Arsenic is recognized to be a nonmutagenic carcinogen because it induces DNA damage only at very high concentrations. However, many more DNA strand breaks could be detected by digesting the DNA of arsenite-treated cells with endonuclease III, formamidopyrimidine-DNA glycosylase, and proteinase K. By doing so, arsenite could be shown to induce DNA damage in human cells within a pathologically meaningful concentration range. Oxidized guanine products were detected in all arsenite-treated human cells examined. DNA-protein cross-links were also detected in arsenite-treated NB4 and HL60 cells. In human umbilical vein endothelial cells, the induction of oxidized guanine products by arsenite was sensitive to inhibitors of nitric oxide (NO) synthase but not to oxidant modulators, whereas the opposite result was obtained in vascular smooth muscle cells. On the other hand, the arsenite-induced oxidized guanine products and DNA-protein cross-links in NB4 and HL60 cells were sensitive to modulators of calcium, NO synthase, oxidant, and myeloperoxidase. Therefore, although oxidized guanine products were detected in all the human cells treated with arsenite, the pathways could be different in different cell types. Because the sensitivity and the mechanism of arsenic intoxication are cell specific, it is important that target tissues and target cells are used for investigations. It is also important that pathologically or pharmacologically meaningful concentrations of arsenic are used. This is because in most cases we are dealing with the chronic effect rather than acute toxicity.
We report here that sequential digestion with endonuclease III, formamidopyrimidine-DNA glycosylase, and proteinase K in Tris buffer markedly increased the sensitivity for detecting DNA damage in arsenic-treated cells. These three enzymes increased DNA strand breaks in an additive manner. By using this sequential-enzyme-digestion comet assay, we demonstrated that trivalent inorganic arsenic induced more DNA damage than monomethylarsonous acid, monomethylarsonic acid, and dimethylarsinic acid in human blood cell lines. However, trivalent inorganic arsenic was far less potent than monomethylarsonous acid in inhibiting pyruvate dehydrogenase activity. Therefore, different mechanisms are involved in inhibiting pyruvate dehydrogenase activity and inducing DNA damage. Our results also indicate while trivalent inorganic arsenic induced more endonuclease III-digestible adducts, monomethylarsonous acid and monomethylarsonic acid induced more proteinase K-digestible adducts. These results suggest there is a difference in the mechanism for inducing DNA damage between inorganic and organic methylated arsenic compounds.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.