TRAP1通过调节线粒体功能及内质网应激对糖尿病心肌病的保护作用及机制研究

Tumor necrosis factor receptor-associated protein 1 (TRAP1), a member of the heat shock protein family, is located predominantly in mitochondria. TRAP1 is found to have essential cytoprotective and antiapoptotic effects under stress. The antiapoptotic mechanisms of TRAP1 are complicated including preservation of mitochondrial function, a quality control on specific mitochondrial client proteins and regulation of endoplasmic reticulum stress. However, the role of TRAP1 in diabetic cardiomyopathy (DCM) has not been explored. Our previous study showed decreased expression of TRAP1 in cardiomyocytes under high glucose condition, accompanied with lower cell viability and increased levels of oxidative stress, which got deteriorated with further inhibition of TRAP1. Based on these prior outcomes, we hypothesize that TRAP1 protected myocardium in DCM by stabilizing client protein Sorcin, inhibiting mitochondrial permeability transition pore (MPTP) opening and relieving endoplasmic reticulum stress. The present project aims to investigate whether TRAP1 can impair myocardial lesion in DCM, both in vitro and in vivo, and to explore the associated mechanisms, by manipulating TRAP1 expression in high glucose condition and diabetic model, for which neonatal rat cardiomyocytes and DCM mice model will be adopted. The results will improve understandings of the pathogenesis of DCM and will provide evidence for new targets of DCM treatment.

TRAP1是一种主要定位于线粒体的热休克蛋白，在应激条件下发挥重要的细胞保护、抗凋亡作用。TRAP1的抗凋亡机制是多方面的，包括维持线粒体功能、参与线粒体中客户蛋白的调控及调节内质网应激等。目前尚未见TRAP1在糖尿病心肌病（DCM）的相关研究。我们的前期研究发现：高糖环境中心肌细胞的TRAP1表达下降，伴随有细胞活性的下降及氧化应激的增强；且这一变化随着TRAP1表达的进一步抑制而更加显著。据此，我们提出假说：TRAP1通过维持客户蛋白Sorcin的稳定、阻断线粒体通透性转换孔（MPTP）开放及减轻内质网应激，发挥对DCM心肌的保护作用。本课题拟采用原代心肌细胞及DCM小鼠模型，通过调控高糖环境及糖尿病模型中的TRAP1表达，从细胞分子、组织及动物整体水平探究TRAP1在DCM中是否有减轻心肌损伤的作用及其作用机制。本研究将有助于进一步阐明DCM的发病机制，并为寻找新的治疗靶点提供依据。

线粒体功能障碍及内质网应激增强诱导的细胞凋亡在糖尿病心肌病(DCM)的发生发展中起着重要作用。Sestrin2是一种重要的氧化应激反应蛋白，被认为参与线粒体功能的维护，特别是在应激条件下。而内质网应激亦与Sestrin2的表达调节密切相关。本研究旨在探讨Sestrin2在DCM中的作用，并探讨其潜在机制。建立体外DCM模型及DCM小鼠模型。通过转染Sestrin2 shRNA及Eupatilin干预等方式，调控Sestrin2的表达。研究发现，DCM细胞活力和线粒体功能受损，内质网应激、细胞凋亡和氧化应激增加。在高糖条件下，DCM小鼠心肌组织和H9c2心肌细胞中的Sestrin2表达显著上调。在初诊2型糖尿病患者中，左心舒张功能功能不全的发生率较高。与心功能正常组相比，合并左心舒张功能功能不全的人群外周血Sestrin2的水平更高。在H9c2心肌细胞中，采用Sestrin2 shRNA直接下调Sestrin2，可提高细胞活力，减少细胞凋亡，减轻氧化应激。且直接下调Sestrin2可减轻高糖诱导的线粒体损伤。采用Eupatilin干预高糖条件下的H9c2心肌细胞，亦可明显提高细胞活力，减少细胞凋亡，减轻氧化应激，且可减轻高糖诱导的线粒体损伤及内质网应激。Eupatilin诱导的Sestrin2表达下降及线粒体融合蛋白2（Mfn2）-PERK通路可能在其中起着重要的桥梁作用。我们的研究结果表明，Sestrin2在糖尿病心肌病的发生发展中起着重要的作用，通过抑制Sestrin2表达的增强，可有效改善DCM心肌损伤，其机制与减轻线粒体损伤及内质网应激相关。