中文摘要：

研究小鼠胚胎成纤维细胞与新生小鼠心肌细胞共培养、并过表达心肌特异性转录因子对小鼠胚胎成纤维细胞向心肌细胞转分化效率的影响.首先取出生48h的新生小鼠的心肌细胞并进行培养,模拟新生状态下的心脏内环境.在这一基础上,将小鼠胚胎成纤维细胞与新生小鼠心肌细胞共培养,并使用相关转录因子过表达的策略,使用质粒载体在小鼠胚胎成纤维细胞中过表达GATA4,MEF2C,TBX5,HAND,NKX2.5.15d后通过鉴定相关mRNA表达量,免疫荧光观察特定蛋白分布以及流式细胞仪分析,来评估转分化的效率.结果表明,在模拟新生心脏内环境以及过表达心肌相关转录因子的条件下,小鼠胚胎成纤维细胞向心肌细胞转分化的效率得到提高,但是经由转分化得到的心肌细胞的成熟程度依然不足.

英文摘要：

To evaluate the effect of natal cardiac microenvironment in cardiomyocytes transdifferentiation,extraction of natal murine cardiomyocytes was performed 48 hours after birth.The mouse embryonic fibroblast（MEF）were cocultured with murine cardiomyocytes in transwell culturing system and then transfected with a plasmid vector to overexpress 5key cardiac transcription factors,including GATA4,MEF2 C,TBX5,HAND and NKX2.5.On Day 15 after transdifferentiation,real-time qPCR,immunofluorescence and flow cytometry were used to evaluate the efficiency of transdifferentiation.Real-time qPCR showed that co-cultured with murine cardiomyocytes,the expression of cardiac specific gene cTnT was upregulated,and the non-cardiac but a muscle specific gene Myh11 was downregulated.Immunofluorescence demonstrated the distribution of cardiac specific protein cTnT andα-MHC,but no sarcomere detected.Flow cytometry indicated that co-cultured with murine cardiomyocytes,the cTnT＋cell rate increasedmuch.However,with the extension of culture time,the maturity of these cells was inhibited and showed the backward development.Therefore,microenvironment of natal cardiomyocytes could significantly improve the efficiency and specificity of transdifferentiation.