Generation of hepatocytes from cultured mouse embryonic stem cells

Abstract

Embryonic stem (ES) cells are pluripotent cells derived from the inner cell mass of fertilized blastocysts in vitro. ES cells can be induced to undergo differentiation into potentially all cell types. The aim of this study is to examine the differentiating potential of mouse ES cells into hepatocytes in the presence of retinoic acid (RA), hepatocyte growth factor (HGF), and β-nerve growth factor (β-NGF). RA, HGF, and β-NGF were added to the cell culture. Hepatocyte induction was confirmed morphologically, as well as biochemically, through immunohistochemical assays of α1-antitrypsin (α1-AT) and alfafetaprotein (AFP) expression and reverse-transcriptase polymerase chain reaction tests for the presence of albumin, transthyretin, glucose 6 phosphates, hepatic nuclear factor 4, and SAPK/ERK kinase-1 (SEK1) messenger RNA, produced only by functioning hepatocytes. Fifteen days after the addition of HGF and β-NGF to the cell culture, many epithelioid cells were noticed. α1-AT, AFP, albumin, transthyretin, glucose 6 phosphates, hepatic nuclear factor 4, and SEK1 messenger RNA expression also was detected, indicating successful ES cell differentiation into functioning hepatocytes. However, in the presence of RA alone, only transthyretin messenger RNA was positive, whereas no other expression pertaining to functioning hepatocytes could be detected. In the presence of HGF and β-NGF, mouse ES cells can differentiate into functioning hepatocytes, whereas RA function is limited. (Liver Transpl 2003;9:1094-1099.)