Liver regeneration and alpha-fetoprotein messenger RNA expression in the retrorsine model for hepatocyte transplantation

Recently, we described a new model for hepatocyte transplantation with near ly total replacement of the liver by exogenous hepatocytes (E, Laconi et at , Am. J. Pathol., 153: 319-329, 1998). The model is based on the mitoinhibi tory effect of the pyrrolizidine alkaloid retrorsine on hepatocytes in the resident liver while transplanted hepatocytes proliferate. In this study, w e exploit this novel approach to address the important and controversial is sue of whether hepatocytes, when proliferating extensively, undergo dediffe rentiation and give rise to foci of undifferentiated hepatocytes. Genetical ly marked hepatocytes (isolated from normal Dipeptidyl peptidase IV+ Fische r 344 rats) were delivered intraportally (2 x 10(6) cells) into the liver o f retrorsine-treated Dipeptidyl peptidase IV- mutant Fischer 344 rats in co njunction with partial hepatectomy. Transplanted hepatocytes were detected histochemically or immunohistochemically, and cell proliferation was studie d by in situ hybridization for histone-3 mRNA. Expression of alpha-fetoprot ein (AFP) mRNA, a marker of hepatocyte dedifferentiation, was also revealed by ill situ hybridization. One day after partial hepatectomy and hepatocyt e transplantation, endogenous hepatocytes and oval cells expanding in the l iver expressed histone-3 mRNA (cells had entered S phase); 2 days later, tr ansplanted hepatocytes and nonparenchymal cells also expressed histone-3 mR NA. Although the majority of endogenous hepatocytes did not divide and beca me arrested as quiescent megalocytes, the exogenous hepatocytes, as well as newly formed small hepatocytes, most probably derived from liver progenito r cells, underwent extensive proliferation. After 7-14 days, the nonparench ymal cells stopped proliferating, but transplanted hepatocytes and small en dogenous hepatocytes continued to proliferate for 1 month, forming foci of dividing parenchymal cells. Although many of the hepatocytes in clusters we re in S phase (histone-3 mRNA positive), none expressed AFP mRNA. In contra st, high expression of AFP mRNA was observed in proliferating oval and tran sitional cells, forming duct-like structures of cytokeratin-19-positive cel ls. From these studies, we conclude that hepatocyte proliferation in the ad ult liver is not associated with dedifferentiation.