Human bone marrow-derived mesenchymal stem cells (BM-MSCs) are expected to be a potential source of cells for transplantation. Although recent reports have shown that isolated MSCs can differentiate into hepatocytes, the efficiency of differentiation is insufficient for therapeutic application. To circumvent this problem, it is necessary to understand the mechanisms of hepatic differentiation of human BM-MSCs. Hepatocyte nuclear factor 3 (HNF3), a forkhead/winged helix transcription factor, is essential for liver development. In the present study, we established a tetracycline (Tet)-regulated expression system for HNF3 in UE7T-13 BM-MSCs. HNF3 expression significantly enhanced expression of albumin, ␣-fetoprotein (AFP), tyrosine amino transferase (TAT) and epithelial cell adhesion molecule (EpCAM) genes. The differentiated cells showed hepatocyte-specific functions including glycogen production and urea secretion. During treatment with the Tet-on system for 8 days, over 80% of UE7T-13 cells turned out to express albumin. Furthermore, the combination of Tet with basic fibroblast growth factor (bFGF) efficiently induced the genes such as albumin and TAT, which are associated with maturity of hepatocytes; however, it suppressed genes such as AFP and EpCAM, which are associated with immaturity of hepatocytes, suggesting that Tet-induced HNF3 expression sensitizes BM-MSCs to bFGF signals. Finally, the results of the present study suggest that down-regulation of Wnt/ -catenin signals caused by translocation of -catenin to cytoplasmic membrane is associated with hepatic differentiation of human BM-MSCs. Conclusion: HNF3 expression induced efficient differentiation of UE7T-13 human BM-MSCs. (HEPATOLOGY 2008;48:597-606.)
H uman bone marrow cells can be expanded in vitro and are expected to be a potential source for stem cell therapy without the risk of immune rejection. The bone marrow cells contain mesenchymal stem cells (MSCs) as well as hematopoietic stem cells. 1 The multipotency of MSCs has been shown by differentiating into various cell types, such as mesodermal, neuroectodermal, and endodermal cells. [1][2][3][4][5][6][7][8] Recently, Lee et al. 5 showed that both human bone marrowderived MSCs (BM-MSCs) and umbilical cord bloodderived MSCs have a potency to differentiate into hepatocytes in vitro.
In our previous reports, the protocol to induce hepatic differentiation of UE7T-13 BM-MSCs has been developed. 6 These cells were immortalized by infection with a retrovirus carrying human telomerase reverse transcriptase (hTERT) and one of the early genes of the human papilloma virus, E7. Although hTERT is introduced into UET7T-13 cells, it has been reported that the differentiation potential of the cells is not affected. 4 Indeed, it has been reported that human fetal hepatocytes immortalized by hTERT do not disrupt their differentiation potential. 9