[Establishment and mechanistic characterization of SV40 T antigen immortalized human fetal hepatocytes]

Human fetal liver cells were cultured in serum free media enriched with hydrocortisone and infected with retrovirus containing SV40 large T gene. Replicating colonies with G-418 resistance developed in 50 days through 23 passages, while none grew in control dishes. After a crisis of 3 months duration, three colonies resumed active proliferation for over 15 months through 60 passages and became immortalized. These cells had epithelioid morphology, and were stained positively for CK-18. Southern blot and immunocytochemistry demonstrated the presence and expression of SV40 T-antigen in the immortalized cell lines. The cells expressed human albumin, especially in those while in cycle. Accumulation of p53 protein in the cell nuclei and strong expression of TGF-alpha as shown by immunocytochemistry explained at least partly the mechanism of unlimited growth of these immortalized human hepatocytes. These cells did not show anchorage-dependent growth in soft agar, nor did tumor form when inoculated into nude mice. These immortalized, differentiated, non-malignant human fetal hepatocyte lines may be useful for further studies.