To examine neutrophil transepithelial migration in the basolateral-to-luminal direction, bronchial epithelial cells (16HBE) were grown at an air-medium interface on the lower face of permeable supports, and resistance across each membrane was recorded before measuring neutrophil transmigration over 2 h. Subconfluent monolayers (resistance < 250 Omega) permitted high spontaneous migration of neutrophils (7.4+/-1%), which was further enhanced (29.7+/-3%) in response to interleukin (IL)-8 (100 ng/ml). Confluent monolayers (250 to 700 Omega) showed low spontaneous migration (2+/- 0.5%) but responded markedly to IL-8 (12.4+/-1.3%). Left in culture, 16HBE resistances continued to increase and were associated with minimal spontaneous migration (< 0.5%) or responses to IL-8. Using cells in the 250 to 700 Omega range, neutrophil migration to IL-8 was dose-dependent and was enhanced when epithelial cells were incubated with a combination of tumor necrosis factor-alpha and interferon-gamma. Neutrophil migration was stimulus-specific and was reduced by preincubation of epithelial cells with a F(ab')(2) anti-intercellular adhesion molecule (ICAM)-1, or by preincubation of neutrophils with anti-CD18, anti-CD11a, anti-CD11b, or anti-CD11c, but not by anti-CD11d, indicating a role for beta(2)-integrin-ICAM-1 interaction in the migration process.