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Ozone (O3) exposure of rats increases airway epithelial permeability. We hypothesized that this increased permeability may be mediated by the epithelial cell cytoskeleton. To test this hypothesis, we studied the effect of cytoskeletal disruption on the transmucosal transport of tracers from airway lumen to blood and compared the results with the effects of O3 exposure. No increase in transport occurred following disruption of microtubules by vinblastine, but disruption of microfilaments with cytochalasin D resulted in increased transport of radiolabeled tracers [99mTc- and 111In-labeled diethylenetriamine-pentacetate (DTPA) and 125I-labeled bovine serum albumin (BSA)]. In control rats, both horseradish peroxidase (HRP) and BSA, localized by cytochemistry and autoradiography, respectively, were detected on the epithelial cell surfaces and in endocytic vesicles. In rats treated with cytochalasin D or exposed to O3, the tracer molecules also penetrated the intercellular spaces, though the apical tight junctions remained devoid of the tracers. Increased numbers of endocytic vesicles containing HRP and aggregation of 125I-labeled BSA autoradiographic grains in the subepithelial region were also seen after either treatment. We conclude that destabilization of cytoskeletal elements following O3 exposure is a possible mechanism of increased transmucosal transport, which may be a combined effect of accelerated transport through both endocytic and paracellular pathways.

Full-length tracheas from Sprague-Dawley rats were exposed to cytoskeleton-active drugs in short-term organ culture, and the permeability of the tracheal epithelium was measured by instilling radiotracers into the lumen and assay of the radioactivity appearing in the external bathing medium. In vitro treatment with cytochalasin D (cyto D, 2-10 x 10(-6) M) increased the rate of movement of [14C]mannitol across the epithelium. Exposure to vinblastine (VB, 10(-4) M) alone had no significant effect. However, VB in combination with cyto D increased the permeability in a dose-dependent manner. In vivo exposure to ozone (O3, 0.8 or 2.0 ppm, 2 h) had only a slight effect on the rate of movement of the tracer as measured in vitro immediately after exposure. At 24 h postexposure there was no significant difference in permeability between ozone- and air-exposed tracheas. Prior in vivo O3 exposure sensitized the tracheas to the in vitro effects of cyto D; treatment of O3-exposed tracheas with cyto D immediately after O3 exposure produced a greater than additive effect on permeability measured in vitro. VB at concentrations up to 10(-4) M had no enhancing effect on permeability in O3-exposed tracheas. Sham exposure to clean air did not affect permeability compared to untreated (shelf) controls. Electron microscopic studies demonstrated penetration of horseradish peroxidase into intercellular spaces in the tracheas treated in vitro with cyto D or cyto D plus VB. Cyto D is known to affect intracellular microfilaments that have attachments at or near the cell surface, while VB affects microtubules associated with internal cellular structures. Therefore, the synergistic effect on tracheal permeability observed with O3 and cyto D, but not with O3 and VB, suggests that O3 may change cell surface structures associated with the microfilamentous cytoskeleton.