骨髓间充质干细胞转化为肝样细胞的体外实验研究

目的评价骨髓间充质干细胞（BMSCs）向肝样细胞诱导的可行性。方法2008年1月-2009年1月,以肝细胞生长因子（HGF）20ng/mL,成纤维细胞生长因子4（FGF-4）10ng/mL为诱导剂,从细胞形态变化,并通过RT-PCR、免疫组化方法分别对诱导第7、14、21及28天的细胞进行白蛋白（ALB）、甲胎蛋白（AFP）、细胞角蛋白18（CK18）等检测。人L-02肝细胞及未诱导的BMSCs分别为阳性和阴性对照。结果BMSCs诱导7d出现类圆形或多角形细胞,并出现铺路石样结构;诱导14d细胞呈现典型的铺路石状;诱导21d,同前;诱导28d,细胞排列紊乱,局部细胞的形态不规则、细胞边界不清。BMSCs诱导第7、14、21天ALB、CK18、AFP等mRNA表达阳性;未诱导BMSCs均为阴性;肝细胞ALB、CK18、AFP等mRNA表达均阳性。免疫细胞化学检测结果同RT-PCR。结论以HGF及FGF-4为主的诱导体系可有效诱导BMSCs向肝样细胞转化,BMSCs可以作为一种新的肝细胞来源。

Study on Bone Marrow Mesenchymal Stem Cells Differentiated into Hepatoid Cells in vitro

Objective To investigate the differentiation of bone marrow mesenchymal stem cells（BMSCs）into hepatocytes by induction of fibroblast growth factor-4（FGF-4）and hepatocyte growth factor（HGF）.Methods From January 2008 to January 2009,BMSCs were isolated by gradient density centrifugation and divided into two groups,one was cultured by Dulbecco＇s minimum essential medium（DMEM）,another was cultured by DMEM supplemented with FGF-4（10 ng/mL）and HGF（20 ng/mL）.Expression of cytokeratin-18（CK18）,fetoprotein（AFP）and albumin（ALB）were detected by immunohistochemistry and RT-PCR respectively.Un-induced BMSCs and L-02 hepatocytes were chosen as negative and positive controls respectively.Results BMSCs remained unchanged morphology during 7 days cultured with FGF-4 and HGF,and morphological changes into round and epithelial could be seen on d14.The protein and mRNAs of ALB,CK18 and AFP could be detected on days 7,14,21 just as L-02.No expression of ALB,CK18 and AFP were detected in the un-induced BMSCs.Conclusion BMSCs can differentiate into hepatocyte by induction of FGF-4 and HGF.BMSCs are a new source of cell types for cell transplantation therapy of hepatic disease.