in EndoC-Bh1 cells cultured with cytokine cocktail while insulin gene expression gradually increased in the control group over time (Day 14: control=14.73 ± 3.580 vs cytokine=4.000 ± 1.361 au; p=0.0487; N=3). On the other hand, amylin gene expression was increased in EndoC-Bh1 cells treated with cytokine cocktail, while amylin gene expression was low throughout the treatment duration in the control group (Day 14: control=0.0210 ± 0.005686 vs cytokine=1.202 ± 0.3927 au; p=0.0397; N=3). Global DNA methylation was significantly elevated in EndoC-BH1 cells cultured with cytokines compared with control (Day 14: control=0.04870 ± 0.001881 vs cytokine=0.1377 ± 0.01584; p=0.0051; N=3). We conclude that long-term exposure of human beta cells to cytokines as a model of inflammatory stress in diabetes may lead to increased DNA methylation. Increased global DNA methylation is a likely mechanism that alters gene expression in beta cells leading to cell dysfunction. Our result which showed reduced insulin and increased amylin gene expression is an indicator of beta cell death. This data may provide a basis for future therapies for improving beta cell function and survival in patients with diabetes by modifying DNA methylation.

 

Nothing to Disclose: JPV, MGS, LAK, SDA, RT, EMA