Mutations in C57BI/6J and Metallothionein Knock-out Mice Induced by Chronic Exposure of Sodium Arsenate in Drinking Water

摘要

Inorganic arsenic compounds have been classified as Group 1 carcinogens by IARC. Despite its strong human epidemiological evidence of arsenic carcinogenicity, the evidence for its carcinogen-icity in animals is limited. As part of the mechanistic study of arsenic carcinogenesis, we investigated the competitive alkylation of DNA using arsenic alone and in presence of fotemustine, a known potent alkylation agent. Our results support that sodium arsenite [As(Ⅲ)], sodium arsen-ate [As(Ⅴ)], monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA), even at orders of magnitude higher than physiological concentrations, do not alkylate DNA in vitro. In an in-vivo experiment, mice were given up to 20 mg/kg of arsenic in the form of sodium arsenite by the intraperitoneal route, no DNA adducts were detected in the liver of mice 0-48 h after dosing using the ~(32)P post-labelling technique. We exposed virgin C57B1/6J and metallothionein knock-out (MT~(-1)) female mice to drinking water containing 500 μg As/l for over 2 years; high incidences of tumours were developed in a variety of tissues, particularly in lung, intestinal tract and liver. Twenty-six tissues of various organs systems from 12 C57B1/6J and 11 MT~(-1) tumour-bearing mice were selected for detection of mutation in exons 5 and 7 of the p53 gene using PCR, SSCP and cycle sequencing techniques. 9/12 (75.0%) of C57B1/6J and 8/11 (72.7%) of MT~- mice developed one or multiple mutations in exon 5 of the p53 gene. Mutation in codon 163 of exon 5 was observed from 9/12 (75%) and 10/14 (71.4%) of the tissues tested in C57B1/6J and MT~- mice, respectively. Other mutations of lower frequencies were also observed in codons 148, 149 of exon 5 and codon 233 of exon 7. To our knowledge, this is the first point mutation observed resulting from arsenic exposure in an in-vivo animal model. In conclusion, our studies suggest that point mutation in the p53 gene is not likely due to the direct binding reaction of arsenic and DNA because arsenic per se does not appear to alkylate the DNA nor does it form DNA adducts. Mutation patterns in other genes induced by chronic exposure of arsenic require further investigation.