Novel whole wood smoke exposure and imaging system for human bronchial epithelial cells cultured at the air-liquid interface
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Exposure to air pollutants is a leading cause of human morbidity and mortality. The frequency and severity of wildland fires, significant contributors to ambient air particulate matter, are projected to increase. The conventional in vitro approach to studying wood smoke effects is to expose submerged cell line cultures to wood smoke condensates or particle extracts, which fails to capture the complexity of wood smoke, especially its volatile fraction. Fully differentiated primary human bronchial epithelial cells cultured at the air-liquid interface (HBEC-ALI) closely approximate the in vivo morphology of the bronchial epithelium. Coupled with genetically encoded fluorescence sensors such as Grx1-roGFP2, the high temporal resolution of live-cell microscopy is well suited for monitoring intracellular redox events that underlie the toxicity of air pollution. In order to integrate real-world wildland fire exposures, high temporal resolution microscopy, and the in vivo relevance of HBEC-ALI, we set out to develop a system to enable live-cell imaging of redox events in HBEC-ALI as they undergo exposure to wood combustion emissions generated in real-time. Wood is pyrolyzed in a tube furnace at controlled temperature and airflow to generate emissions that are conditioned with humidity, CO2, and temperature before entering a custom-built chamber that permits confocal imaging of HBEC-ALI expressing Grx1-roGFP2. Early results show that real-time exposure to wood smoke induces glutathione oxidation, a marker of oxidative stress, in HBEC-ALI in a dose-dependent manner. These findings demonstrate the feasibility of studying redox events in the human bronchial epithelium during exposure to wildland fire smoke. this abstract of a proposed presentation does not necessarily reflect epa policy.