Effects of ozone on DNA single-strand breaks and 8-oxoguanine formation in A549 cells

Animal studies have demonstrated that ozone exposure can induce lung tumors. Recent epidemiological studies have also shown that increased ozone exposure is associated with a greater risk of lung cancer. This study used single-cell gel electrophoresis (the Comet assay) and flow cytometry to investigate DNA damage in A549 cells exposed to ozone levels below the current ambient standard. Cells were exposed to ozone at levels of 0, 60, 80, and 120 ppb, and then DNA single-strand breaks and 8-oxoguanine levels were measured. Additionally, the formamidopyrimidine glycosylase (Fpg) repair enzyme was added to the Comet assay to enhance detection of oxidative damage. Vitamins C and E were also added to determine their inhibitory effects on ozone-induced 8-oxoguanine. Measurements of tail length, tail intensity, and tail moment of the Comet assay were shown to correlate with each other. However, tail moment appeared to be more sensitive than the other two indicators in detecting DNA single-strand breaks. Tail moments of cells exposed to 80 and 120 ppb of ozone were significantly higher than those exposed to 0 ppb (P<0.05). These three indicators of DNA single-strand breaks with Fpg were shown to be increased and more sensitive than those without Fpg. After Fpg was introduced, the tail moments at ozone levels of 60, 80, and 120 ppb were significantly higher than those at 0 ppb (P<0.05). Furthermore, 8-oxoguanine levels, determined by fluorescence intensity, at 80 and 120 ppb of ozone exposure were significantly higher than the level at 0 ppb. Pretreatment with vitamins C and E reduced the 8-oxoguanine levels caused by ozone. We conclude that ozone levels below current ambient standards may induce DNA breaks and oxidative DNA damage. Moreover, the Fpg repair enzyme in the Comet assay can increase the sensitivity of oxidative damage detection in vitro.