Differential toxicity in an alveolar epithelial cell line of fine particulate matter from brakewear, roadwear, and diesel exhaust

Introduction: Airborne fine particulate matter (diameter <2.5μm; PM2.5) is a leading risk factor for lung disease. Improved exhaust technology and car electrification are decreasing exhaust-derived PM emissions but without a similar decrease in poorly characterised, unregulated “non-exhaust” PM, predominantly brakewear PM (BWPM). This study aimed to characterise the toxicology of PM2.5 from 4 brake pad types compared to roadwear and diesel exhaust (DEP). Methods: An alveolar type-II epithelial cell line (ATII^ER:KRASV12) was exposed to 12.5-100µg/ml PM2.5 from 4 brake pad types (low-metallic, semi-metallic, non-asbestos organic, ceramic), roadwear, or DEP for 2, 6 or 24hrs. Cytotoxicity was measured by LDH/MTT assays. Cytokine release (IL-6, IL-8) was assayed by ELISA. ROS generation was assessed using DCFDA. Expression of antioxidant gene haemoxygenase-1 (HMOX1) and metal-binding metallothionein 1G and 2A (MT1G, MT2A) was determined by RT-qPCR. Results: Exposure to all PM2.5 types elicited concentration-dependent increases in both cytotoxicity, IL-6/IL-8 release, and ROS generation with non-asbestos organic and ceramic BWPM being the most potent inducers of these endpoints. Furthermore, at non-cytotoxic concentrations, only the non-asbestos organic and ceramic BWPM elicited significant upregulation of genes involved in oxidative stress (HMOX1), metal-binding (MT1G/MT2A), and inflammation (IL-6, IL-8). Conclusions: ATII^ER:KRASV12 responses were dependent on PM source, with non-asbestos organic and ceramic BWPM most potent. Future work will further characterise the cellular effects of these PM types and how these relate mechanistically to their composition.