Inhibitors of ADP-ribosyl transferase suppress the mitogenic actions exerted by tumour promoters, but not those evoked by peptide mitogens, in primary neonatal rat hepatocytes

A significant stimulation of the 24-h (between day 4 and 5 in vitro) new DNA synthetic activity was elicited in primary neonatal rat hepatocytes kept in low-calcium (0.01 mmol/l) HiWoBa2000 synthetic medium by the addition of a single dose (10(-10) mol/l) of each of several tumour promoters [i.e. 12-O-tetradecanoylphorbol-13-acetate (TPA), phenobarbital, nafenopin, saccharin, teleocidin, benzoyl peroxide butylhydroxytoluene (BHT), dichlorodiphenyltrichloroethane (DDT), lindane, clofibrate and melittin]. Even hormones [e.g. epidermal growth factor (EGF), glucagon and insulin at 10(-10) mol/l] and EGF-like acting drugs (i.e. imidazole and indomethacin, at 10(-11) mol/l) similarly enhanced with respect to untreated controls the 24-h flow into S phase of the primary hepatocytes on condition, however, that the cells were incubated in a high- (i.e. 1.8 mmol/l) and not a low-calcium HiWoBa2000 medium. Xenobiotics, peptide mitogens and EGF-like acting drugs also enhanced the in vitro hepatocellular mitotic activity. The growth-stimulatory effects of the aforementioned eleven tumour promoters were entirely suppressed by the simultaneous addition to the growth medium of a fully effective dose (10(-4) - 10(-3) mol/l) of agents, such as 3-aminobenzamide (3-ABA), 3-methoxybenzamide (3-MBA) or nicotinamide (NA), that are known to inhibit the activity of ADP-ribosyl transferase (ADPRT). However, under the same conditions these inhibitors hampered neither the basal DNA synthetic and mitotic activities of spontaneously cycling hepatocytes nor the stimulation of the hepatocellular growth processes evoked by peptide mitogens and EGF-like acting drugs. Quantitative autoradiographic investigations showed that the incorporation of the ADP-ribose precursor and ADPRT substrate [3H]NAD into nuclear macromolecules of gently digitonin-permeabilized hepatocytes was negligible in the untreated cultures, whereas it was strikingly and nearly steadily increased by a 2-, 8- and 24-h exposure to a fully mitogenic dose (10(-10) mol/l) of TPA, thereby revealing that an early, significant and roughly steady activation of the nuclear ADPRT had taken place in the phorbol ester-treated liver parenchymal cells. The simultaneous addition of 3-ABA (10(-4) mol/l) not only fully checked the mitogenic effects of TPA, but even suppressed about two-thirds of the TPA-elicited nuclear incorporation of [3H]NAD by the permeabilized hepatocytes, thus showing that a significant curtailment of the TPA-activated ADPRT did occur is association with the abatement of the mitogenic effects of TPA by this inhibitor.(ABSTRACT TRUNCATED AT 400 WORDS)