A human T cell leukemia virus-I infected T cell line, ATL-2, produces an interleukin-2 receptor inducing factor, adult T cell leukemia (ATL)-derived factor (ADF). In the conditioned medium (CM) of ATL-2, we found an inhibitory activity on the epidermal growth factor (EGF)-dependent proliferation of primary cultured rat hepatocytes, measured by cell number and [3H]thymidine incorporation. ATL-2 CM dose-dependently inhibited hepatocyte proliferation. This activity was fractionated by gel filtration at a molecular size of 15,000 to 40,000 and was tentatively called hepatocyte growth inhibitory factor (HGI). Further fractionation with the ion-exchange column indicated that HGI was separable from ADF. Nevertheless, there was a positive correlation between HGI and ADF production, because the HGI activity was also detected in the CM of another ADF producer cell line (HUT102), while no significant HGI activity was detected in the CM of low ADF producer cell lines, ED and MOLT4.