Relationship of histochemically detectable altered hepatocyte foci to hepatic tumorigenesis

A new experimental system was used to examine the stages of chemically induced hepatic neoplasia in the rat. The treatment protocol involved the intraperitoneal injection of a single non-necrogenic dose of carcinogen [N-nitrosodiethylamine (NDEA) or benzo[a]pyrene (BP)] into male and female rats within one day after birth, followed by dietary exposure to promoter (0.05% phenobarbital) from weaning. Rats were killed at intervals, and their livers were examined for tumours and for histochemically detectable foci of altered hepatocytes. Six histochemical markers were used. Through serial frozen-sectioning techniques and computer-assisted image analysis, foci containing between one and six markers were identified, and their average sizes were calculated. The same complement of histochemical tests was applied to the primary hepatic tumours observed in this study. The data showed that (1) the new treatment protocol was highly efficient in foci and tumour production; (2) growth rates and incidence levels of foci were directly related to hepatocarcinogenic effectiveness (NDEA greater than BP), whereas both carcinogens had similar effects on foci phenotypic properties; (3) after their formation, foci at a given level of phenotypic complexity did not become progressively more complex; (4) incidence levels of foci were sex-dependent (females greater than males), but growth rates of foci were the same for both sexes; (5) growth rates and growth capacities (ranges of possible growth rates) of foci were directly related to phenotypic complexity levels of foci; (6) frequencies and phenotypic complexities of foci were inversely related; the reverse was true for tumours, although 10% of the tumours were relatively simple (three markers or fewer); (7) marker frequency distribution patterns were completely different in foci and in tumours. From these observations, we suggest that the foci are not direct tumour progenitors but, instead, are manifestations of a mosaic of subtumorigenic effects of the carcinogenic stimulus on cellular functions associated with the control of cell division and phenotypic character. The observed foci and tumour characteristics suggest that these carcinogen-induced cellular changes define elements of the mechanism whereby a specific neoplastic transformation site is rendered more accessible to carcinogenic attack.