title: Transient permeabilization of airway epithelium by mucosal water.
abstract: We describe a simple hyposmotic shock procedure whereby the apical membrane of airway epithelium can be made transiently leaky to proteins and other macromolecules. Bovine or human tracheal epithelial cells were grown as confluent polarized cell sheets on porous inserts. While physiological saline was maintained on the basolateral surface, the mucosal surface was exposed to water. This led to marked increases in the uptake of [14C]mannitol across both apical and basolateral membranes. On restoring saline to the mucosal surface, the [14C]mannitol permeability returned to preexposure levels with a half-life of approximately 5 min. Mucosal water also increased efflux of lactate dehydrogenase and the uptakes of fluorescent albumin and dextran (2,000 kDa). Water-induced increases in mannitol permeability were similar at 4 and 37 degrees C, suggesting that pinocytosis was not the mechanism. Detailed time courses of the uptake of dextran and the loss of lactate dehydrogenase and 36Cl showed that the bulk of the permeability increase occurred during the first 2- to 4-min exposure to water. Transepithelial resistance was reversibly decreased by exposure to water, but short-circuit current responses to transport blockers and secretagogues remained qualitatively normal. The hyposmotic shock procedure also successfully permeabilized apical membranes of primary cultures of nasal epithelial cells from a patient with cystic fibrosis (CF) and of JME/CF 15 cells, a cell line derived from CF bronchial epithelium. This simple and efficient procedure may prove useful in studies on the cell and molecular biology of airway and other epithelia.
authors.affiliation: Children's Hospital Oakland Research Institute, California 94609-1809, USA.
authors.forename: J H
authors.initials: JH
authors.lastname: Widdicombe
authors.forename: F
authors.initials: F
authors.lastname: Azizi
authors.forename: T
authors.initials: T
authors.lastname: Kang
authors.forename: J F
authors.initials: JF
authors.lastname: Pittet
mesh_terms: D000818:Animals; D001834:Body Water / Q000378:metabolism; D002417:Cattle; D002460:Cell Line; D004594:Electrophysiology; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D005456:Fluorescent Dyes; D006207:Half-Life; D006801:Humans; D007477:Ions; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008353:Mannitol / Q000378:metabolism; D009077:Mucins / Q000378:metabolism; D010539:Permeability; D011506:Proteins / Q000378:metabolism; D012137:Respiratory System / Q000378:metabolism*; D014132:Trachea / Q000166:cytology
chemical_list: D005456:Fluorescent Dyes; D007477:Ions; D009077:Mucins; D011506:Proteins; D008353:Mannitol; D007770:L-Lactate Dehydrogenase