DNA oxidation: investigating its key role in environmental mutagenesis with the comet assay

Abstract

DNA oxidation, which can have potentially serious mutagenic consequences, commonly accompanies exposure to environmental mutagens. Oxidised bases can be measured chromatographically, but spurious oxidation during sample preparation leads to serious over-estimation of low levels of damage. A more reliable approach is to employ endonucleases specific for oxidised bases, to introduce breaks in cellular DNA that are then most commonly measured using the comet assay (alkaline single cell gel electrophoresis). The two enzymes in general use are formamidopyrimidine DNA glycosylase, which detects primarily 8-oxo-7,8-dihydroguanine (8-oxoGua), and endonuclease III which recognises oxidised pyrimidines. We give a brief account of the recommended experimental procedures, and then describe applications in various areas of environmental research. Cultured cell lines or white blood cells have been exposed to a range of environmental mutagens, including natural products, industrial chemicals, radiation and nanoparticles. In vivo exposure of animals and humans to pollutants is more challenging but can give particularly valuable information in relation to real life exposure. Possibly the most useful application is in biomonitoring of human population groups suffering actual exposure to environmental or occupational mutagens. Finally, the potential use of this technique to monitor effects of contaminants in the natural environment has yet to be fully exploited.