NAD依赖性Sirt5在心肌缺血再灌注损伤中的作用及其对SDH和琥珀酸代谢的调控研究

Aberrant activation of succinate dehydrogenase(SDH) induced succinate metabolic disturbance acts as an important mechanism underlying the myocardial ischemia/reperfusion injury. Sirt5, a NAD-dependent desuccinylase, desuccinylates and inactivates the SDH. In the ischemia/reperfused myocardial tissue, no significant change in the Sirt5 expression is detectable (our pilot research), but NAD is markedly depleted, which leads to the decline of the Sirt5 activity. We speculate that restoring the Sirt5 activity by increasing the NAD content in the ischemia/reperfused myocardial tissue may desuccinylates and inactivates the SDH, consequently may attenuate the ischemia/reperfusion injury through regulating the succinate metabolism. The ex vivo langendorff heart and new born rat primary cardiomyocytes are utilized to simulate the myocardial ischemia/reperfusion injury. The effects of the NAD-dependent Sirt5 on the oxidative injury, inflammatory injury, the release of the cardiac enzymes and the cardiac function are determined. Then, utilizing the gene silence technique, co-immunoprecipitation, protein expression detection and other methods, the interaction of the Sirt5 and SDH, the succinylated level of the SDH and the variance of the succinate level are investigated. After that, the effect of Sirt5-induced succinate level variance on the radical oxygen species production is further determined, as well as the effect on the proteins that regulating the pro-inflammatory factors expression. This research aims to provide a new strategy of protecting the myocardial tissue from ischemia/reperfusion injury.

琥珀酸脱氢酶（SDH）异常激活导致琥珀酸代谢紊乱是心肌缺血再灌注中重要的损伤机制。NAD依赖的去琥珀酰化酶Sirt5降低SDH活性。缺血再灌注心肌组织内Sirt5表达量无显著变化（前期研究发现），但NAD含量降低，Sirt5活性无法维持。因此推测增加心肌组织NAD含量以维持Sirt5活性，可能通过SDH-琥珀酸代谢通路而减轻心肌缺血再灌注损伤。我们采用Langendorff离体灌注心脏和乳鼠原代心肌细胞模拟心肌缺血再灌注损伤模型，探讨NAD依赖性Sirt5对缺血再灌注心肌细胞氧化和炎性损伤和心功能的影响。并采用基因沉默、免疫共沉淀和蛋白表达检测等技术，探讨Sirt5和SDH的相互作用关系、SDH的琥珀酰化水平变化和琥珀酸含量的改变。并进一步探讨Sirt5导致的琥珀酸含量变化对活性氧产生以及炎性因子上游调控因子表达的影响。本研究从物质代谢调节角度为缓解心肌缺血再灌注损伤提供新的思路。