Immunohistochemical measurement of cell proliferation as replicative DNA synthesis in the liver of male Fischer 344 rats following a single exposure to nongenotoxic hepatocarcinogens and noncarcinogens.

We aimed to modify an in vivo/in vitro hepatocyte replicative DNA synthesis (RDS) test using autoradiography of [3H]methylthymidine (3HTdR) into a nonradioactive in vivo version by applying 5-bromo-2'-deoxyuridine (BrdU) immunohistochemistry. The effects of 12 nongenotoxic hepatocarcinogens and 4 noncarcinogens on RDS induction and histological changes in the liver of male Fischer 344 (F344) rats were investigated 24 or 48 hrs after a single oral administration at diverse dose levels. A statistically significant, dose-related elevation in BrdU labeling indices (LI) was obtained for all of the 12 nongenotoxic hepatocarcinogens with only one exception of D,L-ethionine at relatively lower dose levels. No increase was observed in LI for any of the four noncarcinogens examined. From the results of histopathological evaluation, the increase in hepatocyte proliferation by carbon tetrachloride (CCl4), chloroform (CHCl3), and thioacetamide (TAA) was confirmed to be a regenerative liver response following cytotoxicity. Conversely, safrole, tannic acid, and urethane yet hepatotoxicants did not show inflammatory reaction or necrosis under the condition of the present study, and hence their mode of action by which they induce proliferative response was not obvious. The results of this study showed that the in vivo version of RDS test efficiently discriminated nongenotoxic hepatocarcinogens from noncarcinogens, while it did not well clarify the induction mode of cell proliferation.