Hippo/YAP信号调控心外膜祖细胞向冠脉平滑肌细胞分化的机制研究

Coronary smooth muscle cells (CoSMCs) play an important role in the development of coronary heart disease. It is of great clinical significance to recognize the differentiation and regulation mechanism of CoSMCs for the treatment of coronary heart disease. The study indicated that the embryonic epicardial progenitor cell (EpiCs) is the main source of CoSMCs, but the mechanism of specific differentiation and regulation is not clear. The early epicardial membrane of embryonic heart is in low oxygen state. The Hippo signaling pathway key factor YAP is expressed in the embryo epicardial membrane and is closely related to the development of cardiovascular system. YAP can regulate the expression of differentiated genes by combining the key factors of Notch signal, NICD and RBPJ. We hypothesized that the Hippo signal may regulate the differentiation of EpiCs into the CoSMCs via the "YAP-NICD-RBPJ" signal axis in hypoxic microenvironment. Our study intends to reveal the role and regulation mechanism of Hippo signal in the differentiation of epicardial progenitor cells to the CoSMCs, by tracing the differentiation of EpiCs in vivo, constructing conditional epicardial YAP or RBPJ gene knockout mouse model and in vitro hypoxic culture of EpiCs. Thus, new targets are provided for the prevention and treatment of coronary heart disease.

冠脉平滑肌细胞（CoSMCs）在冠心病发生发展进程中发挥着重要作用。识别CoSMCs的分化调控机制对冠心病的治疗具有重要临床意义。研究表明胚胎心外膜祖细胞（EpiCs）是CoSMCs的主要来源，但具体分化调控机制尚不清楚。胚胎心脏发育早期心外膜处于低氧状态。Hippo信号通路关键因子YAP在胚胎心外膜表达，可能参与调控CoSMCs分化。YAP可通过与Notch信号关键因子NICD、RBPJ结合调控分化相关基因的表达。故提出假说：Hippo信号可能通过“YAP-NICD-RBPJ”信号轴调控了低氧微环境中EpiCs向CoSMCs的分化进程。本课题通过构建Tbx18谱系示踪小鼠、条件性心外膜YAP基因及RBPJ基因敲除小鼠模型，以及体外低氧培养EpiCs，在组织、细胞及功能水平分别探索该信号通路在EpiCs向CoSMCs分化过程中的具体调控机制，从而为冠心病的防治提供新的靶点。

研究冠脉平滑肌细胞（CoSMCs）的分化调控机制对冠心病的防治具有重要意义。本课题最初提出假说：Hippo信号可能通过“YAP-NICD-RBPJ”信号轴调控了低氧微环境中胚胎心外膜祖细胞（EpiCs）向CoSMCs的分化进程。为证实上述假说，本课题从以下三个方面进行了研究：（1）生理状态下 EpiCs 向 CoSMCs 分化过程中 Hippo /YAP 信号、 Notch 信号时空表达模式及低氧状态。（2）轴向分别干预各信号因子，观察其对 EpiCs 向 CoSMCs 分化的影响。（3）体外培养 EpiCs 并探讨 YAP 与 Notch 信号关键因子（NICD、 RBPJ）在 EpiCs 向 CoSMCs 分化过程中的相互作用位点。（4）低氧培养 EpiCs 并分别干预“YAP-NICD-RBPJ”信号轴各因子，观察其对 CoSMCs 分化比率及功能的影响。主要研究成果及进展情况如下：（1）构建 Tbx18 谱系示踪小鼠模型，在体示踪胚胎 EpiCs 向 CoSMCs 分化过程，并揭示在体状态下胚胎心脏心外膜的低氧状态。（2）体外培养及研究EpiCs增殖机制，揭示Agrin通过YAP促进胚胎心外膜细胞的增殖。（3）建立自噬的诱导/抑制模型，初步探讨了自噬参与调控EpiCs 向 CoSMCs 分化的机制。综述所述，本课题初步验证假说：Hippo 信号可能通过“YAP-NICD-RBPJ”信号轴调控了低氧微环境中 EpiCs 向 CoSMCs 的分化过程。其具体调控机制，尚需进一步研究。