SUMO化修饰PPARγ/RXR受体及其cross-SUMOylation抑制糖尿病心肌病氧化应激反应的分子机制研究

Oxidative stress triggered cardiomyocyte apoptosis is an important pathophysiological mechanism of diabetic cardiomyopathy (DCM). And it is the main cause of heart failure and death in patients with DCM. Inhibitory effect of PPARγ/RXR on the expression of NADPH oxidase and iNOS can significantly reduce reactive oxygen/nitrogen species generated and inhibit DCM myocardial oxidative stress. However, the regulation mechanism of these key enzymes is still not clear. SUMO modification of transcription factor plays an important role in the trans-repressor genes expression regulation. Combined with the basis of preliminary studies, we speculate that the SUMOylation of PPARγ/RXR inhibiting of NADPH oxidase and iNOS expression is a key mechanism of their anti-oxidative stress activity. To confirm this hypothesis, we will use RNAi, Co-IP and ChIP to investigate SUMO modification and cross-SUMOylation regulate PPARγ/RXR, which in turn trans-repress the expression of NADPH oxidase and iNOS, inhibit oxidative stress and induce apoptosis and remodeling in high glucose-treated cardiomyocyte in vitro and in vivo SUMO-KO DCM mice model. Exploring this possible mechanism, we can provide new clues to clarify the role of oxidative stress in DCM.

氧化应激触发心肌细胞凋亡是糖尿病心肌病(DCM)重要病理生理机制，是导致DCM患者心力衰竭和死亡的主要原因。PPARγ/RXR抑制NADPH氧化酶和iNOS表达可以显著减少活性氧(氮)自由基生成，减轻DCM心肌氧化应激反应。然而调控这些关键酶表达的分子机制目前尚不清楚。SUMO化修饰在调控转录因子反式阻遏基因表达中发挥重要作用。结合前期研究基础，我们推测SUMO化修饰PPARγ/RXR抑制NADPH氧化酶和iNOS表达是其发挥抗氧化应激作用的关键机制。为证实以上假说，本课题拟通过体外高糖培养心肌细胞和在SUMO基因敲除DCM小鼠模型中，采用RNAi、Co-IP及ChIP等技术明确SUMO化修饰和"cross-SUMOylation"调控 PPARγ/RXR反式阻遏NADPH氧化酶和iNOS表达，从而抑制氧化应激、细胞凋亡与重构的作用并探讨其可能机制，为阐明DCM的氧化应激机制提供新线索。

背景：核受体超家族是一类配体依赖的转录因子，其成员近些年被显示在心血管疾病的发病过程中扮演着极其重要的角色。SUMO化修饰的PPARγ/RXR受体如何参与氧化应激反应的调控，尤其是其信号通路尚不明确。方法：本项目从离体和整体两个水平，采用分子生物学和细胞生物学技术，结合分子荧光探针激光扫描共聚焦显微镜、siRNA干扰、流式细胞检测等实验方法，检测氧化应激反应关键酶系及其下游产物，细胞凋亡的相关指标、活性氧水平等以及通过同位素成像技术和心脏超声评估整体心功能。结果：发现心血管细胞中SUMO化修饰的PPARγ/RXR受体通路是该核受体调控参与氧化应激反应重要机制。NcoR是SUMO化修饰的PPARγ/RXR受体是其调控氧化应激关键酶系的重要靶点，激活该通路能有效抑制氧化应激所导致的炎症反应和细凋亡。同时进一步探索了RXR和其异源二聚体核受体在心肌缺血再灌注损伤中改善氧化应激和细胞损伤的保护作用。结论：心血管细胞中SUMO化修饰的PPARγ/RXR受体通路是该核受体参与调控氧化应激反应重要机制。RXR和其相关的异源二聚体核受体家族是心血管系统重要内源性保护因子，该通路网络的阐明将为治疗心脏疾病提供新的技术路线和筛选新的基因靶点。