Prostaglandin E(2) (EP(1)) receptor agonist-induced DNA synthesis and proliferation in primary cultures of adult rat hepatocytes: the involvement of TGF-alpha.

Abstract

We investigated the effects of prostaglandin (EP) receptor subtype agonists on DNA synthesis and proliferation in primary cultures of adult rat hepatocytes to elucidate their mechanisms of action. Maintained in short-term cultures (i.e. 3.5 h) in a serum-free, defined medium, hepatocyte parenchymal cells underwent DNA synthesis and proliferation in the presence of sulprostone (10(-6) M), PGE(2) (10(-6) M), and 17-phenyl-trinor-PGE(2) (10(-9) M) in a time- and dose-dependent manner. PGE(2) was less potent than 17-phenyl-trinor-PGE(2) in stimulating hepatocyte mitogenesis. Sulprostone (10(-6) M) and 11-deoxy-PGE(1) (10(-6) M) showed weak and insignificant stimulation, respectively, for hepatocyte mitogenesis. These effects of PGE(2), 17-phenyl-trinor-PGE(2), and sulprostone were abolished by treatment with a specific EP(1) receptor antagonist, SC-51322, or the PLC inhibitor U-73122. The effects of these EP(1) receptor agonists were potentiated by ionomycin and blocked by verapamil. Hepatocyte mitogenesis was almost completely blocked by specific inhibitors of growth-related signal transducers, such as genistein, wortmannin, PD98059, and rapamycin. A monoclonal antibody against TGF-alpha dose-dependently inhibited PGE(2)- and 17-phenyl-trinor-PGE(2)-induced hepatocyte mitogenesis. Treatment with the EP(1) receptor agonists significantly increased the secretion of TGF-alpha, reaching a maximum within 5 min. The increase in TGF-alpha secretion was blocked by SC-51322, U-73122, somatostatin, and verapamil and potentiated by ionomycin. These results indicate that the proliferative mechanisms of action of EP(1) receptor agonists are mediated through an increase in the autocrine secretion of TGF-alpha, which is dependent on the EP(1) receptor/G-protein involved in PLC regulation/PLC/Ca(2+) system. The locally secreted TGF-alpha, in turn, acts as a complete mitogen that stimulates the tyrosine kinase/MAPK pathway in these cells.