P044 Pollutant particulate matter acts on bronchial epithelial and dendritic cells to promote a pro-inflammatory environment

Abstract

Introduction Airborne particulate matter (PM) from ambient pollution is believed to exacerbate asthmatic disease, although the mechanisms underpinning this are unclear. Dendritic cells (DC) orchestrate immune responses in the airways, and lie directly underneath human bronchial epithelial cells (HBEC) that line the airways. HBEC are the first cells to encounter PM entering the lungs. This study explores the relationship of DC and HBEC following exposure to foreign noxious material such as PM. Methods Primary PM samples were obtained from the KCL Environmental Research Group. Pollutants were extracted from filters set up within the London Low Emission Zone to monitor the association of PM composition with firework activity. NIST SRM1648a (standard reference material) was obtained from National Institute of Standards and Technology. Due to the limit of primary sample supply, NIST was used as a surrogate prototypical urban ambient pollution PM. Oxidative potentials of PM were quantified by depletion rate of 2 μg/ml ascorbic acid.Cytokines TNF-α, GM-CSF, IL-6, IL-8, IL-10 and IL-1β in supernatants were determined by Cytokine Bead Array (CBA). Viability was measured using propidium iodide and Annexin V staining. Myeloid dendritic cells (Cd11c+) were isolated from peripheral blood mononuclear cells. Dendritic cells were cultured alone, or together with epithelial cells. Supernatants are collected after 24 h. CD80 surface staining was analysed by flow cytometry. Results The capacity of individual primary PM extracts to stimulate IL-6, IL-8 and GM-CSF production by primary bronchial epithelial cells strongly correlate with the oxidative potential of individual PM extracts, as measured by ascorbic depletion rates. Furthermore, by inducing antioxidants using sulforaphane, stimulation of these cytokines by PM was suppressed due to enhanced protection against oxidative stress. Dendritic cells alone produce a similar pro-inflammatory response to a high oxidative potential PM. In co-culture, stimulation with PM greatly increases the cytokine production compared to individually cultured epithelial cells or mDCs. IL-10 and IL-1β, which were both non-detectable in epithelial cells are also increased in dendritic cells co-cultured with HBEC and stimulated with PM. Conclusion Oxidative stress has been identified as a major determinant of the capacity of PM to induce pro-inflammatory cytokines in primary human bronchial epithelial cells. Epithelial cells interact with dendritic cells in co-cultures stimulated with PM to greatly potentiate pro-inflammatory responses.