骨骼肌脂滴比较蛋白组学研究及骨骼肌脂滴蛋白OXPAT对胰岛素抵抗的影响

The unbalanced neutral lipid metabolism in skeletal muscle is closely associated with the development of Type 2 Diabetes Mellitus (T2DM).Lipid droplet(LD)-associated PAT family proteins play a pivital role in regulation of the activity and location of lipid metabolic enzymes.However, till now our understanding of the role of LDs in regulating metabolism in skeletal muscle is less well developed, and the function of PAT protein - - OXPAT in regulating insulin action in skeletal muscle of obese and diabetic individuals has not been well delineated. This project will explore the role of OXPAT on skeletal muscle triglycerol(TG) metabolism and insulin sensitivity. We have identified the comparison proteome of skeletal muscle LD from normal and db/db mice. The primary results of proteome and immunoblotting presented that OXPAT was significantly decreased on LD, which suggested that the reduced OXPAT may be implicated in the lipid metabolic disorder in skeletal muscle. By overexpression and interference of OXPAT in skeletal muscle cell and db/db mouse skeletal muscle tissue, we will observe the changes of the insulin signal and the amount of TG DG and ceramide. Using Co-IP, FERT and confocal assay,we will investigate the regulation of OXPAT on the activity and location of lipid metabolic enzymes. This research will provide the theory basis for the unbalanced lipid metablism in T2DM skeletal muscle, and may promote new directions in future research concerning the etiology and treatment of T2DM.

骨骼肌脂代谢不平衡与2 型糖尿病（T2DM）的发生发展密切相关，脂滴PAT家族蛋白对脂代谢酶的定位与活性起关键调节作用，但骨骼肌主要的PAT家族蛋白- - OXPAT在骨骼肌脂代谢和胰岛素抵抗中的作用还未阐明。本项目将探索OXPAT对骨骼肌甘油三酯代谢，脂滴的大小和胰岛素抵抗的影响及其机制。预实验结果表明正常与糖尿病小鼠骨骼肌脂滴上OXPAT的定位有显著差异，提示OXPAT的表达降低和脂滴定位的减少可能与T2MD时骨骼肌的脂代谢紊乱相关。接下来我们将在糖尿病骨骼肌细胞和糖尿病动物模型的骨骼肌中增加和干扰OXPAT的表达，考察血糖血脂及胰岛素敏感性的变化，脂滴大小及脂代谢变化。并利用免疫共沉淀、荧光共振能量转移和激光共聚焦实时观察OXPAT在骨骼肌中对脂代谢酶定位及活性的调节机制。本研究将为T2DM骨骼肌脂代谢失衡提供理论依据，并为T2DM的防治提供新的思路。

脂滴是一种几乎在各种生物中普遍存在的细胞器。它在脂质合成、储存、 转运和代谢中发挥着重要的作用。脂滴蛋白是脂滴功能的重要执行者，本项目从解析脂滴蛋白的角度出发，分析不同脂肪酸处理的骨骼肌细胞，心脏、肝脏、棕色脂肪等组织在不同状态下的脂滴差异蛋白谱，另外，我们通过差速离心的方法，在小鼠肝脏中成功分离并得到了中性脂成分不同的脂滴。这些有关心脏、肝脏和棕色脂肪组织脂滴分析研究结果为今后的脂滴功能研究提供了重要的方法和信息，尤其将有助于推动脂质异常储存引起的代谢性疾病和心血管疾病的机理研究。此外，还依托此课题进行了一系列代谢性疾病及脂质异常导致的血管功能障碍的机制研究。