Identification of differentially expressed genes in aflatoxin B1- treated cultured primary rat hepatocytes and Fischer 344 rats

Aflatoxin B1 (AFB1), a mutagen and hepatocarcinogen in rats and humans, isa contaminant of the human food supply, particularly in parts of Africa andAsia. AFB1-induced changes in gene expression may play a part in thedevelopment of the toxic, immunosuppressive and carcinogenic properties ofthis fungal metabolite. An understanding of the-role of AFB1 in modulatinggene regulation should provide insight regarding mechanisms of AFB1-inducedcarcinogenesis. We used three PCR- based subtractive techniques to identifyAFB1-responsive genes in cultured primary rat hepatocyte RNA: differentialdisplay PCR (DD-PCR), representational difference analysis (RDA) andsuppression subtractive hybridization (SSH). Each of the three techniquesidentified AFB1- responsive genes, although no individual cDNA was isolatedby more than one technique. Nine cDNAs isolated using DD-PCR, RDA or SSHwere found to represent eight genes that are differentially expressed as aresult of AFB1 exposure. Genes whose mRNA levels were increased in culturedprimary rat hepatocytes after AFB1 treatment were corticosteroid bindingglobulin (CBG), cytochrome P450 4F1 (CYP4F1), alpha-2 microglobulin,C4b-binding protein (C4BP), serum amyloid A-2 and glutathione S-transferaseYb2 (GST). Transferrin and a small CYP3A-like cDNA had reduced mRNA levelsafter AFB1 exposure. Full-length CYP3A mRNA levels were increased. Whenliver RNA from AFB1-treated male F344 rats was evaluated for transferrin,CBG, GST, CYP3A and CYP4F1 expression, a decrease in transferrin mRNA andan increase in CBG, GST, CYP3A and CYP4F1 mRNA levels was also seen.Analysis of the potential function of these genes in maintaining cellularhomeostasis suggests that their differential expression could contribute tothe toxicity associated with AFB1 exposure.