目的 研究microRNA-1(miRNA-1)能否诱导骨髓间充质干细胞(MSCs)向心肌样细胞分化.方法 构建大鼠miRNA-1表达载体,分离扩增培养及鉴定大鼠MSCs.脂质体法转染大鼠第4代MSCs,实时定量RT-PCR(qRT-PCR)检测转染miRNA-1质粒后MSCs的miRNA-1表达水平.分别于转染2、4和6d后用RT-PCR检测心肌重要转录因子GATA4、NKx2.5和MEF2C的mRNA表达,免疫荧光检测心肌特异蛋白I(cTnI)的表达.结果 90%以上的MSCs表达MSCs重要标志物CD29、CD44;未检测到造血前体细胞标志抗原CD34、白细胞标志抗原CD45的表达.转染miRNA-1质粒后,miRNA-1表达水平明显上调.转染miRNA-1质粒2、4和6 d后,GATA4、NKx2.5和MEF2C的mRNA表达逐渐增强.第4和6天后可见cTnI阳性表达细胞.结论 miRNA-1能诱导大鼠MSCs向心肌样细胞分化.%Objective To investigate may miRNA-1 induce rat bone marrow mesenchymal stem cells (rMSCs) differentiation into cardiomyocyte-like cells. Methods miRNA-1 expression vector was constructed, rMSCs were isolated, amplified and identified. The fourth passage of rMSCs were transfected with Lipofectamine2000 and miRNA-1 expression was then detected by quantitative real-time RT-PCR (qRT-PCR). The mRNA expression of cardiac-specific transcription factor GATA4, NKx2. 5 and MEF2C were detected by RT-PCR,cTnI expression was measured by immunofluorescence,respectively,2,4,6 days. Results Over 90％ of the cultured rMSCs was positive for CD29 and CD44, as two significant markers of MSCs and negative for hematopoetic precursor cell markers CD34, leukocyte antigen CD45. After transfection of miRNA-1 in MSCs, the expression level of miRNA-1 was markedly enhanced. The mRNA expression of GATA4, NKx2. 5 and MEF2C gradually increased from day 2 to day 6. The cTnI expression was detected at day 4,6. Conclusion miRNA-1 can induce MSCs differentiation into cardiomyocyte-like cells.
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