Proliferation of Adult Rat Hepatocytes in Primary Cultures Induced by Platelet-Derived Growth Factor Is Potentiated by Phenylephrine

Abstract

We investigated whether or not proliferation of adult rat hepatocytes induced by plateletderived growth factor (PDGF) is affected by α1adrenoceptor agonists such as phenylephrine during the early and late phases of primary culture. Adult rat hepatocytes underwent significant DNA synthesis after culture with 10 ng/ml of PDGF for 2 hr at a low cell density (3.3 × 104 cells/cm2). Under these culture conditions, the number of nuclei increased significantly during the 3.5-hr culture period. Hepatocyte DNA synthesis and proliferation induced by 10 ng/ml of PDGF decreased slightly as a result of increasing the initial plating density. An α1adrenoceptor agonist, phenylephrine (10-6 and 10-5 M), alone did not affect hepatocyte DNA synthesis and proliferation, but markedly potentiated PDGF-induced hepatocyte DNA synthesis and proliferation. The phenylephrine effect was mimicked by phorbol myristate acetate (10-7 M), but not by ionomycin (10-5 M). The mitogenic effects of PDGF were almost completely blocked by treating hepatocytes with genistein (5 × 10-6 M), U-73122 (3 × 10-6 M), sphingosine (10-5 M), wortmannin (10-7 M) and rapamycin (10 ng/ml). These results demonstrate that PDGF can induce the proliferation of adult rat hepatocytes rapidly in primary culture, regardless of the initial plating density. The present results also suggest that following stimulation with PDGF, activation of tyrosine kinase, phospholipase C, phosphatidylinositol 3-kinase, protein kinase C (PKC) and p70 ribosomal protein S6 kinase is essential for the proliferation of adult rat hepatocytes. The co-mitogenic effects of phenylephrine may involve PKC activation.