miR-21促进脐带血间充质干细胞血管向分化的体外研究

Effect of miR-21 in promoting angiogenesis differentiation of umbilical cord blood mesenchymal stem cells in vitro

目的:体外研究miR-21对脐带血间充质干细胞(umbilical cord blood derived mesenchymal stem cells,UCBMSCs)血管向分化的促进作用。方法:培养UCBMSCs(来源于同济大学干细胞库);构建Lenti-Lac Z-Luciferase和LentimiR-21-Luciferase慢病毒载体;目的基因转染UCBMSCs后0、1、4、7及14d,分别采用实时荧光定量PCR及Western印迹法检测关键成血管因子的表达。用第3代UCBMSCs分别转染Lenti-Lac Z-Luciferase和Lenti-miR-21-Luciferase后,接种于Matrigel上培养,显微镜下观察其管样结构形成情况。采用SPSS13.0软件包对数据进行统计学分析。结果:成功培养脐带血骨髓间充质干细胞并成功转染目的基因;q PCR以及Western印迹法结果表明,miR-21组HIF-1α和VEGF在第4天开始显著高表达,并持续到7 d和14 d;Matrigel结果显示,Lenti-miR-21-Luciferase组管状结构最多,长度最长。结论:Mi R-21具有促进UCBMSCs血管向分化的作用。

PURPOSE： To investigate the effect of miR-21 on promoting vascular differentiation of umbilical cord blood der/ved mesenchymal stem cells （UCBMSCs） in vitro. METHODS： UCBMSCs （from Stem Cell Bank of Tongji University） were cultured; Lenti-LacZ-Luciferase and Lenti-miR-21-Luciferase Lentiviral vector were reconstructed; Real-time fluorescence quantification PCR and Western blot were performed to detect the expression of key angiogenic factors at 0, 1, 4, 7 and 14 d after target gene was transduced into UCBMSCs; UCBMSCs of the third generation were transduced with Lenti-LacZ-Luciferase and Lenti-miR-21- Luciferase; matrigel tube assay was performed to observe the tube-like structure under microscope. The data were statistically analyzed using Student＇s t test and variance analysis with SPSS13.0 software package. RESULTS： UCBMSCs were successfully cultured; The results of qPCR and Western blot showed the expression of HIF-1α and VEGF in miR-21 group significantly increased at 4 d and continued until 7 d and 14 d; Matrigel results showed the tubular structures in Lenti-miR-21-Luciferase group were more than other groups both in amount and in length. CONCLUSIONS： MiR-21 can promote vascular differentiation of UCBMSCs.