人骨髓来源多能成体祖细胞与人肝细胞系体外间接共培养向肝样细胞分化的实验研究

Differentiation of ZHJ-MAPO into hepatocyte-like cells by indirect co-culture with human hepatocyte line L02

目的探索人骨髓来源多能成体祖细胞ZHJ-MAPCs与人肝细胞系L02在体外间接共培养条件下诱导其横向分化为肝细胞的可行性，为其在组织工程学研究和临床医学中的应用奠定基础。方法将分别接种于盖玻片上的ZHJ-MAPCs和人肝细胞系L02共置于直径10cm培养皿中实现间接共培养。于间接共培养第1、3、5、7天免疫细胞化学鉴定ZHJ-MABCs的ALB、AFP、CK-18、CK-19等肝细胞特征性表型表达变化情况，并设阳性对照（单独培养的L02细胞）和阴性对照（单独培养未经诱导的ZHJ-MAPCs），计数阳性细胞比率。结果AFP在ZHJ-MAPCs间接共培养第1天即表现为强阳性。ALB在共培养第3天出现较强的阳性，第5天达到高峰。CK-18在培养第5天开始出现阳性。CK-19在各时间点均为阴性着色。阳性对照细胞ALB及CK18有较强的阳性表达，AFP弱阳性表达，CK19阴性表达。阴性对照细胞均为阴性表达。结论与人肝细胞系L02间接共培养能够诱导人骨髓来源的ZHJ-MAPCs向成熟肝样细胞横向分化。

Objective To investigate the possibility of differentiation of ZHJ-MAPCs into hepatocytes by indirect co-culture with human hepatocyte line L02 in vitro and evaluate the potential use of ZHJ-MAPCs in tissue engineering either experimentally or clinically. Methods ZHJ-MAPCs and L02 hepatocytes were separated on coverslip （both with cell density of 1 （10^5/ml）, then they were put in a culture dish （10 cm）. The expression of ALB, AFP, CK-18 and CK-19 in ZHJ-MAPCs was determined with immunocytochemistry at different time phases. The separated culture of L02 hepatocytes was employed to serve as the positive control and that of ZHJ-MAPCs as the negative control. Results AFP was strongly positive on day 1 after the co-culture. ALB was positive on day 3 and reached its peak on day 5 after the coculture. CK-18 was positive on day 5. However, CK-19 was negative at different time phases. The positive control cells had high expression of ALB and CK-18 hut weak expression of AFP and negative one of CK-19. The negative control cells had no expression of the 4 markers. Conclusions ZHJ-MAPCs can be induced to differentiate into mature hepatocyte-like cells by co-culture without cell-to-cell contact.