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目的探讨5-氮胞苷作为诱导剂体外诱导人骨髓间充质干细胞(human mesenchymal stem cells,hMSCs)分化为心肌样细胞,并对其进行鉴定。方法采用密度梯度离心法与贴壁培养法相结合分离培养hMSCs,以5-氮胞苷诱导第3代hMSCs,24h后继续培养,观察细胞形态学的变化;培养4周,采用免疫细胞化学法对分化的心肌样细胞特异性蛋白结蛋白、心肌肌钙蛋白I进行鉴定。结果培养7d后hMSCs贴壁呈集落生长,有成纤维细胞样外观;5-氮胞苷诱导7d后细胞呈梭形,排列方向渐趋一致,有肌管样结构形成,免疫细胞化学法检测人心肌特异性蛋白结蛋白、心肌肌钙蛋白I表达阳性,而未经诱导的相同培养时间的hMSCs中均未见表达。结论 hMSCs体外在5-氮胞苷诱导下可定向分化为心肌样细胞。
Objective To investigate the differentiation of human mesenchymal stem cells (hMSCs) into cardiomyocyte-like cells induced by 5-azacytidine as an inducer and to identify them. Methods The hMSCs were isolated and cultured by density gradient centrifugation and adherent culture. The 3rd generation hMSCs were induced by 5-azacytidine and cultured 24h later. The morphological changes of the cells were observed. After 4 weeks of culture, hMSCs were cultured with immunocytochemistry Differentiated cardiomyocyte-like protein-specific protein, cardiac troponin I was identified. Results After cultured for 7 days, hMSCs adherent colonies grew in the form of fibroblast-like appearance. After 7 days of induction, the cells were fusiform in shape and arranged in a consistent direction with the formation of myotubes and immunocytochemistry Myocardial specific protein desmin, cardiac troponin I expression was positive, but did not induce hMSCs were not expressed in the same culture time. Conclusion hMSCs can be induced to differentiate into cardiomyocytes in vitro under the induction of 5-azacytidine.