Using the DNA alkaline unwinding assay to detect DNA damage in laboratory and environmentally exposed cells and tissues

Abstract

The DNA alkaline unwinding assay is being evaluated for use in the detection of DNA damage in marine animals exposed to environmental pollutants. In preliminary work, DNA unwinding methods were used with in vitro cell systems to demonstrate DNA strand breaks. Cultured mammalian fibroblasts and sperm from marine fish and invertebrates ( Pseudopleuronectes, Arbacia and Mytilus) showed concentration-dependent increases in DNA strand breaks after brief exposures to alkylating agents. DNA unwinding methods were also used on DNA extracted from marine animals injected with genotoxicants and from animals exposed in situ at an estuarine site. Gills from blue mussels caged at the New Bedford Harbor Superfund Site (MA, USA) highly contaminated with many organic (e.g. PCBs) and inorganic contaminants, were also examined. A significant increase in DNA strand breaks was seen in gill tissues of animals held in the contaminated site for as little as 3 days. Although not as severe, an increase in strand breaks was also seen in animals held at the control site for 28 days. This apparent damage may have resulted from low-level contamination and/or from generally poor tissue condition. These results suggest that DNA alkaline unwinding results from controlled experiments using cells briefly exposed to toxicants can be used to identify DNA strand breaking agents. While whole-animal exposures to toxicants may affect DNA integrity, unwinding results may not specifically reflect genotoxicity.