脂肪组织ClC-3容积调节性氯通道在胰岛素抵抗中的作用与机制研究

以前脂肪细胞凋亡为中心的脂肪组织更新障碍是胰岛素抵抗的重要起始环节。我们前期研究在ClC-3基因敲除2型糖尿病小鼠等整体模型上发现，脂肪组织ClC-3容积调节性氯通道与胰岛素抵抗发生相关；体外实验发现ClC-3参与饱和游离脂肪酸诱导前脂肪细胞凋亡。本项目拟在已有工作基础上，应用ClC-3 siRNA和ClC-3 cDNA全长转染技术, 阐明ClC-3容积调节性氯通道在饱和游离脂肪酸诱导前脂肪细胞凋亡中的作用和机制；结合不同机制2型糖尿病模型 （包括ClC-3基因敲除和自发性肥胖2型糖尿病小鼠等），明确ClC-3容积调节性氯通道能否通过调控前脂肪细胞凋亡参与胰岛素抵抗发生，并揭示其与胰岛素抵抗过程中脂肪组织增殖与分化、炎症和胰岛素信号通路变化的内在联系。本研究将从ClC-3容积调节性氯通道这一新角度，揭示胰岛素抵抗发生新机制，并为寻找改善胰岛素抵抗的新药靶提供实验室依据和新思路。

The enhanced apoptosis in preadipocyte of type 2 diabetes patients has been shown in the recent work. Our preliminary study have shown that ClC-3 volume-regulated chloride channel is associated with the insulin resistance by using ClC-3 KO mice subjected to type 2 diabetes mellitus (T2DM). The ClC-3 KO mice have indicated the significant reduction in HOMA-IR and serum glucose levels, but no significant change in serum insulin levels.These data suggested that ClC-3 should be involved in the development of insulin resistence, but not be interpreted by the modulation of insulin secretion.We further found that there is an increase in the ClC-3 protein expression in adipose tissue in T2DM rat models induced by high-sucrose-high-fat diet combined with a low dose of STZ,and which has a positive correlation with HOMA-IR levels,but such correlation could not been found in liver and skeletal muscle. In 3T3-L1 preadipocyts, we have shown that palmitate, the main component of saturated free fatty acids(FFA), induced an increase in ClC-3 protein expression, in parallel with promoting preadipocyte apoptosis in a concentration- dependent manner, ClC-3 siRNA silencing significantly reversed the above both change induced by palmitate.These preliminary data suggested that ClC-3 chloride channel may play a role in free fatty acid-related preadipocyte apoptosis. Our research group has been focused on the role of ClC-3 volume-regulated chloride channel for two decades. We have shown that ClC-3 channels are important mediator contributing to cellular volume regulation, proliferation and apoptosis and thereby accounting for many pathological processes in response to volume stress,such as hpertension,stroke and atherosclerosis. Enlarged adipocytes is taken as an independent predictor for T2DM, and which is mainly due to reduced differentiation ability of preadipocytes or the enhanced apoptosis of preadipocytes. Based on our preliminary data and the suitable molecular tools including ClC-3 KO mice, we will investigate the role and the molecular mechanism of ClC-3 chloride channel during obesity-related insulin resistance. The main objectives of this project are: 1) to investigate the role and molecular mechanisms that ClC-3 volume-regulated chloride channel in FFA-induced preadipocyte apoptosis;2) to examine whether ClC-3 volume-regulated chloride channel is therefore involved in adipose tissue dysfuntion and IR developmemt by using several in vivo T2DM models(including KKAy and/or db/db mouse model, ClC-3 KO T2DM mouse model); and the related signaling pathway,such as preadipocyte apoptosis, proliferation and differentiation ability,inflammation and insulin signal in adipose tissue. The novelty of this proposal is to identify ClC-3 volume-regulated chloride channel as a novel molecular mechanism for adipose tissue derived insulin resistance, and will help in the search of new drug target for improving insulin sensitivity.