Hepatocyte growth factor is a key cytokine for in vitro inducing the differentiation of bone marrow mesenchymal stem cells into liver stem cells. Basic fibroblast growth factor can not only increase the proliferation rate of mesenchymal stem cells and its life, but also can maintain the multilineage differentiation potential of mesenchymal stem cells in the proliferation process.OBJECTIVE:
To investigate the possibility of rat bone marrow mesenchymal stem cells to differentiate into hepatocyte-like cells induced by hepatocyte growth factor and basic fibroblast growth factor in vitro.METHODS:
Bone marrow mesenchymal stem cells were collected from femora of Sprague-Dawley rats. The harvested bone marrow mesenchymal stem cells were separated and purified by whole bone marrow adherent culture method, and passaged in vitro. The bone marrow mesenchymal stem cells were identified by flow cytometry and osteogenic induction. The cells were divided into groups: (1) M0 group: as a negative control, treated without any factor; (2) M1 group: as the positive control group, treated with 20 μg/L hepatocyte growth factor; (3) M2 group: treated with 20 μg/L hepatocyte growth factor+5 μg/L basic fibroblast growth factor; (4) M3 group: treated with 20 μg/L hepatocyte growth factor+10 μg/L basic fibroblast growth factor; (5) M4 group: treated with 20 μg/L hepatocyte growth factor+20 μg/L basic fibroblast growth factor. The morphological changes were observed under inverted microscope. At different stages of differentiation, the album expressions of of mature hepatocyte phenotype marker and alpha fetoprotein of immature hepatocyte phenotype marker were detected by immunohistochemical staining.RESULTS AND CONCLUSION:
The harvested bone marrow mesenchymal stem cells showed morphologic changes of hepatocyte after induction. The album was positively stained at 7 days after induction, its expression level was reduced at 14 days after induction, and then the expression changed into negative at 21 days after induction. The expression of alpha fetoprotein began to appear at 14 days after induction and then continued. No positive staining could be seen in the M0 group, and at the same time point, the positive staining rate in the M3 and M4 group was higher than that in the M2 group (P < 0.05). It indicates that hepatocyte growth factor and basic fibroblast growth factor can induce bone marrow mesenchymal stem cells to differentiate into hepatocyte-like cells, and there is synergistic effect between them.