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We studied the genotoxic potency of motorcycle exhaust particles (MEP) by using a bacterial reversion assay and chromosome aberration and micronucleus tests. In the bacterial reversion assay (Ames test), MEP concentration-dependently increased TA98, TA100, and TA102 revertants in the presence of metabolic-activating enzymes. In the chromosome aberration test, MEP concentration-dependently increased abnormal structural chromosomes in CHO-K1 cells both with and without S9. Pretreatment with antioxidants (alpha-tocopherol, ascorbate, catalase, and NAC) showed varying degrees of inhibitory effect on the MEP-induced mutagenic effect and chromosome structural abnormalities. In the in vivo micronucleus test, MEP dose-dependently induced micronucleus formation in peripheral red blood cells after 24 and 48 h of treatment. The increase of micronucleated reticulocytes induced by MEP was inhibited by pretreatment with alpha-tocopherol and ascorbate. The fluorescence intensity of DCFH-DA-loaded CHO-K1 cells was increased upon the addition of MEP. Our data suggest that MEP can induce genotoxicity through a reactive oxygen species-(ROS-) dependent pathway, which can be augmented by metabolic activation. Alpha-tocopherol, ascorbate, catalase, and NAC can inhibit MEP-induced genotoxicity, indicating that ROS might be involved in this effect.

The effect of alpha-naphthoflavone (alpha-NF) on vascular function was studied in isolated ring segments of the rat thoracic aorta and in primary cultures of human umbilical vein endothelial cells (HUVECs). alpha-NF induced concentration-dependent relaxation of the phenylephrine-precontracted aorta endothelium-dependently and -independently at lower and higher concentrations, respectively. The cGMP, but not cAMP, content was increased significantly in alpha-NF-treated aorta. Pretreatment with N(omega)-nitro- l-arginine methyl ester (L-NAME) or methylene blue attenuated both alpha-NF induced vasorelaxation and the increase of cGMP content significantly. The increase of cGMP content induced by alpha-NF was also inhibited by chelating extracellular Ca(2+) with EGTA. These results suggest that the endothelium-dependent vasorelaxation induced by alpha-NF is mediated most probably through Ca(2+)-dependent activation of NO synthase and guanylyl cyclase. In HUVECs, alpha-NF induced concentration-dependent formation of NO and Ca(2+) influx. alpha-NF-induced NO formation was abolished by removal of extracellular Ca(2+) and by pretreatment with the Ca(2+) channel blockers SKF 96365 and Ni(2+), but not by the L-type Ca(2+) channel blocker verapamil. The Ca(2+) influx, as measured by (45)Ca(2+) uptake, induced by alpha-NF was also inhibited by SKF 96365 and Ni(2+). Our data imply that alpha-NF, at lower concentrations, induces endothelium-dependent vasorelaxation by promoting extracellular Ca(2+) influx in endothelium and the activation of the NO-cGMP pathway.