Tumor necrosis factor (TNF) receptor-2-mediated DNA synthesis and proliferation in primary cultures of adult rat hepatocytes: The involvement of endogenous transforming growth factor-α

We investigated the effects of tumor necrosis factor (TNF)-α on DNA synthesis and proliferation, and its signal transduction pathways in primary cultures of adult rat hepatocytes. TNF-α induced time- and dose-dependent increases in hepatocyte DNA synthesis and proliferation. The hepatocyte proliferation stimulated by 30 ng/ml TNF-α was significantly inhibited by anti-TNF receptor 2 antibody, but not by anti-TNF receptor 1 antibody. TNF-α-induced hepatocyte DNA synthesis and proliferation were blocked by AG1478 (10− 7 M), PD98059 (10− 6 M), LY 294002 (10− 7 M), and rapamycin (100 ng/ml). TNF-α at 30 ng/ml significantly increased phosphorylation of receptor tyrosine kinase (175 kDa) and p42 mitogen-activated protein (MAP) kinase. This data suggests that the proliferative signal for primary cultured hepatocytes induced by TNF-α is mediated by TNF receptor 2 and the receptor tyrosine kinase/MAP kinase pathway. In addition, TNF-α-induced hepatocyte mitogenesis was significantly blocked by somatostatin (10− 6 M), adenylate cyclase inhibitor dideoxyadenosine (10− 7 M), protein kinase A inhibitor H-89 (10− 7 M), and neutralizing antibody to transforming growth factor (TGF)-α in culture. Indeed, 30 ng/ml TNF-α was found to rapidly stimulate secretion of TGF-α, and this secretion was also blocked by anti-TNF receptor 2 antibody. Moreover, TGF-α secretion induced by TNF-α was suppressed by dideoxyadenosine, H-89, and somatostatin. Together, these results indicate that stimulation of TNF receptor 2 by 30 ng/ml TNF-α induces autocrine secretion of TGF-α via the adenylate cyclase/protein kinase A pathway, after which TGF-α induces hepatocyte DNA synthesis and proliferation through the TGF-α receptor-linked tyrosine kinase (175 kDa)/MAP kinase signaling system.