Putrescine as a comitogen of epidermal growth factor in rat liver growth

Putrescine can stimulate regeneration of the remnant liver after partial hepatectomy in rats when exogenously administered, but its mitogenic action has not been shown in cultured hepatocytes. To find its action site(s) in the regulation of hepatocyte proliferation, we examined its effect on hepatocyte DNA synthesis in relation to mitogenic action of epidermal growth factor in vitro and in vivo. When putrescine was added to the medium of adult rat hepatocytes in primary culture 36 hr after plating, DNA synthesis at 50 hr induced by addition of epidermal growth factor at 24 hr was significantly enhanced. This enhancement disappeared by removal of epidermal growth factor at the time of putrescine addition. Putrescine added to the medium was taken up in a dose-related manner by hepatocytes, irrespective of the presence of epidermal growth factor, whereas 125I-epidermal growth factor binding to hepatocytes was not affected by addition of putrescine. When rats received epidermal growth factor at 2-hr intervals until 10 hr, 5-bromo-2'-deoxyuridine labeled and mitotic hepatocytes were increased in number at 48 hr with increased hepatic DNA content. These increases were not affected by concomitant administration of putrescine until 10 hr, but significantly enhanced by additional administration of putrescine and epidermal growth factor from 20 to 30 hr. We conclude that putrescine may stimulate proliferation of hepatocytes that have entered the G1-phase of the cell cycle as a comitogen of epidermal growth factor, probably through action at the molecular levels to enhance its mitogenic activity.