Hepatocytic differentiation of mesenchymal stem cells in cocultures with fetal liver cells.

摘要

AIM: To investigate the hepatocytic differentiation of mesenchymal stem cells (MSCs) in co-cultures with fetal liver cells (FLC) and the possibility to expand differentiated hepatocytic cells. METHODS: MSCs were marked with green fluorescent protein (GFP) by retroviral gene transduction. Clonal marked MSCs were either cultured under liver stimulating conditions using fibronectin-coated culture dishes and medium supplemented with stem cell factor (SCF), hepatocyte growth factor (HGF), epidermal growth factor (EGF), and fibroblast growth factor 4 (FGF-4) alone, or in presence of freshly isolated FLC. Cells in co-cultures were harvested, and GFP+ or GFP- cells were separated using fluorescence activated cell sorting. Reverse transcription-polymerase chain reaction (RT-PCR) for the liver specific markers cytokeratin-18 (CK-18), albumin, and alpha-fetoprotein (AFP) was performed in different cell populations. RESULTS: Under the specified culture conditions, rat MSCs co-cultured with FLC expressed albumin, CK-18, and AFP-RNA over two weeks. At wk 3, MSCs lost hepatocytic gene expression, probably due to overgrowth of the cocultured FLC. FLC also showed a stable liver specific gene expression in the co-cultures and a very high growth potential. CONCLUSION: The rat MSCs from bone marrow can differentiate hepatocytic cells in the presence of FLC in vitro and the presence of MSCs in co-cultures also provides a beneficial environment for expansion and differentiation of FLC.