Carbon Nanotubes Induce Oxidative DNA Damage in RAW 264.7 Cells

The induction of DNA and chromosome damage following in vitro exposure to carbon nanotubes (CNT) was assessed on the murine macrophage cell line RAW 264.7 by means of the micronucleus (MN) and the comet assays. Exposures to two CNT preparations (single-walled CNT (SWCNT > 90%) and multiwalled CNT (MWCNT > 90%) were performed in increasing mass concentrations (0.01-100 microg/ml). The frequency of micronuclei was significantly increased in cells treated with SWCNT (at doses above 0.1 microg/ml), whereas MWCNT had the same effect at higher concentrations (1 microg/ml) (P < 0.05). The results of the comet assay revealed that the effects of treatment with SWCNT were detectable at all concentrations tested (1-100 microg/ml); oxidized purines increased significantly, whereas pyrimidines showed a significant increase (P < 0.001) only at the highest concentration (100 microg/ml). In cells treated with MWCNT, an increase in DNA migration due to the oxidative damage to purines was observed at a concentration of 1 and 10 microg/ml, whereas pyrimidines showed a significant increase only at the highest mass concentration tested. However, both SWCNT and MWCNT induced a statistically significant cytotoxic effect at the highest concentrations tested (P < 0.001). These findings suggest that both the MN and comet assays can reliably detect small amount of damaged DNA at both chromosome and nuclear levels in RAW 264.7 cells. Moreover, the modified version of the comet assay allows the specific detection of the induction of oxidative damage to DNA, which may be the underlying mechanism involved in the CNT-associated genotoxicity.