B97-09 Impact of Aeroallergens and Woodsmoke on Differentiated Airway Epithelial Cells

Abstract Rationale Climate change is intensifying aeroallergen exposure by prolonging pollen seasons, increasing pollen counts, and shifting the growth and distribution of fungal species. These aeroallergens may act synergistically with climate-driven increases in wildfire smoke, creating complex exposures that may compromise airway epithelial barriers. Understanding how these stressors converge at the airway epithelium is key to clarifying their impact on respiratory health. Methods Human bronchial epithelial cells (HBECs) were obtained from five healthy donors (3 male: 2 female) undergoing bronchoscopies and expanded in PneumaCult-NGEx media (STEMCELL Technologies) to passage 3-5. HBECs were differentiated at air-liquid interface (ALI) for 21 days using Pneumacult-ALI media, as per the manufacturer’s recommendation, and ciliary movement and mucus production were visually confirmed. In submerged condition, HBECs were exposed to four types of allergens (Cladosporium cladosporioides, Birch Mix, Ragweed Mix, and Aspergillus fumigatus), at varying concentrations. RNA lysates were collected at 4h post-allergen exposure. qPCR was performed to determine interleukin (IL)-8 and IL-33 responses to allergen exposures. Differentiated HBECs were, alternatively, exposed to ragweed (diluted in PBS) for 1h, followed by exposure to photochemically aged pine wood smoke (WS; PM2.5 concentration of 140µg/m3) for 2h using the Cultex RFS system (Cultex Technology). Cell culture media samples from the basolateral chamber were collected at 24h post-exposure, and IL-8 expression was quantified by ELISA. Transepithelial electrical resistance (TEER) was measured before and after each exposure. Results To determine the appropriate allergen exposure dose for ALI experiments, we determined that 0.137 allergen units (AU)/mL ragweed exposure induced IL-8 and IL-33 responses of the same magnitude as the lipopolysaccharide control (0.01 µg/mL) in submerged culture. In differentiated ALI cultured HBECs, 0.137 AU/mL of ragweed exposure for 1hr increased IL-8 concentration in cell culture media (basolateral compartment) (ragweed alone (mean±SEM): 287.8±68.7 pg/mL, p = 0.05; ragweed+WS: 326.74±95.7, p = 0.01; vs negative control 206.9±65.6 pg/mL) at 24h. Aged WS exposure alone did not increase IL-8 (209.1±67.6 pg/mL, vs control, p 0.05). TEER increased with WS exposure (TEER (W/cm2) in WS-alone: 977±184 vs control: 648±110, p = 0.03), but not with allergen exposure (ragweed-alone: 771±119, p 0.05). Conclusions Our results reveal distinct yet complementary effects of allergens and wildfire smoke on epithelial inflammation and barrier function, both of which may contribute to the development or exacerbation of respiratory disease. Ongoing multi-omics and functional analyses will further delineate the molecular circuits underpinning these interactions and reveal their implications for climate-related respiratory health. This abstract is funded by: GenomeBC SIP033