The objective of this proposal is to elucidate the cellular and molecular mechanisms by which ozone causes asthma exacerbations, with the overarching goal of informing the rationale design of clinical trials targeting pathways identified by this investigation. Our preliminary data have led us to the overall hypothesis that following ozone exposure, ATP released by airway epithelia leads to mast cell activation, and mast-cell-derived proteases, lipid mediators, and cytokines augment airway inflammation and AHR. To test this hypothesis we will first measure ATP and other nucleotide/side levels in airway epithelial cell cultures form normal volunteers and asthmatics and in the airways of nave and house dust mite (HDM)-allergic mice exposed to ozone. We will determine if ATP is more potent than its degradation products at inducing AHR and airway inflammation in allergic mice; we will manipulate ATP levels in vivo, using both mice lacking proteins important in extracellular ATP release and degradation as well as pharmacological inhibitors, and determine the impact of these changes on ozone-induced phenotypes. Next, to determine if mast cells are critical intermediaries between ATP and AHR/airway inflammation, we will measure mast cell degranulation in the murine airway after ATP and ozone exposures; we will expose mast cell deficient mice to ATP and ozone; and we will determine if ATP released by epithelial cell cultures exposed to ozone induces mast cell activation in vitro. Finally, we will determine the indirect and direct molecular mechanisms by which epithelial-derived ATP activates mast cells by measuring known mast cell activators released by epithelial cell cultures and examining their capacity to stimulate mast cells alone and in combination. Since the cellular and molecular mechanisms by which ozone contributes to asthma exacerbation remain incompletely understood, this work will advance our understanding of environmental asthma pathogenesis and reveal new avenues of therapy.
Asthma attacks are important causes of acute illness in asthmatics, resulting in lost time from school or work, physician visits, and hospital admissions, all at a substantial cost to the public. Ozone is an important cause of asthma attacks by causing inflammation and airway constriction. This project seeks to understand the ways in which ozone contributes to asthma attacks so that new therapies can be developed.