心肌脂滴比较蛋白组学研究及心肌脂滴蛋白CGI-58对心脏功能的影响

Myocardial metabolic disorder especially the aberrant metabolism in triglyceride (TG) hydrolyzation plays a pivotal role in the pathogenesis of cardiac dysfunction. Lipid droplet (LD) is the main organelle in TG storage and metabolism and plays a pivotal role in sustaining intracellular energy homeostasis. LD protein CGI-58 is the key regulator of adipose triglyceride lipase (ATGL) through determining the activity and LD location of ATGL. However, till now, the research about LDs in cardiomyocyte is less well developed, and the effect of CGI-58 on cardiomyocyte lipid metabolism and heart function is not well delineated. This project will explore the role of CGI-58 in cardiomyocyte triglycerol（TG）metabolism as well as in heart function. We have identified the different LD proteins from normal and diastolic dysfunction rat hearts. The preliminary results of proteome and immunoblotting presented that CGI-58 was significantly decreased on LD of diastolic dysfunction rat hearts, suggesting that the reduced CGI-58 may be implicated in the lipid metabolic disorder in cardiac dysfunction. Through constructing heart specific CGI-58 transgenic mouse and manipulating the expression level and location of CGI-58 by using molecular biology techniques, we will detect the variation of the amount and size of LD, the amount and metabolism of TG and fatty acid as well as heart function in vivo and in vitro under conditions of normal and cardiac dysfunction. Using co-immunoprecipitation (Co-IP), fluorescent energy resonance transfer (FERT) and confocal assay, we will investigate the regulation mechanism of CGI-58 on the activity and location of lipid metabolic enzyme ATGL. This research will provide the theory basis for the aberrant lipid metabolism in cardiac dysfunction, and may promote new directions in future research concerning the etiology and treatment of cardiac dysfunction.

心脏能量代谢紊乱与心功能低下密切相关，其中甘油三酯代谢障碍是主要环节。脂滴是细胞内甘油三酯代谢的主要场所，在维持细胞能量平衡中发挥关键作用。而决定甘油三酯水解酶定位及活性的脂滴蛋白CGI-58在心肌脂代谢和心脏功能中的作用还未见阐述。本项目将探索CGI-58对心肌脂代谢和心脏功能的影响及其机制。预实验结果表明正常与心功能不全鼠心肌细胞脂滴上CGI-58定位有显著差异，提示CGI-58表达降低和脂滴定位减少可能与心功能不全心肌能量代谢紊乱相关。我们将利用分子生物学和转基因手段考察增加和干扰CGI-58对心肌细胞和心功能低下动物模型的心肌中脂滴大小数量，甘油三酯含量，脂代谢及心脏功能的影响。并利用免疫共沉淀，荧光共振能量转移和激共聚焦显微镜实时观察CGI-58对心肌细胞脂代谢酶定位及活性的调节机制。本研究将为心功能不全能量代谢失衡提供理论依据，并为心功能不全的防治提供新的思路和靶标。

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，分析心脏在心功能不全和正常状态下脂滴差异蛋白谱，发现ATGL和Dysferlin蛋白在功能不全的心肌脂滴上含量增加。心脏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，将有助于推动脂质异常储存引起的心脏疾病的机理研究。除此之外，还依托本课题开展了血清外泌体差异蛋白谱研究及代谢性血管功能障碍的机制研究。