The use of cultured human hepatocytes as a test system to evaluate cell proliferation as a key event in nongenotoxic carcinogenesis

The objective of this study was to evaluate the applicability of replicative DNA synthesis (RDS) in cultured human hepatocytes as an in vitro model to evaluate the carcinogenicity of nongenotoxic chemicals and species differences. Investigations were performed with 39 cryopreserved human hepatocyte preparations from predominantly Caucasian male and female donors aged 10 months to 80 years and were conducted by two separate laboratories, which employed different culture conditions and methodology for evaluating effects on hepatocyte RDS. For all male and female human hepatocyte preparations of all ages examined, treatment with either epidermal growth factor (EGF) and/or hepatocyte growth factor (HGF) resulted in a stimulation of RDS. In contrast, the treatment of human hepatocytes with the constitutive androstane receptor (CAR) activators phenobarbital and CITCO and the peroxisome proliferator-activated receptor alpha (PPARα) activator WY-14,643 did not result in any increases in RDS. These findings are in agreement with previous studies where, unlike EGF and HGF, nongenotoxic CAR and PPARα activators are mitogenic agents in rodent but not in human hepatocytes. While some donor to donor variability was observed, the qualitative inducibility of RDS by EGF and/or HGF in human hepatocytes was not sex-, age-, or, based on a limited number of samples examined, ethnicity-dependent. These studies demonstrate that cultured cryopreserved human hepatocytes are an established, reproducible and relevant in vitro test system for investigating the nongenotoxic carcinogenic potential of chemicals and species differences, and worth progressing to formal validation according to OECD principles.