细胞自噬在氧化三甲胺（TMAO）促进动脉粥样硬化斑块易损性中的作用及机制研究

The rupture of vulnerable atherosclerotic plaque may lead to acute vascular events, even death. It is very important to increase plaque stability. We have found that the level of plasma TMAO in patients with acute myocardial infarction was significantly higher than that of normal people in clinical study. The plasma TMAO level in patients with vulnerable plaque was significantly higher than that in patients with stable plaque. Further, it was found that TMAO could decrease the repair function of endothelial cells and promote the adhesion of monocytes. TMAO could also reduce the level of autophagy in smooth muscle cells. The level of autophagy is one of the factors that affect the instability of plaque. Therefore, this study intends to investigate whether TMAO can promote the vulnerability of plaque by inhibiting autophagy pathway. Our research team will measure plasma TMAO levels in patients with stable or vulnerable atherosclerotic plaques by mass spectrometry. We will study the stability of atherosclerotic plaque in Beclin-1 and ApoE double-knockout mice, and mechanism of TMAO effect on autophagy in plaque macrophages, smooth muscle cells, endothelial cells and plaque stability by cell experiment. We investigate the role and mechanism of TMAO in promoting the vulnerability of plaque by inhibiting the level of autophagy integrated with clinical, animal and cellular levels.

动脉粥样硬化易损斑块破裂导致急性血管事件甚至危及生命，故增加斑块稳定性有至关重要临床意义。本课题组前期临床研究发现急性心肌梗死患者血浆TMAO水平明显高于正常人，动脉粥样硬化易损斑块患者血浆TMAO水平比稳定斑块患者明显升高。进一步通过细胞实验发现，TMAO可降低内皮细胞的修复功能，并且促进单核细胞粘附。TMAO会降低斑块平滑肌细胞自噬水平，而细胞自噬水平是影响斑块不稳定因素之一。故本研究拟探讨TMAO是否可能通过抑制相关细胞自噬途径促进斑块易损性。我们将通过质谱测定存在稳定或易损动脉粥样硬化斑块患者的血浆TMAO水平，观察Beclin-1、ApoE基因双敲除小鼠动脉粥样硬化斑块稳定性，并通过细胞实验探讨TMAO对斑块巨噬细胞、平滑肌细胞、内皮细胞自噬和斑块稳定性的影响和机制。结合临床患者、动物和细胞水平探讨TMAO通过抑制相关细胞自噬水平而促进斑块易损性的作用及机制。

动脉粥样硬化是导致心血管疾病的主要原因之一，而肠道菌群代谢产物TMAO在动脉粥样硬化发生发展进程中具有潜在的助推作用。本课题组通过大量临床研究发现，血浆TMAO随年龄增长呈升高趋势；女性血浆TMAO水平略高于男性；血浆TMAO水平与冠心病、糖尿病、体质量指数（BMI）、总胆固醇、颈动脉斑块的易损性及斑块负荷呈正相关，与高密度脂蛋白呈负相关；冠心病合并脑血管狭窄与血浆TMAO水平升高无关。TMAO前体物质胆碱、肉碱、三甲胺乙内酯和三甲氨基丁内盐的浓度与冠心病无显著相关性，甜菜碱与低密度脂蛋白呈负相关。另外，我们发现瑞舒伐他汀治疗后的TMAO水平较低，但TMAO前体水平较高。本课题组通过动物实验探索FMO3基因敲除对左旋肉碱（L-carnitine）代谢为TMAO的影响，以及TMAO在促进动脉粥样硬化中的作用，结果发现，FMO3基因敲除后，L-carnitine代谢为TMA及TMAO均有升高趋势；TMAO能升高血压、增加血管僵硬度，影响心功能（降低心输出量和每搏输出量），能增加内皮细胞凋亡，促进动脉粥样硬化。课题组在动物实验基础上，通过体外细胞实验，发现TMAO能抑制细胞自噬，增加内皮细胞凋亡。本课题组进一步通过体外内皮细胞实验，通过自噬抑制剂和过表达beclin-1探索细胞自噬与凋亡在TMAO促进动脉粥样硬化中的作用机制，并探索其作用通路。本项目的发现为动脉粥样硬化不稳定斑块的相关机制提供依据，为临床提供可能的干预靶点。