Ozone injury to alveolar epithelium in vitro does not reflect loss of antioxidant defenses.Toxicol Appl Pharmacol 1994; 125(1):59-69TA
The objective of this study was to characterize an in vitro model of oxidant gas toxicity, using primary cultures of alveolar type II cells maintained in serum-free medium, by evaluating (1) epithelial barrier function, (2) the stability of cellular antioxidant defenses, and (3) the response of alveolar epithelial barrier properties to ozone exposure. Antioxidant enzyme activities and glutathione levels were measured in rat type II cells that were freshly isolated, cultured for 1 day in serum-supplemented medium, and subsequently grown in serum-free nutrient medium. After measurement of peak bioelectric properties on Day 4 in primary culture, alveolar epithelial monolayers were exposed to ozone at various concentrations and lengths of exposure. Ozone-induced alterations in monolayer bioelectric properties and impairment of cellular organization were used to evaluate oxidant injury. The primary effect of ozone exposure was a dose-dependent increase in monolayer permeability, which resulted from damage to intercellular junctions and/or loss of epithelial integrity. Extensive and persistent permeability increases correlated with focal areas of epithelial degradation. The focal nature of ozone injury to alveolar epithelium in vitro suggests that individual cell susceptibility to oxidant stress may account for the overall decrement in barrier function. However, this sensitivity does not result from overall loss of antioxidant defenses associated with cell culture, as these monolayers (when cultured in serum-free medium) maintained their antioxidant enzyme activities and glutathione content at levels found in freshly isolated cells. We conclude that the sensitivity of these monolayers to ozone injury in vitro reflects a disproportionate degree of oxidant stress on cell membranes relative to intracellular antioxidant defenses, i.e., cellular susceptibility to oxidant injury may depend on the ratio of the surface area of the cell to its cytoplasmic volume.