Ex vivo toxicity of nitrogen dioxide in human nasal epithelium at the WHO defined 1-h limit value

Current pollution limits indicating potential harm to human health caused by nitrogen dioxide have prompted a variety of studies on the cytotoxicity and genotoxicity of nitrogen dioxide (NO₂) in vitro. The present study focuses on toxic effects of NO₂ at the WHO defined 1-h limit value of 200 μg NO₂/m(3) air, equivalent to 0.1 ppm NO₂. Nasal epithelial mucosa cells of 10 patients were cultured as an air-liquid interface and exposed to 0.1 ppm NO₂ for 0.5 h, 1 h, 2 h and 3 h and synthetic air as negative control. After exposure, analysis of genotoxicity was performed by the alkaline single cell microgel electrophoresis (comet) assay and by the micronucleus test. Depression of proliferation and cytotoxic effects were checked by the micronucleus assay and the trypan blue exclusion assay. The experiments demonstrated significant DNA fragmentation even at the shortest exposure duration of half an hour in the comet assay. The amount of DNA fragmentation significantly increased with extended NO₂ exposure durations. The amount of DNA fragmentation increased with extended exposure durations to synthetic air at a significantly lower level as compared to NO₂ exposure. Micronucleus inductions were seen only at the longest exposure duration of 3h. There were no changes in proliferation seen in the micronucleus assay under any experimental setup. Moreover, no signs of necrosis, apoptosis or changes in viability were detected. Data demonstrate genotoxicity of NO₂ at concentrations found in the urban atmosphere during short exposure durations. DNA alterations in the micronucleus assay at an exposure time of 3h indicate a significant DNA alteration possibly being hazardous to humans.