The influence of phenobarbital and butylated hydroxytoluene on the ploidy rate in rat hepatocarcinogenesis.

The effect of the 'promoters' phenobarbital (PB) and butylated hydroxytoluene (BHT) on the ploidy changes during hepatocarcinogenesis in rats was compared in a densitometric analysis of Feulgen-stained nuclei on paraffin-embedded tissue slices. The triphasic Gerlans protocol for liver-cancer induction was applied. Initiation with a single dose of diethylnitrosamine (DEN), and selection with 2-acetylamino-fluorene (2-AAF) combined with a proliferative stimulus (CCl4 administration), was followed by a treatment with PB or BHT for periods up to 22 weeks. Control animals received no treatment after the initiation and selection procedure. Despite intra- and inter-individual variations, an increase in the amount of 2N nuclei is found in the putative preneoplastic lesions of animals that received initiation and selection (I-S) and 3 weeks basal diet (BD). When the diet is supplemented with PB (after I-S), the increase of diploid nuclei starts earlier. At the time carcinoma arise (22 weeks PB treatment) a decrease in the frequency of 2N nuclei is found. BHT-treated animals which develop no carcinoma within the considered timespan, show a clear increased amount of 2N nuclei in the precancerous lesions only after 14 weeks treatment. It seems that there is a positive correlation between the outgrowth of putative preneoplastic foci and nodules in rat liver and an increase of diploid nuclei in these lesions. PB, as promoter used after initiation and selection, speeds up the development of carcinoma in rat liver, and therefore also the shift to diploidization in these rats starts earlier in comparison with I-S-treated rats. Although BHT does not promote liver carcinogenesis, an increase of diploid nuclei is also observed here during lesion formation. It may, therefore, be concluded that the phenomenon of diploidization is closely linked to and probably necessary for preneoplastic development, but that it is not an absolute indicator for neoplastic transformation.