Prostaglandin E2 Regulation of Cystic Fibrosis Transmembrane Conductance Regulator Activity and Airway Surface Liquid Volume Requires Gap Junctional Communication
Stimulation of the cystic fibrosis transmembrane conductance regulator (CFTR) by protease-activated receptors (PARs) at the basolateral membranes and by adenosine receptors (ADO-Rs) at the apical membrane maintain airway surface liquid (ASL) volume, which is required to ensure hydrated and clearable...
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Published in | American journal of respiratory cell and molecular biology Vol. 44; no. 1; pp. 74 - 82 |
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Main Authors | , , , , , , |
Format | Journal Article |
Language | English |
Published |
American Thoracic Society
01.01.2011
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Abstract | Stimulation of the cystic fibrosis transmembrane conductance regulator (CFTR) by protease-activated receptors (PARs) at the basolateral membranes and by adenosine receptors (ADO-Rs) at the apical membrane maintain airway surface liquid (ASL) volume, which is required to ensure hydrated and clearable mucus. Both pathways involve the release of prostaglandin E
2
(PGE
2
) and the stimulation of their basolateral receptors (EP-Rs). We sought to determine whether gap junctions contribute to the coordination of these pathways for modulating CFTR activity and mucus hydration. We used RT-PCR and Western blotting to determine connexin (Cx), CD73, and EP-R expression in a Calu-3 airway epithelial cell line grown on Transwell (Corning Costar, Cambridge, MA) inserts. We used dye coupling to evaluate gap junctional intercellular communication (GJIC). We used Ussing chamber studies and X-Z confocal microscopy to monitor Cl
−
secretion and ASL volume regulation. We found that connexin 43 (Cx43)–mediated GJIC was increased either by endogenous ADO after the hydrolysis of purine nucleotides by CD73 or by the direct activation of ADO-Rs. Inhibition of phospholipase A2 and cyclooxygenase prevented ADO-dependent increases in GJIC, suggesting the involvement of PGE
2
. PGE
2
was found to increase GJIC markedly by stimulating EP4-Rs. The modulation of ADO signaling also affected the PAR-dependent activation of CFTR. The reduction of GJIC by CD73 or Cx43 inhibition prevented PAR-evoked CFTR currents in Ussing chambers. The inhibition of GJIC resulted in a failure of PGE
2
to increase ASL volume in Calu-3 cells and in primary cultures of well-differentiated human airway epithelial cells. Thus, gap junctions coordinate a signaling network comprising CFTR, ADO-Rs, PARs, and EP-Rs, and are required for ASL volume homeostasis. |
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AbstractList | Stimulation of the cystic fibrosis transmembrane conductance regulator (CFTR) by protease-activated receptors (PARs) at the basolateral membranes and by adenosine receptors (ADO-Rs) at the apical membrane maintain airway surface liquid (ASL) volume, which is required to ensure hydrated and clearable mucus. Both pathways involve the release of prostaglandin E
2
(PGE
2
) and the stimulation of their basolateral receptors (EP-Rs). We sought to determine whether gap junctions contribute to the coordination of these pathways for modulating CFTR activity and mucus hydration. We used RT-PCR and Western blotting to determine connexin (Cx), CD73, and EP-R expression in a Calu-3 airway epithelial cell line grown on Transwell (Corning Costar, Cambridge, MA) inserts. We used dye coupling to evaluate gap junctional intercellular communication (GJIC). We used Ussing chamber studies and X-Z confocal microscopy to monitor Cl
−
secretion and ASL volume regulation. We found that connexin 43 (Cx43)–mediated GJIC was increased either by endogenous ADO after the hydrolysis of purine nucleotides by CD73 or by the direct activation of ADO-Rs. Inhibition of phospholipase A2 and cyclooxygenase prevented ADO-dependent increases in GJIC, suggesting the involvement of PGE
2
. PGE
2
was found to increase GJIC markedly by stimulating EP4-Rs. The modulation of ADO signaling also affected the PAR-dependent activation of CFTR. The reduction of GJIC by CD73 or Cx43 inhibition prevented PAR-evoked CFTR currents in Ussing chambers. The inhibition of GJIC resulted in a failure of PGE
2
to increase ASL volume in Calu-3 cells and in primary cultures of well-differentiated human airway epithelial cells. Thus, gap junctions coordinate a signaling network comprising CFTR, ADO-Rs, PARs, and EP-Rs, and are required for ASL volume homeostasis. |
Author | Bacchetta, Marc Losa, Davide Scheckenbach, K. E. Ludwig O'Grady, Scott Dudez, Tecla Crespin, Sophie Chanson, Marc |
AuthorAffiliation | 1 Laboratory of Clinical Investigation III, Department of Pediatrics, Geneva University Hospitals and University of Geneva, Geneva, Switzerland; and 2 Department of Integrative Biology and Physiology, University of Minnesota, St. Paul, Minnesota |
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Notes | Originally Published in Press as DOI: 10.1165/rcmb.2009-0361OC on February 18, 2010 Author Disclosure: None of the authors has a financial relationship with a commercial entity that has an interest in the subject of this manuscript. This work was supported by the Swiss National Science Foundation (grant 310000–119739), Vaincre la Mucoviscidose, the Schweizerische Gesellschaft für Cystische Fibrose, and the National Institutes of Health (grant DK07401). K.E.L.S. was supported by the Schmidheiny Foundation and the Novartis Consumer Health Foundation. D.L. was supported by the Italian Cystic Fibrosis Research Foundation (grant FFC#19/2009), adopted by the FFC delegation “La Bottega delle Donne.” |
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Title | Prostaglandin E2 Regulation of Cystic Fibrosis Transmembrane Conductance Regulator Activity and Airway Surface Liquid Volume Requires Gap Junctional Communication |
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