ROS在调控心肌衰老过程中Beclin 1-Vps34复合体功能和自噬流的机制研究

In spite of advances in preventive and therapeutic strategies, cardiovascular disease (CVD) remains the leading cause of death and disability in the elderly.Heart beat need lots of oxygen and produce a large amount of Reactive Oxygen Species (ROS), cause a high level oxidative stress to the organ. Autophagy is the basic catabolic mechanism that involves cell degradation of unnecessary or dysfunctional cellular components through the lysosomal machinery. However, it is down-regulate in the aging. Beclin-1 and its binding partner class III phosphoinositide 3-kinase (PI3K), also named Vps34, are required for the initiation of the formation of the autophagosome in autophagy. How oxidative stress regulate Beclin 1 complex component and function is not clear yet. Based on our previous research, we are going to study the relationship between autophagy and cardiac aging. Therefore, we propose to re-establish the balance in the redox status in the aging heart by TEMPO. We are going to use western blot, Quantitative PCR, confocal microscopy, RNA interference and knock-in mouse to study the mechanism. Mitochdrial ATP consumption, O2 consumption and ROS produce will also be used to address how oxdiative stress regulate Beclin 1 complex to affect autophagy in the aging heart. In toto, our study will provide mechanistic insight into the cardioprotection of elderly and underscore its protection by antioxidant treatment where could be a therapeutic strategy.

心脏在耗氧同时产生大量自由基，造成氧化应激增加，而老年机体自噬功能下调。Beclin 1 是自噬的特异性基因，通过与不同分子相互作用形成复合体而发挥作用。衰老过程中的氧化应激如何调节Beclin 1复合体的结构和功能需要进一步研究。本项目在既往工作基础上将探讨心肌衰老与自噬间的交互调节作用。将用TEMPO抑制超氧离子从而降低老年大鼠机体氧化应激，通过（1）western、荧光定量PCR、免疫共沉淀和共聚焦显微镜阐述Beclin 1-Vps34复合体结构和功能变化。（2）测定长半衰期蛋白，运用mCherry-GFP-LC3结合RNAi、基因敲入等阐述氧化应激对自噬潮的调节作用。（3）验证抗氧化处理后老年心肌抗缺血再灌输损伤能力，线粒体ATP、O2耗量、ROS产量、心肌组织抗氧化能力，探讨自噬对心肌衰老的调控作用。本研究成果可为老年心肌保护提供新的思路，并可横向推广到其他衰老相关的疾病中去。

心血管疾病发病率随年龄增长而明显增加，心脏在耗氧同时产生大量自由基，造成氧化应激增加，而老年机体自噬功能下调。Beclin 1 是自噬的特异性基因，通过与不同分子相互作用形成复合体而发挥作用。衰老过程中的氧化应激如何调节Beclin 1复合体的结构和功能需要进一步研究。本项目在既往工作基础上将探讨心肌衰老与自噬间的交互调节作用。本研究通过（1）检测对不同氧化应激态下老年大鼠心肌组织Beclin 1-Vps34复合体功能。（2）检测不同氧化应激态下老年大鼠心肌自噬潮的特点。（3）检测不同氧化应激态下老年大鼠心肌的抗氧化能力和线粒体功能。探讨自噬对心肌衰老的调控作用。.我们发现老年大鼠心肌氧化应激压力明显高于成年大鼠，心肌损伤后自噬潮通路受阻，受损线粒体的清除功能较成年大鼠明显不足。这些原因可能是由于老年大鼠Beclin 1-Vps34复合体功能下降，Beclin1 的分子交互作用发生改变，凋亡通路异常活跃而引起。.本项目的实施阐明ROS与衰老心肌自噬的交互作用。论证自噬在抗氧化治疗改善老年心肌易损性的地位和作用。从而形成进一步指导临床抗氧化治疗的策略，为衰老性疾病的防治工作提供新的思路，具有较高的临床价值。并可横向推广到其他衰老相关的疾病中去。