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Scientific article
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Prostaglandin E₂regulation of cystic fibrosis transmembrane conductance regulator activity and airway surface liquid volume requires gap junctional communication

Published inAmerican journal of respiratory cell and molecular biology, vol. 44, no. 1, p. 74-82
Publication date2011
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 E₂ (PGE₂) 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 PGE₂. PGE₂ 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 PGE₂ 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.

Keywords
  • 5'-Nucleotidase/metabolism
  • Adenosine/metabolism
  • Blotting, Western
  • Cell Communication/drug effects
  • Cell Line
  • Cell Polarity
  • Chlorides/metabolism
  • Connexins/metabolism
  • Cyclooxygenase Inhibitors/pharmacology
  • Cystic Fibrosis Transmembrane Conductance Regulator/genetics/metabolism
  • Dinoprostone/metabolism
  • Epithelial Cells/drug effects/metabolism
  • GPI-Linked Proteins/metabolism
  • Gap Junctions/drug effects/metabolism
  • Homeostasis
  • Humans
  • Membrane Potentials
  • Microscopy, Confocal
  • Mucociliary Clearance/drug effects
  • Mucus/metabolism
  • Phospholipases A2/antagonists & inhibitors/metabolism
  • RNA Interference
  • Receptors, Prostaglandin E/metabolism
  • Receptors, Purinergic P1/metabolism
  • Respiratory Mucosa/drug effects/metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Signal Transduction/drug effects
  • Surface Properties
  • Time Factors
Citation (ISO format)
SCHECKENBACH, K E Ludwig et al. Prostaglandin E₂regulation of cystic fibrosis transmembrane conductance regulator activity and airway surface liquid volume requires gap junctional communication. In: American journal of respiratory cell and molecular biology, 2011, vol. 44, n° 1, p. 74–82. doi: 10.1165/rcmb.2009-0361OC
Main files (1)
Article (Published version)
accessLevelPublic
Identifiers
ISSN of the journal1044-1549
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356downloads

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