心肌高表达mi-99a改善心肌梗死后心室重构及其机制研究

Myocardial infarction is a primary cause of morbidity and mortality in the worldwide. Attenuates the cardiac pathologic remodeling has become the predominant strategies after myocardial infarction. Previous studies has shown that both the MEK1-ERK1/2 and mTOR signaling pathway are activated in response to myocardial hypoxia and hypertrophic stimuli. The activation of MEK1-ERK1/2, increase myocyte thickness, decrease myocyte lengthening, reduce left ventricular dilatation, and improve cardiac function. While, the activation of mTOR, increases cardiac pathological hypertrophy and inflammation, decreases autophagy ,worsen cardiac function. One recent study reveals that MEK1-ERK1/2 can down-regulating mir-99a expression indirectly to increased the expression of mTOR in tumor cells, suggesting that MEK1-ERK1/2 and mTOR interact with each other and reach a blance. To observe and explore the role of mir-99a during the ventricular remodeling, we plan to inject mir-99a plasmid into the myocardium in a mouse model of myocardial infarction, and to suppress the mTOR without the damage of MEK1/2, which contribute to attenuates the cardiac pathologic remodeling after Myocardial infarction. This study will provide the new evidence for theoretical basis and practical applications to improve the prognosis of myocardial infarction.

心肌梗死是危及人民健康的致死性疾病之一,减少病理性心室重构是心肌梗死后治疗的关键。MEK1-ERK1/2、mTOR在心肌缺氧、肥大刺激时均激活。MEK1-ERK1/2激活利于心肌细胞增厚（thickness），抑制心肌细胞延长，减少左心室扩大，改善心功能；mTOR激活增加心肌梗死后病理性肥厚、心肌炎症，减少自噬，促进心功能恶化。最近在肿瘤细胞中的研究发现MEK1-ERK1/2通过下调mir-99a表达间接引起mTOR蛋白表达增加，提示MEK1-ERK1/2、mTOR之间相互作用，相互平衡。申请者计划在小鼠心肌梗死模型中通过心肌内直接注射高表达mir-99a的质粒，抑制mTOR而不减少MEK1/2的表达，使心肌梗死后心肌向有利于减少病理性重构方向发展。研究结果将为临床改善心肌梗死预后提供新的理论与方法。

项目背景：MicroRNAs（miRNAs） 在正常细胞功能的维持中有很多重要的作用，其中包括细胞的凋亡和自噬。该实验的目的是探索microRNA-99a (miR-99a)在心肌梗死后心室重构中的作用及其相关机制。. 主要研究内容：运用实时荧光定量逆转录聚合酶链反应Taqman探针的方法，我们检测了体外培养的乳鼠心室肌细胞在缺氧状态下miR-99a的表达，发现miR-99a在心肌细胞缺氧时表达水平明显下降。因此，我们在在体和离体水平促进心肌细胞高表达miR-99a，观察miR-99a对心肌细胞凋亡和自噬, 心肌梗死后心脏结构和功能的影响。发现在体外培养的乳鼠心室肌细胞中高表达miR-99a能够减少心肌细胞缺氧时的凋亡发生。成年的雄性C57/BL6小鼠行心肌梗死造模时在心肌内直接注射高表达miR-99a（lenti-99a-GFP）和仅含有阴性对照序列（lenti-GFP）的慢病毒，发现术后4周lenti-99a-GFP组的小鼠左心室功能、生存率等较阴性对照组(仅注射阴性病毒lenti-GFP）明显改善。进一步研究表明，miR-99a高表达组心肌mTOR蛋白表达降低,心肌细胞的凋亡减少,自噬增加。. 重要结果：心肌梗死后心肌高表达miR-99a改善心肌梗死动物的生存率和心室重构的进程。这些作用通过减少心肌细胞的凋亡，增加自噬发生等发挥, 由抑制mTOR/P70/S6K信号通路介导。. 关键数据：术后4周lenti-99a-GFP组的小鼠左心室功能、生存率等较阴性对照组(仅注射阴性病毒lenti-GFP）明显改善。. 科学意义：本研究提示在心脏中高表达miR-99a可能成为缺血性心脏病的新的治疗手段。