Citation:

McCullough, S. Breaking the Barrier: Adverse Cellular and Molecular Effects of Air Pollutant Exposure Beyond the Airway Epithelium. 2018 Genetics and Environmental Mutagenesis Society Fall Meeting, Durham, NC, November 07, 2018.

Impact/Purpose:

Presentation of data on the adverse impacts of air pollutant exposures on different cell types within the human airway. Mechanistic data that feed into single- and multi-dimensional adverse outcome pathway (AOP) development were also presented.

Description:

Air pollutant exposure causes the deaths of approximately eight million people and contributes to adverse health outcomes of millions more worldwide every year. These effects are driven by the exposure-mediated induction of oxidative stress and inflammation; however, despite the impact on public health, our ability to identify susceptible populations, establish risk mitigation strategies, and develop therapeutic interventions is hindered by an incomplete understanding of the associated cellular and molecular mechanisms. Advancing the protection of public health will require the use of robust and broadly-applicable in vitro organotypic models to develop single- and multi-dimensional adverse outcome pathways by providing insight into the role of key cell types in the airway microenvironment, which cannot be readily accomplished with traditional in vivo animal and in vitro monoculture cell-based models. Here we tested the hypothesis that “trans-epithelial” exposure to diesel exhaust particulates (DEP), a ubiquitous air pollutant, will alter oxidative stress and pro-inflammatory signaling within the airway microenvironment using a novel physiologically-relevant in vitro organotypic model. Our “trans-epithelial exposure model” (TEEM) incorporates human bronchial epithelial cells (HBEC) barrier grown on a collagen matrix, which serve as a functional barrier between the host and the environment, in the apical compartment of a Transwell insert and human lung fibroblasts (HLF), which play a critical role in regulating airway structure, function, and homeostasis, in the basolateral compartment. Time course (2-24 hours) analysis of oxidative stress responsive and pro-inflammatory gene expression in both the HBEC and HLF following exposure of the HBEC to DEP (“trans-epithelial DEP,” or TE-DEP, exposure) demonstrated that the kinetics and magnitude of oxidative stress-responsive gene expression is similar between the two cell types; however, peak pro-inflammatory gene induction in HLF was delayed, but more prolonged in HLF relative to HBEC. Genes involved in glutathione homeostasis and hydrogen peroxide (H2O2) signaling (NQO1, TRX1, PTGS2 and GCLM1) were also alternatively regulated in response to DEP exposure, and their induction was attenuated by pre-treatment with the free radical scavenger, N-acetyl-cysteine (NAC). Using a novel dual fluorescent biosensor to measure intracellular glutathione oxidation and H2O2 accumulation (roGFP2/HyPER red) we observed an increase in glutathione oxidation and H2O2 accumulation in HLF following TE-DEP exposure. The findings presented here demonstrate that DEP exposures have trans-epithelial effects on the airway microenvironment and suggest the need for further study to characterize the associated molecular mechanisms to develop higher resolution adverse outcome pathways and improve the protection of public health.

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