低氧/HIF对小鼠成纤维细胞直接重编程为心肌细胞的作用及机制的研究

项目名称： 低氧/HIF对小鼠成纤维细胞直接重编程为心肌细胞的作用及机制的研究

项目编号： No.31271287

项目类型： 面上项目

立项/批准年度： 2013

项目学科： 生物科学

项目作者： 刘慧雯

作者单位： 哈尔滨医科大学

项目金额： 80万元

中文摘要： 直接诱导成纤维细胞重编程为心肌细胞有望为心肌细胞移植提供新的细胞来源。细胞微环境不仅支持细胞发育，也可以通过影响细胞的极化来调节改变信号转导通路从而指导细胞的分化方向。本研究拟构建αMHC启动子驱动的EGFP标记转基因小鼠，以确保只有定向分化的心肌细胞才能表达GFP。通过特定转录因子（Gata4、Mef2c和Tbx5）和中药单体淫羊藿苷直接重编程成纤维细胞为GFP+的心肌细胞。通过调节细胞培养过程中不同氧气浓度，检测相应条件下诱导的成纤维细胞表型、功能基因组和表观遗传学改变来评估其对重编程的作用，以期探讨诱导成纤维细胞定向分化为心肌细胞的作用机制。利用EYFP-HIF1α标记的小鼠成纤维细胞系，活体动态观察HIF-1α表达与定向诱导分化过程中成纤维细胞表观遗传学变化，获得HIF-1α表达与成纤维细胞重编程相关性的内在关系及可能的信号转导通路，并研究低氧或低HIF活性对直接重编程的作用机制。

中文关键词： 低氧；低氧诱导因子；成纤维细胞；直接重编程；心肌细胞

英文摘要： The development by the Yamanaka group of a method to reprogram somatic cells into induced pluripotent stem cells (iPS) by over expression of pluripotency factors hold considerable promises for the development of stem cell therapies. Here, we propose to directly induce fibroblast differentiate into myocardial cells through the same or similar mechanisms for fibroblast reprogramming. Our working hypothesis is that fibroblast can be induced and differentiated into myocardial cell through manipulated cellular microenvironment. The resultant fibroblast cells with myocardial cell characteristics could become important myocardial cell source used for myocardial cells transplantation. To test our hypothesis, first, we will establish transgenic mice harburing αMHC promoter driven EGFP-IRES-Puromycin (αMHC-EGFP) as report gene. This biomarker system will eusure only differenatiated cell with myocardial characteristics cells positively expressed the green fluorescent protein (GFP). Secondly, Myocardial specific transcriptional factors, Gata4、Mef 2c and Tbx5, Combining traditional Chinese medicine monomer ICA, will be introducd into fibroblast, we expect directly reprogramming of fibroblasts into functional myocardial cells will occur. To enhance efficiency of reprogramming, we will modify O2 concentration to optimize idea

英文关键词： hypoxia；hypoxia-inducible factor-1；fibroblast；direct reprogramming；cardiomyocytes