Toxicity of polycyclic aromatic hydrocarbons involves NOX2 activation

Abstract

Polycyclic Aromatic Hydrocarbons (PAHs) are environmental pollutants.The present study compares the toxic effects of BaP alone and a mixture of PAHs on human breast cancer cells. We hypothesize that PAH mixture is more toxic than BaP alone, and an increased NOX2 activation is related to PAH-induced oxidative stress. Initially, we exposed cultured human breast cancer cells to BaP alone (125 ng/mL and 500 ng/mL) and a mixture of PAHs (125 ng/mL and 500 ng/mL). After 24 h of exposure, the PAH mixture demonstrated a significant (P < 0.05) reduction in cell viability. The higher concentration of BaP alone (500 ng/mL) and both 125 ng/mL and 500 ng/mL PAH mixture significantly (P < 0.05) increased lactate production by MDA-MB-231 cells. We had observed an identical level of increased lactate levels when the cells were exposed to PAHs for 48 h. Flow cytometric analysis revealed that only PAHs mixture (both 125 ng/mL and 500 ng/mL) suppressed S phase significantly (P < 0.05). Finally, immunofluorescence microscopy was undertaken to examine the role of NOX2 due to PAHs toxicity. Colocalization of GP91phox and P47phox, a hallmark of NOX2 activation in the cell membrane of macrophage Kupffer cells demonstrated that higher concentration of BaP or PAH mixture showed increased colocalization events. These data suggest that the mixture of PAHs is more toxic and perturbing to DNA synthesis than BaP alone in cultured cells, and the toxicity is accompanied by NOX2 activation. Thus PAHs can lead to the increased burden of oxidative stress and alter the cellular redox status. Keywords: Polycyclic aromatic hydrocarbons, Benzo[a]pyrene, Toxicity, NOX2, Flow cytometry