Dose and time dependence of the cellular phenotype in rat hepatic preneoplasia and neoplasia induced by single oral exposures to N-nitrosomorpholine.

The dose and time dependence of the cellular phenotype in preneoplastic and neoplastic liver lesions was studied quantitatively in groups of male Sprague-Dawley rats exposed to single oral doses of 0, 200 and 320 mg/kg body wt of N-nitrosomorpholine (NNM) and killed at different time points between 7 and 80 weeks. Compared with the untreated controls, NNM-treated rats showed a dose- and time-dependent increase in the total number and volume of preneoplastic foci of altered hepatocytes (FAH) and in the incidence of hepatocellular adenomas (HCA). In controls, two major phenotypes of FAH were found in low numbers, namely clear and combined clear/acidophilic cell foci which both store excessive amounts of glycogen. The increase in the total number and volume of FAH by single doses of NNM was associated with changes in their cellular phenotype. Clear cell foci were more frequent in the lower dose group than in the higher dose group throughout the observation period. With the exception of the first 15 weeks, combined clear/acidophilic cell foci represented the most frequent phenotype at both dose levels and all time points studied. In contrast, mixed cell foci, which were completely lacking in controls, showed a high relative frequency after a single NNM dose of 320 mg/kg body wt throughout the observation period, but emerged at later time points and in lower numbers when the lower single dose was given. At both dose levels, mixed-cell foci represented the largest phenotype of FAH; their volume fraction correlated positively with the incidence of HCA, indicating a direct precursor-product relationship between these lesions. The size class distribution and temporal appearance of the different phenotypes of FAH and of the adenomas suggest a progression-linked phenotypic instability of the altered cellular phenotypes, resulting in a predominant sequence of cellular changes which leads from glycogenotic clear and acidophilic cell foci to mixed and basophilic cell populations, the latter being poor in glycogen. In addition to these types of FAH, tigroid cell foci, which were very rare in controls, developed in significantly greater numbers after single-dose treatment with NNM at both dose levels. This type of focus may either represent a side lineage of the predominant lineage or an intermediate stage in an alternative lineage of hepatocellular changes, but in any case has the potential to give rise to HCA.