title: Mutations associated with in vivo aflatoxin B1-induced carcinogenesis need not be present in the in vitro transformations by this toxin.
abstract: Ingestion of aflatoxin B1 is implicated in the high incidence of human liver cancers in several developing countries. An association has been detected between human exposure to aflatoxins, and mutations in the third base of codon 249 of the p53 gene in hepatomas. In vitro experiments using human cell line cells and aflatoxin B1 have demonstrated the induction of p53 mutations in codon 249 and adjacent codons. It was therefore of interest to see if this correlation between the in vivo and in vitro situations held for other species. The present study examined a rat liver-derived cell line, transformed in vitro with aflatoxin B1, for the presence of mutations associated with in vivo aflatoxin-induced hepatocarcinogenesis. In an in vivo rodent model systems using the aflatoxin B1-sensitive male F344 rat, previous studies have shown that hepatocarcinogenesis is accompanied by significant incidences of codon 12 mutations in K-ras and codon 13 mutations in N-ras genes, but in contrast to the human, apparently not by mutations in codon 243 of the p53 gene (which corresponds to codon 249 in the human gene). In contrast to the situation in humans, mutation in the third base of codon 243 in the rat would not result in any changes in amino acid sequence, but mutations in codon 250, as seen in in vitro human systems, would be expressed in the rat p53 protein. In the present study, an immortalised, non-transformed liver epithelial cell line derived from a male F344 rat was transformed in vitro by aflatoxin B1 as demonstrated by tumour formation in nude mice. The transformation was dependent on metabolic activation of the aflatoxin B1. Transfection of DNA, extracted from these tumours, into NIH 3T3 fibroblasts conferred a stable, malignant transforming capacity. However, no mutations in codon 12 of the K-ras or codon 13 of the N-ras genes were detected in any of these tumours. These results indicate that in vitro transformation does not necessarily involve the same mutations, as those observed in vivo. Also, no mutations in codon 243 or adjacent codons of the p53 gene, paralleling those observed in the human cell line treated with aflatoxin B1, were detected. The results serve to emphasise the in vivo and in vitro variation in the oncogene activation in the same target organ or cell lines derived from that organ, even when using a single carcinogen activated by a known metabolic pathway.
authors.affiliation: Department of Pharmaceutical Sciences, De Montfort University, Leicester, UK.
authors.forename: L A
authors.initials: LA
authors.lastname: Stanley
authors.forename: H G
authors.initials: HG
authors.lastname: Mandel
authors.forename: J
authors.initials: J
authors.lastname: Riley
authors.forename: S
authors.initials: S
authors.lastname: Sinha
authors.forename: F M
authors.initials: FM
authors.lastname: Higginson
authors.forename: D J
authors.initials: DJ
authors.lastname: Judah
authors.forename: G E
authors.initials: GE
authors.lastname: Neal
mesh_terms: D016475:3T3 Cells; D016604:Aflatoxin B1 / Q000633:toxicity*; D000818:Animals; D015139:Blotting, Southern; D015197:Carcinogenicity Tests; D002460:Cell Line; D002471:Cell Transformation, Neoplastic / Q000139:chemically induced / Q000235:genetics*; D004252:DNA Mutational Analysis; D004847:Epithelial Cells / Q000187:drug effects; D016158:Genes, p53 / Q000187:drug effects / Q000235:genetics; D011905:Genes, ras / Q000187:drug effects / Q000235:genetics; D008099:Liver / Q000737:chemistry / Q000187:drug effects*; D008297:Male; D051379:Mice; D008819:Mice, Nude; D009153:Mutagens / Q000633:toxicity*; D009154:Mutation / Q000187:drug effects*; D051381:Rats; D011916:Rats, Inbred F344; D017422:Sequence Analysis, DNA; D014162:Transfection
chemical_list: D009153:Mutagens; D016604:Aflatoxin B1