Liver cells from six donors have been cultured; all underwent 3 successive (1:4 split ratio) subculturings, with a cell doubling time of 3 days, before division ceased. The replicating cells were uniformly positive for keratin 18 but negative for keratin 19, a keratin species not found in hepatocytes but present in ductal cells. In addition, 50% exhibited human albumin. Cells from the second, third and fourth donor received the SV40 T antigen gene in order to extend their population doubling potential. One of these, denoted as THLE-1, continued to replicate 40 population doublings before growth ceased. The other two, THLE-2 and THLE-3 respectively, appear to have indefinite life spans. When THLE-1 cells were examined at the 3rd passage, they were uniformly positive and negative for keratins 18 and 19 respectively. However, even though 100% of the cells exhibited keratin 18 at the 10th passage, 30% of the cells now also displayed keratin 19. Transferrin, fibrinogen and albumin expression was measured in 10th passage cells. None of the cells showed evidence of transferrin or fibrinogen. On the other hand, 20% of the cells were positive for albumin. These albumin expressing cells were always found in clusters of 8-12 cells. In addition, the number of albumin positive cells increased to 30-40% if the THLE-1 cells were incubated for 48 hrs in medium supplemented with 10% serum. Comparable characterizations of THLE-2 and THLE-3 cells are in progress.
