Figure 3

Antioxidant response and DNA damage following VOC exposure. (A) Measurement of reduced glutathione levels. Primary keratinocytes were exposed to VOCs (toluene, hexane, acetaldehyde, formaldehyde, acetone, 20 ppmV, each) for 4 hours. 24 hours after exposure they were incubated with ThiolTrackerTM Violet dye for 30 minutes and GSH level was assayed using a microplate fluorimetric reader. GSH level is presented as a percentage of non-treated cells. Data, mean ± SEM from three independent cultures, **P < 0.01 (B) GPx2 induction following VOC exposure. Primary keratinocytes were exposed to VOCs (toluene, hexane, acetaldehyde, formaldehyde, acetone, 20 ppmV, each) for 4 hours. 24 hours later GPx2 expression was detected by immunoblotting against GPx2 protein using cell lysates. The data shown is representative of three separate cultures. The blot was cropped and full-length blot is included in Supplementary file information. (C) For the same VOC exposure DNA damage was evaluated by flow cytometry using cells stained with Anti-phospho Histone H2A.X (Ser139) Alexa Fluor 488 antibody. Results are presented as the mean fluorescent intensity of the cells. Data, mean ± SEM from three independent cultures, **P < 0.01. For the same VOC exposure DNA damage was evaluated by western blot analysis using polyclonal antibody against Phospho-Histone H2AX. The blot was cropped and full-length blot is included in Supplementary file information.