人胚胎干细胞向心房肌细胞和心室肌细胞定向分化的转录因子调控网络

Utilizing cardiomyocytes derived from human embryonic stem cells has promising application perspectives in the fields of drug development and cell transplantation therapy of myocardial infarction. The discovery of key transcription factor network, from embryonic stem cells to atrial and ventricular muscle cell, can not only improve the differentiation procedure of embryonic stem cells, but also reveal the pathogenesis of congenital heart disease. In our previous work, we have established a basic protocol for direct differentiation of embryonic stem cells into atrial and ventricular myocytes. We will take advantage of the developed differentiation system, combined with the sequencing of single cell gene expression method (single cell RNA-seq), to compare early atrial and ventricular myocytes differences in gene expression. Meanwhile based on the occurrence frequency of DNA transcription factors' binding sites by bioinformatics analysis, we want to find the distinct transcription factors regulatory network in atrial and ventricular myocytes respectively. Furthermore, we will use over-expression and RNA knock-down techniques to validate these key regulation factors, finally unveil the molecular mechanisms of atrial and ventricular myocytes differentiation.

利用人胚胎干细胞分化获得人心肌细胞对药物的研发和利用细胞移植治疗心肌 梗死具有重要意义。了解和发现从胚胎干细胞到心房和心室肌细胞分化过程中关键转录因子 网络不但可以完善胚胎干细胞的分化方法，也对揭示先天性心脏病的发病机制有重要的指导 意义。在前期工作中我们已经建立了一个可将胚胎干细胞分化成心房肌细胞和心室肌细胞的 定向分化培养体系。我们将利用已建立的心肌定向分化系统，结合单细胞基因表达测序的方 法(single cell RNA-seq) ，比较早期心房肌细胞和心室肌细胞基因表达的差异，并通过生 物信息学分析转录因子的 DNA 结合位点出现的频率为依据发现调节心房肌细胞和心室肌细 胞分化的转录因子调控网络，同时利用过表达和 RNA knock-down 技术来发现和验证在心肌 前体细胞向心房肌细胞和心室肌细胞分化的关键调控因子，从而揭示心房肌细胞和心室肌细 胞的分子机制。

利用人胚胎干细胞分化获得人心肌细胞对药物的研发和利用细胞移植治疗心肌梗死具有重要意义。了解和发现从胚胎干细胞到心房和心室肌细胞分化过程中关键转录因子网络不但可以完善胚胎干细胞的分化方法，也对揭示先天性心脏病的发病机制有重要的指导意义。在前期工作中我们已经建立了一个可将胚胎干细胞分化成心房肌细胞和心室肌细胞的定向分化培养体系。本项目中，我们通过人及非人类灵长类诱导性多能干细胞的分化体系，进一步验证了RA信号通路在心房定向分化中的重要作用，探讨了BMP信号通路在心室肌细胞特化中的作用机制。通过microarray的方法比较了RA和BMP处理得到的心房肌细胞和心室肌细胞基因表达的差异，并通过生物信息学方法预测了心房肌细胞和心室肌细胞分化的转录因子调控网络，从而为揭示心肌前体细胞向心房肌细胞和心室肌细胞分化分子机制提供潜在的靶标。