40 Establishing a Physiologically Relevant in Vitro Airway Model to Assess the Health Impacts from Different Fungal Species

Abstract Allergic airway disease (AAD) is a collective term for respiratory disorders that can be exacerbated upon exposure to airborne allergens, including fungi and other air pollutants. Fungi are of particular interest as they are present in both outdoor and indoor air and are associated with asthma and allergy. A recent literature review identified key pathways contributing to the allergic responses to fungal allergens. However, due to the heterogeneity of models and methods used, it is not clear whether there are common or specific pathways induced by different fungi/fungal components. Air-liquid interface (ALI) culture was used to induce differentiation of nasal (RPMI 2650), bronchial (HBEC3-KT, BEAS-2B, Calu-3) and small airways (A549) epithelial cells into a columnar stratified epithelium with cilia, goblet cells, and the formation of tight junctions. Cultures were harvested at 0, 7, 14, 21 and 28 days, and the expression of molecular markers of basal, ciliated and mucus producing cells were assessed with reverse-transcription quantitative PCR (RT-qPCR). Different cell lines produce subtly different differentiated models, with downregulation of basal (undifferentiated) cell markers and upregulation of markers of ciliated and mucus producing cells from day 7. While no ciliated cells were observed in any of the differentiated cell lines, mucus was produced in some. The differentiation of primary cell lines at ALI, and expression of genes of interest (including known fungal receptors and genes identified in driving fungal allergy) were also measured to inform selection of appropriate model(s) to better understand the allergenic response to fungi/fungal components.