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目的:探讨小分子水凝胶对大鼠骨髓间充质干细胞(MSCs)心肌分化过程中早期分化基因Desmin和α-actin的表达影响。方法:流式细胞仪检测鉴定P9 MSCs细胞表面标记物,分别应用10μmol/L5-aza(5-aza组)、小分子水凝胶+5-aza(水凝胶组)对第9代MSCs进行联合诱导24 h,诱导4周后,QRTPCR和Westem-blot ting法检测Desmin、α-actin mRNA及蛋白的表达。结果:第9代MSCs CD44阳性表达、CD34阴性表达;诱导4周后,5-aza组细胞难以形成球状细胞团,细胞容易脱落,凋亡及死亡细胞较多;水凝胶组细胞死亡少,数量较诱导前增多,细胞之间的分支连接紧密,有聚集生长的趋势,形成球状细胞团块,个别细胞内形成类肌管状结构。5-aza组和水凝胶组心肌早期分化基因Desmin、α-actin均有阳性表达,水凝胶组mRNA及蛋白表达水平均较5-aza组明显增强(P<0.05)。结论:小分子水凝胶可作为细胞的三维培养支架,有利于MSCs生长,具有促进5-aza诱导的MSCs向心肌样细胞分化的作用。
OBJECTIVE: To investigate the effect of small molecule hydrogels on the expression of early differentiation genes Desmin and α-actin in rat bone marrow mesenchymal stem cells (MSCs) during cardiac differentiation. Methods: The surface markers of P9 MSCs were identified by flow cytometry. The MSCs of 9th generation were treated with 10μmol / L 5-aza (5-aza) and small molecule hydrogel + 5-aza After induction for 24 h and induction for 4 weeks, the expression of Desmin and α-actin mRNA and protein were detected by QRTPCR and Westem-blotting. Results: The MSCs of the 9th generation showed positive expression of CD44 and negative expression of CD34. After 4 weeks of induction, the cells in the 5-aza group were difficult to form the globular cell mass, the cells were easy to fall off, and there were many apoptotic and dead cells; The number increased compared with that before induction, and the branches between the cells were closely connected with each other. There was a trend of aggregation and growth, forming globular cell mass, and the formation of muscle-like tubular structures in individual cells. The mRNA expression of Desmin and α-actin in 5-aza group and hydrogel group were significantly higher than those in 5-aza group (P <0.05). CONCLUSION: Small molecule hydrogels can be used as a three-dimensional culture scaffold for cells, which is beneficial to the growth of MSCs and promoting the differentiation of MSCs induced by 5-aza into cardiomyocyte-like cells.