Isolation and characterization of a putative liver progenitor population after treatment of fetal rat hepatocytes with TGF-beta.

The "in vitro" establishment of a physiological model of bipotential liver progenitors would be useful for analyzing the molecular mechanisms involved in regulating growth and differentiation, as well as studying their potential role/s in liver physiology and pathology. The transforming growth factor-beta (TGF-beta) induces de-differentiation of fetal rat hepatocytes (FH), concomitant with changes in morphology. The aim of this work was to isolate and characterize this population of TGF-beta-treated fetal hepatocytes (TbetaT-FH) and test whether they can behave as liver progenitors. The TbetaT-FH isolated cell lines show high expression of Thy-1 and low expression of c-Kit. They express liver-specific proteins, such as albumin and alpha-fetoprotein, and mesenchymal markers, such as vimentin. TbetaT-FH maintain expression of the hnf3beta gene, but lose expression of hnf1beta, hnf4, and hnf6. They express c-met and show an increase in proliferation in response to HGF. Interestingly, the transdifferentiation process is coincident with changes in the expression of genes related to the oxidative metabolism. TbetaT-FH cultured in the presence of EGF + DMSO change morphology, towards epithelial cells, gaining expression of CK19 and c-Kit, markers found in hepatoblasts and bile duct cells. Furthermore, TbetaT-FH form duct-like structures when cultured on Matrigel. TbetaT-FH show also potential to revert to an hepatocyte phenotype when submitted to a long-term "in vitro" differentiation protocol towards hepatocytic lineage. In summary, our results support the hypothesis that hepatocytes can function as facultative liver stem cells and demonstrate that TGF-beta might play an essential role in the transdifferentiation process.