Transforming growth factor-β1 inhibits the growth of primary adult rat hepatocyte cultures by increasing cAMP levels

Abstract

We investigated the mechanisms of transforming growth factor-β1 (TGF-β1) inhibition on transforming growth factor-α (TGF-α)-induced DNA synthesis and proliferation in primary cultures of adult rat hepatocytes. TGF-α (1.0 ng/ml) produced a 4.2-fold elevation of DNA synthesis during 3 h of culture and a 1.2-fold increase in nucleus number (proliferation) during 4 h of culture. TGF-β1 dose dependently inhibited the TGF-α-induced hepatocyte DNA synthesis and proliferation: half-maximal inhibition occurred at a TGF-β1 concentration of 0.08 ng/ml. The inhibitory effects of 1.0 ng/ml TGF-β1 were almost completely reversed by adenylate cyclase inhibitors, 2,4-dideoxyadenosine (10 −6 M), and somatostatin (3×10 −7 M), or by a specific inhibitor of protein kinase A, H-89 ( N-[2-( p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide dihydrochloride; 10 −7 M). In addition, while TGF-α did not affect the basal cellular adenosine 3′,5′-monophosphate (cAMP) levels, TGF-β1 was found to produce dose-dependent increases in cellular cAMP levels. The cAMP-elevating effects of TGF-β1 were also reversed by 2,4-dideoxyadenosine (10 −6 M), and somatostatin (3×10 −7 M), but not by H-89 (10 −7 M). The present results suggest that the specific mechanisms involved in the growth inhibitory effect of TGF-β1 on TGF-α-induced hepatocyte DNA synthesis and proliferation are via stimulation of adenylate cyclase, which increases intracellular cAMP and subsequently activates protein kinase A.