title: Cellular mechanisms of mainstream cigarette smoke-induced lung epithelial tight junction permeability changes in vitro.
abstract: Mainstream cigarette smoke increases the permeability of human airways; however, the mechanism for this increased permeability is poorly defined. Tight junctions between adjacent epithelial cells constitute the physiological barrier to fluid and macromolecules in epithelium. These structures are highly regulated by phosphorylation and their association with the cytoskeleton. The goal of these studies was to identify the signal transduction pathways that regulate smoke-induced permeability. Using a physiologically relevant air-liquid interface exposure system, electrically tight monolayers of the human bronchial epithelial cell-line Calu-3 were exposed to fresh, whole mainstream cigarette smoke. This exposure results in a regulated, dose-dependent loss of epithelial barrier function in the lung epithelial monolayers. With cigarette smoke exposure, transepithelial electrical resistance (TER) is decreased and albumin flux is increased, indicating a loss in barrier function to ions and macromolecules, respectively; however, both largely recover in 30 min. Smoke-induced losses of macromolecular barrier function are the result of multicellular junctional reorganization, resulting in increased leak volume rather than leak frequency. Inhibiting Rho kinase (ROCK) significantly reduces the smoke-induced permeability to both ions and macromolecules, while inhibiting protein tyrosine kinases (PTK) only reduces smoke-induced macromolecular permeability. Interestingly, inhibiting myosin light chain kinase (MLCK) exacerbates smoke-induced permeability, indicating that MLCK and ROCK have opposing regulatory roles. Our results demonstrate that the smoke-induced loss of epithelial barrier function in human bronchial epithelium is a regulated process rather than a cytotoxic response. Additionally, our results indicate that activation of PTK and ROCK and inactivation of MLCK contribute to the increased airway permeability caused by mainstream cigarette smoke.
authors.affiliation: Program in Toxicology, College of Pharmacy, University of New Mexico Health Sciences Center, Albuquerque, New Mexico, USA.
authors.forename: Dorian S
authors.initials: DS
authors.lastname: Olivera
authors.forename: Susan E
authors.initials: SE
authors.lastname: Boggs
authors.forename: Chris
authors.initials: C
authors.lastname: Beenhouwer
authors.forename: James
authors.initials: J
authors.lastname: Aden
authors.forename: Cindy
authors.initials: C
authors.lastname: Knall
mesh_terms: D000577:Amides / Q000494:pharmacology; D001381:Azepines / Q000494:pharmacology; D018929:Cell Culture Techniques / Q000295:instrumentation; D045744:Cell Line, Tumor; D002463:Cell Membrane Permeability / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D017097:Electric Impedance; D005456:Fluorescent Dyes / Q000378:metabolism; D019833:Genistein / Q000494:pharmacology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D008168:Lung / Q000187:drug effects* / Q000378:metabolism; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D009281:Naphthalenes / Q000494:pharmacology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology; D020545:Respiratory Mucosa / Q000187:drug effects* / Q000378:metabolism; D012710:Serum Albumin, Bovine / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D012906:Smoke / Q000009:adverse effects*; D019108:Tight Junctions / Q000187:drug effects* / Q000378:metabolism; D013997:Time Factors; D014026:Nicotiana; D054460:rho-Associated Kinases
chemical_list: D000577:Amides; D001381:Azepines; D005456:Fluorescent Dyes; D047908:Intracellular Signaling Peptides and Proteins; D009281:Naphthalenes; D047428:Protein Kinase Inhibitors; D011725:Pyridines; D012906:Smoke; C070571:ML 7; C108830:Y 27632; D012710:Serum Albumin, Bovine; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D009219:Myosin-Light-Chain Kinase