AN INVITRO SYSTEM TO MODEL PULMONARY EPITHELIAL BARRIER DYSFUNCTION MEDIATED BY IMMUNE EFFECTOR-CELLS

An assay of epithelial barrier function was developed to monitor immun e-mediated changes in lung permeability that may be occurring during p ulmonary allograft rejection and inflammatory lung diseases. Lung tiss ue was obtained from minipigs, digested with collagenase (1 mg/ml) ove rnight, and propagated in RPMI 1640 tissue culture medium. Cells with an epithelioid morphology were purified by differential detachment usi ng trypsin-ethylenediaminetetraacetic acid and were characterized as e pithelial by positive staining with an anti-cytokeratin monoclonal ant ibody. Monolayers of these epithelial cells were cultured on porous ti ssue culture inserts, and transmonolayer resistance values were measur ed. Transmonolayer resistance values reached a mean of 5487 +/- 2882 O MEGA (mean +/- 95% confidence interval; n = 9) after 5 days in culture . These values indicated the presence of functional intercellular tigh t junctions between the cells. Addition of cytotoxic immune effector c ells to the cultured monolayers caused a rapid reduction in the transm onolayer resistance values, whereas unstimulated splenocytes failed to produce this effect. Comparison of these results with those obtained in parallel experiments performed with standard isotopic cytotoxicity assays indicated the sensitivity of the transmonolayer resistance tech nique. The assay described in this report will enable in vitro modelin g of epithelial permeability damage mediated by both activated lymphoi d cells and their soluble products.