Identification of pathway-based toxicity in the BALB/c 3T3 cell model

•Dose-related toxicity was evidenced in BALB/c 3T3 cells treated with PM2.5 samples.•Cell toxicity depended on PM2.5 sampling season and sampling site.•Gene pathways related to toxic stress response were identified in the treated cells.•No foci formation was evidenced in the cell transformation assay.•Gene pathways related to chronic lung diseases and cancer onset were modulated. The particulate matter represents one of the most complex environmental mixtures, whose effects on human health and environment vary according to particles characteristics and source of emissions. The present study describes an integrated approach, including in vitro tests and toxicogenomics, to highlight the effects of air particulate matter on toxicological relevant endpoints. Air samples (PM2.5) were collected in summer and winter at different sites, representative of different levels of air pollution. Samples organic extracts were tested in the BALB/c 3T3 CTA at a dose range 1–12m3. The effect of the exposure to the samples at a dose of 8m3 on the whole-genome transcriptomic profile was also assessed. All the collected samples induced dose-related toxic effects in the exposed cells. The modulated gene pathways confirmed that toxicity was related to sampling season and sampling site. The analysis of the KEGG’s pathways showed modulation of several gene networks related to oxidative stress and inflammation. Even if the samples did not induce cell transformation in the treated cells, gene pathways related to the onset of cancer were modulated as a consequence of the exposure. This integrated approach could provide valuable information for predicting toxic risks in humans exposed to air pollution.