Embryonic stem (ES) cells have a potential to differentiate into various progenitor cells. Here we investigated the differentiation capacity of mouse ES cells into hepatocytes both in vitro and in vivo. During the culture of embryoid bodies (EBs) derived from ES cells,
albumin (ALB)
messenger RNA (
mRNA) was expressed within 12 days after removal of
leukemia inhibitory factor, and
alpha-fetoprotein (AFP)
mRNA was observed within 9 days without additional exogenous
growth factors. In ES cells and early EBs, by contrast, neither ALB
mRNA nor AFP
mRNA was observed. ALB
protein was first detected at day 15 and the level increased with the culture period. The differentiation of EBs facilitated the synthesis of
urea with the culture period, whereas early EBs and ES cells produced no
urea. These results suggest that cultured EBs contain hepatocytes capable of producing ALB and
urea. ES cells and the isolated cells from EBs were transplanted through portal vein to the liver after 30% partial
hepatectomy of female mice pretreated with
2-acetylaminofluorene. Four weeks after
transplantation with isolated cells from day-9 EBs, ES-derived cells containing Y-chromosome in the liver were positive for ALB (0.2% of total liver cells), whereas
teratoma was found in mice transplanted with ES cells or EBs up to day 6. The incidence of
teratoma was decreased with the culture duration and no
teratoma was observed in the liver transplanted with isolated cells from day-9 EBs. In conclusion, our in vitro and in vivo experiments revealed that cultured EBs contain functional hepatocytes or hepatocyte-like cells.