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目的从脂肪来源的间充质干细胞(ADMSCs)的混合细胞群中分离纯化出CD73阳性的细胞亚群,证明几种不同诱导法诱导CD73细胞的成心肌分化潜能。方法体外分离培养1～3月龄小鼠的ADMSCs,利用流式细胞仪从ADMSCs中分选出CD73+和CD73-两个细胞亚群。分别培养两组细胞,利用5-氮杂胞苷(5-aza)、心肌组织裂解液和5-aza+心肌组织裂解液对两组分选出的细胞分别进行成心肌的诱导。利用免疫细胞化学技术检测3种诱导法的成心肌效果。结果未分选的ADMSCs可分化为成脂和成骨细胞,分选的CD73+细胞具备分化为心肌细胞的良好潜能。3种诱导法均能诱导CD73+ADMSCs向心肌方向分化,5-aza诱导CD73+ADMSCs分化为心肌细胞率为(22.99±6.72)%,心肌组织裂解液组心肌细胞率(14.12±5.42)%,5-aza+心肌组织裂解液组心肌细胞率(26.94±6.11)%。3种方法比较,5-aza+心肌组织裂解液的诱导效果最佳(P<0.05)。结论 CD73+比CD73-的ADMSCs更易于分化为心肌细胞。化学诱导因素和体外模拟心肌微环境,可高效诱导脂肪间充质干细胞分化为心肌细胞。
OBJECTIVE: To isolate and purify CD73-positive cell subpopulations from a mixed population of adipose-derived mesenchymal stem cells (ADMSCs) and to demonstrate that several different induction methods induce the cardiomyogenic potential of CD73 cells. Methods ADMSCs from 1 to 3 months old mice were isolated and cultured in vitro. Two subpopulations of CD73 + and CD73- were sorted out from ADMSCs by flow cytometry. Two groups of cells were cultured respectively, and the cells of both groups were induced into cardiac muscle by using 5-azacytidine (5-aza), myocardial tissue lysis solution and 5-aza + myocardial tissue lysate. The use of immunocytochemistry to detect three kinds of induction of myocardial effect. Results Unsorted ADMSCs differentiated into adipogenic and osteoblastic cells, and sorted CD73 + cells had the potential to differentiate into cardiomyocytes. Three kinds of induction methods could induce the differentiation of CD73 + ADMSCs into cardiac muscle. The percentage of cardiomyocytes differentiated into CD73 + ADMSCs by 5-aza was (22.99 ± 6.72)%, that of cardiomyocytes was (14.12 ± 5.42)%, Cardiomyocytes in 5-aza + myocardial lysate group were (26.94 ± 6.11)%. Among the three methods, 5-aza + myocardial lysate had the best induction effect (P <0.05). Conclusion CD73 + is more easily differentiated into cardiomyocytes than CD73- ADMSCs. Chemically-induced factors and simulated myocardial microenvironment in vitro can efficiently induce adipose-derived mesenchymal stem cells to differentiate into cardiomyocytes.