IN-VITRO AND IN-VIVO RESPONSE OF HEPATOCYTES FROM HEPATIC NODULES TO THE MITOINHIBITORY EFFECTS OF PHENOBARBITAL

One of the proposed mechanisms by which phenobarbital (PB) promotes he patocarcinogenesis in the rat is by differential mitoinhibition. Howev er, our earlier studies indicated that PB inhibited DNA synthesis in v itro in hepatocytes isolated from both surrounding non-nodular liver a nd hepatic nodules promoted by erotic acid (OA). Since nodules generat ed by one promoter need not necessarily be resistant to another promot er, the present study was undertaken to determine whether foci/nodules promoted by PB itself are resistant to the mitoinhibitory effects of PB. Accordingly, rats were initiated with diethylnitrosamine (DENA, 20 0 mg/kg i.p) and promoted with PB (0.07% of PB as its sodium salt) in their drinking water for 16 or 33 weeks, In vitro studies indicated th at PB (3-5 mM) inhibited DNA synthesis induced by epidermal growth fac tor (EGF) in hepatocytes from surrounding non-nodular liver as well as from nodules promoted by PB for 33 weeks. In another experiment, init iated rats exposed to PB for 33 weeks were subjected to either two-thi rds partial hepatectomy (PH) or sham hepatectomy. Hepatocytes were lab elled with tritiated thymidine ill vivo for 48 h. Autoradiographic ana lysis indicated that in the presence of PB, the hepatocytes from both foci/nodules and the surrounding non-nodular liver responded to PH to the same extent. In addition, they both responded to PH less efficient ly as compared to the corresponding controls. Further, initiated rats exposed to PB for 16 weeks when subjected to PH and killed 4 weeks the reafter, the percentage area occupied by gamma-glutamyl-transpeptidase -positive foci/nodules in the PB group increased, but to the same exte nt as in initiated control rats not exposed to PB. The above results r aised an interesting possibility that the lack of resistance of the PB -promoted nodules to the mitoinhibitory effects of PB may be because t he PB-promoted nodule does not express a resistant phenotype. To exami ne this aspect, the response of hepatocytes from 33 week PB-promoted n odules to the mitoinhibitory effects of OA was examined. The results i ndicated that OA (60-120 mu M) inhibited EGF-induced DNA synthesis in hepatocytes isolated from both nodules as well as from surrounding non -nodular liver. These results suggest that PB is a mitoinhibitor but m ay not provide a strong differential growth advantage to foci/nodules in response to a proliferative stimulus. Further, the nodules promoted by PB do not appear to express