基因调控心脏祖细胞过表达Cnot3对心梗后心功能修复作用的研究

Coronary artery disease (CAD) and subsequent myocardial infarction are the most common heart diseases, and claim the leading causes of morbidity and mortality. Stem cell therapy has been considered as a promising strategy to repair damaged myocardium caused by infarction. Despite great breakthrough in deriving cardiomyocytes (CMs) from human induced pluripotent stem cells (hiPSCs), the survival and maturation of those cells after transplantation into infarcted myocardium are still the major obstacles hindering their clinical application. Thus, it is critically important to find novel molecular targets to improve the outcomes of stem cell therapy. Cnot3, a component of Ccr-Not complex, is an important factor in embryonic stem cell (ESC) self-renewal. Most importantly, it has been identified as a crucial factor required for normal heart function in an RNAi screen in Drosophila. In addition, our preliminary data suggested that Cnot3 plays a critical role in cardiac development. In particular, Cnot3 induced the proliferation of cardiac cells at the late stage of differentiation, and thereby greatly enhanced the differentiation efficiency of ESC toward cardiac fate. Herein, we hypothesized that genetic manipulation of Cnot3 could increase proliferation of transplanted cardiac cells in infarcted myocardium and improve the cardiac function. To this end, we plan to test our hypothesis by establishing inducible Cnot3 overexpression hiPSC, then injecting hiPSC-derived cardiac progenitor cells (hiPSC-CPC) into infarcted myocardium of immunodeficient mice, and evaluating the improvement of cardiac function.Findings from this study will further our understanding of the molecular basis governing cardiac development, and shed light on how to improve stem cell therapy by genetic manipulation of key factors.

冠心病致心肌梗死后，残存的心肌细胞再生和修复能力低下是导致心力衰竭的重要原因。干细胞移植已被认为是修复梗死心肌，恢复心脏功能的新疗法，但移植细胞在受体内成熟细胞数量少和存活时间短的问题严重影响了治疗效果；因此，探索新的分子靶点以克服以上问题十分必要。研究发现，Ccr4-Not复合物亚基Cnot3不仅是保持ESC多能性的关键，还是维持果蝇正常心脏功能的重要因子；本研究前期结果显示，在ESC或iPS细胞向心脏祖细胞（CPC）分化的过程中，Cnot3的表达先降低后升高，提示其可能在细胞分化发育的中晚期发挥重要作用；进一步研究发现，Cnot3过表达可显著促进CPC的增殖和成熟。基于此，本研究拟采用iPS细胞定向分化为CPC并调控Cnot3过表达；观察其对梗死后心肌细胞的再生和心功能的改善的作用，同时深入探讨心脏功能相关的分子基础，为干细胞治疗冠心病的临床应用提供新思路和奠定理论基础。

冠心病致心肌梗死后，残存的心肌细胞再生和修复能力低下是导致心力衰竭的重要原因。干细胞移植已被认为是修复梗死心肌，恢复心脏功能的新疗法，但移植细胞在受体内成熟细胞数量少和存活时间短的问题严重影响了治疗效果；因此，探索新的分子靶点以克服以上问题十分必要。和目前众多转录调控的研究不同，细胞命运的转录后调控在控制心肌细胞命运方面起到了重要的作用。研究发现，Ccr4-Not复合物亚基Cnot3不仅是保持ESC多能性的关键，还是维持果蝇正常心脏功能的重要因子；本研究结果显示，在ESC或iPS细胞向心脏祖细胞（CPC）分化的过程中，Cnot3的表达先降低后升高，并且Cnot3在拥有高增殖潜能的人或者小鼠心肌细胞中都有高的表达水平，而Cnot3的敲除也能够导致心肌细胞增殖能力的降低。其机制应为Ccr4-Not复合物通过Cnot3依赖性途径与抗增殖基因相互反应，同时介导了他们的降解。最后，在体外培养的人心肌细胞高表达Cnot3能够增强细胞的增殖能力，且在心梗小鼠心脏体外移植模型中起到重要的作用。我们的结论揭示了在调节心肌细胞增殖过程中一个未曾发现的RNA降解调控机制，为心脏细胞生长发育命运及冠心病的治疗上提出了特殊的分子靶点Cnot3，为细胞治疗冠心病的临床应用提供新思路和奠定理论基础。