Cytoskeletal modulation and tyrosine phosphorylation of tight junction proteins are associated with mainstream cigarette smoke-induced permeability of airway epithelium.
Cigarette smoke increases the permeability of the lung epithelium. Consequences of increased permeability include increased access of toxins and pathogens from the air spaces to the interstitium and even the blood stream, and leakage of fluids into the air spaces. The mechanisms for permeability alterations have not been elucidated for airway epithelia. By analogy with other types of epithelia, we hypothesized that changes in the phosphorylation status and function of tight junction (TJ) or cytoskeletal proteins might mediate the smoke-induced permeability changes. We investigated the effects of exposure to mainstream cigarette smoke (MS) on cultures of Calu-3 cells, an airway epithelial cell line. Specifically, MS exposure caused increases in phosphorylation of the myosin-binding subunit (MBS) of myosin phosphatase and myosin light chain (MLC), proteins involved in the regulation of actin polymerization. These results implicate activation of Rho kinase (ROCK), consistent with previously reported data indicating that inhibition of ROCK activation suppressed MS-induced increases in permeability. MS exposure also increased polymerized (filamentous) actin (f-actin) content and caused redistribution of the TJ proteins from the normal apical circumferential band to a more basal location. The translocation of the TJ proteins was spatially associated with local increases in both f-actin and macromolecular permeability. Finally, MS exposure increased tyrosine phosphorylation of occludin but not ZO-1 and decreased association between the two TJ proteins. These results indicate that MS exposure causes alterations in cytoskeletal and TJ structure and function, resulting in increased macromolecular permeability that may contribute to the adverse health effects of MS.
Division of Pathophysiology, Lovelace Respiratory Research Institute, 2425 Ridgecrest SE, Albuquerque, NM 87108, USA.
Dorian
D
Olivera
Cindy
C
Knall
Susan
S
Boggs
JeanClare
J
Seagrave
D000199:Actins / Q000187:drug effects / Q000378:metabolism; D045744:Cell Line, Tumor; D002463:Cell Membrane Permeability / Q000187:drug effects*; D003598:Cytoskeletal Proteins / Q000187:drug effects; D003599:Cytoskeleton / Q000187:drug effects*; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D010766:Phosphorylation; D020545:Respiratory Mucosa / Q000187:drug effects*; D012906:Smoke / Q000009:adverse effects*; D012907:Smoking / Q000009:adverse effects; D019108:Tight Junctions / Q000187:drug effects* / Q000378:metabolism; D014026:Nicotiana / Q000009:adverse effects* / Q000378:metabolism; D014443:Tyrosine / Q000187:drug effects
D000199:Actins; D003598:Cytoskeletal Proteins; D012906:Smoke; D014443:Tyrosine