The goal of the proposal is to define the cellular interactions between the oxidant air pollutant, ozone, and an inflammatory cell, the neutrophil, in long-term episodic exposure to ozone. Previous studies showed a notable delay in epithelial necrosis and a marked decrease in ciliated cells that remained reduced for an additional day in neutrophil-depleted rats exposed to ozone as compared to rats with neutrophils. We interpret these results as a beneficial effect of the neutrophil on the repair of the central acinar epithelium in ozone-induced injury. However, there is little information regarding the influence of neutrophils on the repair of air way and parenchymal epithelium with long-term exposure to ozone. We will use long-term episodic exposures which show a cumulative effect on the interstitium similar to long-term daily exposures to ozone. We believe the episodic exposure regiment will be the key to understanding the transition from the acute to the chronic ozone lesion. We will evaluate episodic exposures in two species: one which is relatively sensitive to ozone, the Rhesus monkey, and another which is relatively insensitive to ozone injury, the rat. We will evaluate the cellular responses (injury and repair) in the tracheobronchial airways and central acinus to long-term episodic ozone exposure in Rhesus monkeys and rats with and without the influence of neutrophils. We will also evaluate the influence of neutrophils and their components on cellular repair from ozone-induced injury in 1) explant airways isolated as microdissected bronchi or bronchioles from ozone- exposed Rhesus monkeys and rats and 2) purified monkey airway epithelial cells, rat alveolar epithelial cells and transformed human bronchial epithelial cells exposed to ozone in vitro. These studies will define: 1) whether neutrophils specifically kill ozone-injured cells and/or release granule components that indirectly or directly stimulated epithelial cells to proliferate, 2) whether delayed epithelial repair in neutrophil-depleted monkeys or rats will result in enhanced collagen accumulation and smooth muscle hypertrophy in airway walls and a metaplastic epithelium, 3) whether ozone exposure inhibits epithelial barrier function, increases DNA damage and apoptosis, and kills epithelial cells in vitro, 4) whether neutrophils preferentially killing apoptotic epithelial cells previously exposed to ozone in vitro, and 5) whether neutrophil-dependent epithelial cell killing requires a microenvironment established by adherence receptors or other receptors on ozone-injured or -altered (apoptosis) epithelial cells and neutrophils.
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