Influence of DT diaphorase on quinone-mediated genotoxicity in human and fish cell lines

Abstract

The influence of the quinone-reducing enzyme, DT diaphorase [NAD(P)H: (quinone acceptor) oxidoreductase], on the genotoxicity of quinones was examined in two cell lines, namely a human hepatoma cell line, HepG2 and a brown bullhead fibroblast cell line, BB. The quinone-reductive characteristics of these two cell lines were examined using an acetylated cytochrome c reduction assay for enzymatic reductase activity. Subsequently, the influence of DT diaphorase on the genotoxicity of two model quinones, menadione (MND) and 9,10-phenanthrenequinone (PQ) was examined in an alkaline unwinding assay for DNA single-strand breaks. Results revealed that DT diaphorase was the predominant quinone reductase in cytosols of both cell lines, and that levels of specific DT diaphorase activity were generally equivalent in the two species. Despite these similarities, results revealed marked qualitative differences between the two species in terms of the influence of DT diaphorase on quinone-mediated genotoxicity. When pretreated with the DT diaphorase inhibitor, dicoumarol, HepG2 cells exhibited a marked exacerbation of genotoxicity in the presence of either MND or PQ, indicating a protective influence of the enzyme. In contrast, quinone genotoxicity in BB cells was not affected by DT diaphorase inhibition, indicating the lack of a protective effect of DT diaphorase. This study illustrates the manner in which functionally analogous enzymes may have markedly distinct influences on xenobiotic toxicity in different cellular systems.