2013 DM Research Grant Report

DR-1

Glucagon-Like Peptide-1 Prevents ß Cells from Apoptosis through Improving Mitochondrial Function and Decreasing ER Stress via Suppressing Sustained AMPK Activation by Methylglyoxal

1TIen-JYun CHang, 1,2HSING-CHI TSENG, 1,2LEE-MING CHUANG

1Department of Internal Medicine, National Taiwan University Hospital, Taipei, Taiwan 2Institute of Molecular Medicine, National Taiwan University Medical College, Taipei, Taiwan

objective: Accumulation of methylglyoxal (MG) contributes to glucotoxicity, and leads to β-cell apoptosis. Glucagon-like peptide-1 (GLP-1) had the protective ability against β-cell apoptosis. However, the molecular mechanism of protecting β-cell from MG-induced apoptosis by GLP-1 remains unclear. In this study, we investigated the signaling pathway involved in the anti-apoptotic effect of GLP-1.

Methods: Rat insulinoma cell line, RIN-m5F cells, was used. MTT assay, annexin V/PI staining, flow cytometry for sub-G1 fraction (DNA fragmentation), western blot of active form caspase 3 and phosphorylated JNK were used as indicators of apoptosis. Extracellular flux (XF) analyses for oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) were applied to estimate the mitochondrial function. Western blots of phosphorylated JNK and eIF2αwere used as indicator of ER stress. results: MG treatment of RIN-m5F cells induced decreased cell viability with increased apoptotic markers. Besides, MG-treated cells were under increased ER stress as revealed with enhancement in phosphorylated JNK and eIF2α. Mitochondrial dysfunction was also found in the MG-treated cells. Pretreatment of GLP-1 conferred anti-apoptotic effects on MG- treated beta cells by alleviating the above abnormalities. Besides, prolonged AMPK activation was found under chronic MG incubation. Pretreatment of GLP-1 improved ATP production and decreased ER stress upon MG treatment, thus reverting cytotoxic effect from the prolonged AMPK activation.

Conclusion: GLP-1 protected β-cell from MG-induced apoptosis through improving mitochondrial function and decreasing ER stress, which may be caused by prolonged AMPK activation upon MG treatment. How this feature of GLP-1 on beta cell function or beta cell mass remains to be further evaluated.

DR-2

The Role of Serum Prothymosin-α in Diabetes Mellitus

Horng-YIH ou

Department of Internal Medicine, National Cheng-Kung University Medical College and Hospital

Insulin resistance, characterized by a reduced ability of insulin to regulate glucose homeostasis in target tissues, is an important pathophysiologic defect in type 2 diabetes. Previous studies indicated that inflammation plays an important role in the development of insulin resistance; inflammatory reactions disrupt insulin signal transduction and further decrease insulin sensitivity. Previous studies also showed that Toll-like receptor 4 (TLR-4), which is responsible for identifying the pathogenassociated molecular pattern (PAMP) of different pathogens in the original immunity against infection, will affect insulin sensitivity. The activation of TLR-4 can impair insulin sensitivity, disrupt the homeostasis of blood glucose, and then lead to development of diabetes.

Prothymosin-α is a cytokine related to immune-modulation and cell proliferation. In immunemodulation, it is known that Prothymosin-α can compensate the defect of mixed lymphocyte reaction, increase the expression of MHC class II antigen, increase the IL-2 receptor expression in T lymphocytes, and enhance the ability of cytotoxicity in natural killer cells and LAK cells, as well as inhibit the growth of tumor by the indirectly increased secretion of TNF-α. Prothymosin-α also has a function to facilitate cell proliferation, and the level of Prothymosin-α is higher in tumor cells than that of normal cells. In addition, previous studies indicated that Prothymosin-α binds to TLR-4 to stimulate the production of interferons in cells and further kill the pathogens; however, the effects of Prothymosin-α on diabetes and insulin sensitivity are still unknown.

Therefore, in this study, we hypothesize that Prothymosin-α may regulate insulin sensitivity and act as a biomarker of diabetes. To test this hypothesis, we enrolled 300 subjects with different glycemic status from the center of health care and diabetes clinic in the National Cheng Kung University Hospital. We found that subjects with newly diagnosed diabetes (NDD) had significantly higher Prothymosin-α levels than those of impaired glucose tolerance (IGT) and normal glucose tolerance (NGT). Moreover, Prothymosin-α, diabetes, body mass index (BMI), and hypertriglyceridemia are independent predictive factors of HOMA-IR after adjustment for multiple confounding variables. Our results suggest that Prothymosin-α might play an important role in the pathogenesis of insulin resistance and diabetes.