Biochemical analysis of the initial carcinogenic changes that induce preneoplastic and neoplastic cell populations during chemical hepatocarcinogenesis in rats

To analyze the initial carcinogenic changes that induce preneoplastic and neoplastic cell populations in the rat liver, a short-term in vivo promotion assay method was developed. Preneoplastic foci and nodules were quantitated with glutathione S-transferase P-form (GST-P) and γ-glutamyl transpeptidase. Among the four agents tested, benzyl isothiocyanate (BITC) demonstrated the strongest promotor activity, producing very large nodules composed of 2¹⁸ to 2²⁰ cells in the rat liver. In addition, a choline/methionine-deficient (CMD) diet, which strongly inhibits protein synthesis, exhibited lower but distinct promotive activity, giving rise to large nodules composed of 2¹¹ to 2¹³ cells. Based on the collected stereologic and biochemical data as well as the results of DNA microarray analysis, preneoplastic foci and nodules were strongly indicated to grow without cell division. The absence of cell division indicates the absence of mutations in the genetic mechanism, and vice versa; thus, preneoplastic cell induction can be considered nongenetic. Furthermore, the nodules were markedly more susceptible to promoter agents than hepatocytes as to die of necrosis. Based on these experimental findings, neoplastic cell induction was logically deduced to be nongenetic. The present analysis may help improve the knowledge of the "unknowable mechanism of cancer initiation" of rat chemical hepatocarcinogenesis.