脱氧胆酸-TGR5信号通路激活在急性心梗中的保护作用机制研究

Acute myocardial infarction (MI) is a severe disease with complex pathological processes, which is still lack of efficient prevention or treatment procedure. Strategies of modulations in metabolic processes have been recently considered as one of the most promising ways to improve the prognosis of acute MI. Our group previously delivered metabolomics detection of plasma samples from patients with different stages of heart failure after MI, and found other than classic glucose and lipid metabolism, also enrichment of metabolites in bile acid synthesis pathways. With further analysis we first discovered the concentration of deoxycholic acid (DCA) decreased significantly with the decrease of cardiac function. Moreover, supplement DCA by gavage could improve cardiac function and alleviate the infarction area significantly in an acute MI mouse model, suggesting that DCA plays a protective role in the process of repair after MI, but the underlying mechanism is unclear. A G-protein coupled receptor TGR5 is one of the most affinitive receptors of DCA, and we also found that cardiac dysfunction was further decline and infarction area was increase in TGR5 knockout mice after MI. The role of DCA-TGR5 signaling pathway in MI has not been reported yet. On the basis of previous results of experiments, in this study we proposed a hypothesis that DCA could bind to its receptor TGR5 to improve acute MI prognosis. We plan to validate this proposal by establishing MI model with TGR5 knockout mice that have already been constructed, aimed to provide novel strategies for the treatment of acute MI.

急性心肌梗死病理机制复杂，目前缺乏有效的防治手段。近年来针对代谢的调控策略被认为是最具有前景的改善急性心梗预后的方式之一。本课题组前期通过收集心梗后不同心功能分级的患者血浆样本进行代谢组学检测，发现胆汁酸合成通路产物的高度富集，并首次发现脱氧胆酸（DCA）随心功能降低浓度显著下降。在小鼠心梗模型中给予DCA灌胃可以改善心功能，使心梗面积明显减少，提示DCA在心梗中发挥保护作用，其具体机制尚不明确。G蛋白耦联受体TGR5是DCA主要结合的受体之一，TGR5敲除小鼠心梗后心功能显著下降，且心梗面积增加，DCA-TGR5信号通路是否有心脏保护作用尚无报道。本课题根据前期预实验结果提出假说，认为DCA与其受体TGR5结合激活下游信号通路改善心梗后损伤修复。本课题拟用已构建成功的TGR5敲除小鼠建立心梗模型探讨DCA-TGR5信号通路激活在急性心肌梗死中的保护作用，旨在为急性心梗的防治提供新的思路。

G蛋白耦联胆汁酸受体TGR5是介导胆汁酸分子生物学效应最主要的受体之一。本项目探讨了胆汁酸分子对TGR5信号通路的激活在急性心梗过程中发挥的功能及其作用机制，通过收集急性心梗患者的血浆进行胆汁酸代谢组学检测，发现心梗组血浆中脱氧胆酸（DCA）水平显著低于健康对照组；使用TGR5敲除（Tgr5-/-）小鼠建立心梗模型，发现与野生型相比，Tgr5-/-小鼠在心梗后7天心功能较差且梗死面积增加，而给予Tgr5-/-小鼠DCA灌胃不能改善心梗损伤情况。对心脏组织进行转录组学分析，结果显示在心梗后1天DCA可以下调由心梗引起的细胞因子产生、细胞因子与受体相互作用、细胞免疫反应等相关信号通路的基因表达；通过Western blot证实DCA抑制心梗后心脏中NF-κB信号通路的激活以及IL-1β的剪切成熟，ELISA结果显示DCA抑制心梗后血浆中IL-1β分泌水平；DCA抑制心肌细胞由缺氧引起的NF-κB信号通路以及IL-1β的激活，通过TGR5信号通路改善缺氧诱导的ROS产生，并减轻心梗后1天的细胞凋亡水平。超声心动结果显示DCA对于心梗后的心功能保护作用可持续至心梗后28天，培养心脏成纤维细胞发现DCA可以抑制LPS引起的NF-κB信号通路激活，Tgfb基因表达下调以及Mmp9基因表达上调。在小鼠建立心梗模型后7天进行组织取材称重，发现Tgr5-/-小鼠脾脏重量与野生型有显著差异；通过流式细胞分析提示TGR5的免疫调节作用可能参与了心梗后的心脏重塑；利用骨髓移植小鼠建立心梗模型，发现移植了Tgr5-/-骨髓的野生型小鼠在心梗后心功能较差且梗死面积增加；利用CD4特异性TGR5敲除（Tgr5ΔCd4）小鼠建立心梗模型，发现Tgr5ΔCd4小鼠相比于对照小鼠心梗损伤加重，心功能较差且梗死面积增加。本研究通过对胆汁酸代谢产物-TGR5受体功能的研究，为心梗的临床治疗提供新的思路及理论依据。