南方医科大学学报

南方医科大学学报 ›› 2006, Vol. 26 ›› Issue (10): 1439-1442.

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人骨髓间充质干细胞体外分化为肝样细胞的实验研究

谢金敏； 陈建锋； 高毅； 姚坤厚；   

  1. 南方医科大学珠江医院普外科； 南方医科大学珠江医院普外科 广东广州510282； 广东广州510282；
  • 出版日期:2006-10-20 发布日期:2006-10-20
  • 基金资助:
    广东省科技计划项目(2002A3020206)~~

Hepatocyte growth factor and fibroblast growth factor-4-induced differentiation of human bone marrow mesenchymal stem cells into hepatocyte-like cells in vitro

XIE Jin-min, CHEN Jian-feng, GAO Yi, YAO Kun-hou Department of General Surgery, Zhujiang Hospital, Southern Medical University, Guangzhou 510282, China   

  1. 南方医科大学珠江医院普外科； 南方医科大学珠江医院普外科 广东广州510282； 广东广州510282；
  • Online:2006-10-20 Published:2006-10-20

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摘要： 目的探索肝细胞生长因子(HGF)联合成纤维细胞生长因子-4(FGF-4)诱导人骨髓间充质干细胞(HMSCs)向肝细胞方向分化的能力,为生物人工肝及肝细胞移植等找到新的种子细胞来源。方法分别用HGF、FGF-4、HGF+FGF-4诱导HMSCs分化为肝样细胞;以未加诱导因素的HMSCs、L-02人肝细胞株为阴、阳性对照组。相差显微镜观察细胞形态的变化;在不同诱导分化阶段免疫细胞化学染色检测AFP和CK18,并用图像分析法计算HMSCs的分化比率;免疫荧光细胞化学染色法检测AFP、ALB的表达;RT-PCR检测AFP和ALBmRNA表达的情况。结果HMSCs经诱导向肝样细胞转化,变成纺锤形、不规则圆形或多角形细胞。免疫细胞化学染色各诱导组均可检测出AFP和CK18表达,图像分析表明HGF+FGF-4联合诱导分化的AFP、CK18阳性率最高,HGF次之,FGF-4则较弱;免疫荧光细胞化学染色发现各诱导组AFP、ALB均为阳性表达,而阴性对照组则为阴性表达;各诱导组RT-PCR均可检测出AFP和ALBmRNA的表达,而阴性对照组则没有表达。结论HGF、FGF-4、HGF+FGF-4均能诱导HMSCs分化为肝样细胞,分化的肝样细胞能分泌肝细胞特异性产物AFP、ALB、CK18等。以HGF+FGF-4诱导HMSCs分化为肝样细胞的阳性率最高,HGF次之,FGF-4则较弱。 

Abstract: Objective To induce the differentiation of human bone marrow mesenchymal stem cells (HMSCs) into hepatocyte-like cells with hepatocyte growth factor (HGF) and fibroblast growth factor-4 (FGF-4) in vitro. Methods HMSCs were induced to differentiate into hepatocyte-like cells by HGF (group B), FGF-4 (group C) and HGF+FGF-4 (group D) in vitro. Undifferentiated HMSCs and L-02 cells were used as the negative (group A) and positive (group E) controls, respectively. The changes of cell morphology were observed microscopically. The expressions of hepatic markers, alpha fetoprotein (AFP) and CK-18, were detected by immunocytochemical staining at different times after induction, and the differentiation ratios of the various groups of HMSCs were calculated on the basis of image analysis. The expressions of AFP and ALB were detected by immunofluorescence assay in each group at different times after induction, and the expressions of AFP and ALB mRNA by RT-PCR. Results HMSCs gradually transformed into spindle-shaped, round, polygonal or irregular cells after induction. Immunocytochemical staining revealed positive AFP and CK18 expressions in groups B, C, and D after induction as well as in group E. The positive units (PU) of AFP and CK18 in group D calculated according to image analysis were significantly higher than that of groups A, B, and C. The expressions of AFP and ALB detected by immunofluorescence were both positive after induction in all groups except group A, similar to the findings of the expressions of AFP and ALB mRNA by RT-PCR. Conclusion HMSCs can be induced to differentiate into hepatocyte-like cells by HGF, FGF-4 and their combination at certain concentrations, and the hepatocyte-like cells can express some hepatic markers such as AFP, ALB, CK18, etc. HGF+FGF-4 may achieve more effective induction of HMSC differentiation into hepatocyte-like cells, and the efficiency of HGF is greater than that of FGF-4. 

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