Ambient fine particulate matter (PM2.5) induces AhR-dependent proinflammatory responses and reactive oxygen species...

To investigate the cytotoxic effects of airborne particulate matter smaller than 2.5 μm (PM2.5) on human conjunctival epithelial cells and to elucidate whether these effects are mediated through activation of the aryl hydrocarbon receptor (AhR) signaling pathway.

Human conjunctival epithelial cells were exposed to PM2.5 (0, 12.5, 25, and 50 μg/mL) for 24 hours. Cell viability was assessed using an MTT-based colorimetric assay by measuring absorbance at 450 nm, and intracellular reactive oxygen species (ROS) production was evaluated using DCFH-DA staining followed by quantification of DCF fluorescence intensity. AhR activation was evaluated by analyzing cytoplasmic and nuclear fractions of mRNA via Western blot. The mRNA expression of AhR, its downstream target genes (CYP1A1, CYP1B1, AhRR), and inflammatory cytokines (IL-1β, IL-6, TNF-α) was quantified by real-time PCR. AhR involvement was confirmed using siRNA-mediated AhR knockdown cells.

PM2.5 induced dose-dependent reductions in cell viability and significant increases in intracellular ROS at 25 and 50 μg/mL. Western blot analysis showed decreased cytoplasmic AhR and increased nuclear AhR following PM2.5 exposure. PM2.5 significantly upregulated AhR, its target genes, and inflammatory cytokines. In AhR knockdown cells, PM2.5 failed to induce nuclear translocation of AhR, upregulation of target genes, ROS production, or inflammatory cytokine expression, indicating that PM2.5-induced oxidative and inflammatory responses are dependent on AhR signaling.

PM2.5 induces cytotoxicity, oxidative stress, and inflammation in human conjunctival epithelial cells through AhR activation. These findings identify AhR as a key molecular mediator of PM2.5-induced ocular surface damage and suggest that targeting AhR pathways may provide a potential therapeutic strategy for preventing air pollution-related ocular surface disease.