Epithelial sodium channel silencing as a strategy to correct the airway surface fluid deficit in cystic fibrosis

Ambra Gianotti, Raffaella Melani, Emanuela Caci, Elvira Sondo, Roberto Ravazzolo, Luis J V Galietta, Olga Zegarra-Moran

Research output: Contribution to journalArticlepeer-review

Abstract

In the respiratory system, Na+ absorption and Cl- secretion are balanced to maintain an appropriate airway surface fluid (ASF) volume and ensure efficient mucociliary clearance. In cystic fibrosis (CF), this equilibrium is disrupted by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, resulting in the absence of functional CFTR-dependent Cl- secretion. The consequences of defective Cl - transport are worsened by the persistence of Na+ absorption, which contributes to airway surface dehydration. We asked whether normal ASF can be restored to an equal extent by recovering Cl- secretion from mutated CFTR or by reducing Na+ absorption. This is highly relevant in the selection of the best strategy for the treatment of patients with CF. We analyzed the ASF thickness of primary cultured bronchial CF and non-CF epithelia after silencing the epithelial Na+ channel (ENaC) with specific short, interfering RNAs (siRNAs) and after the pharmacological stimulation of CFTR. Our results indicate that (1) single siRNAs complementary to ENaC subunits are sufficient to reduce ENaC transcripts, Na+ channel activity, and fluid transport, but only silencing both the α and β ENaC subunits at the same time leads to an increase of ASF (from nearly 7 μm to more than 9 μm); (2) the ASF thickness obtained in this way is about half that measured after maximal CFTR stimulation in non-CF epithelia (10-14 μm); and (3) the pharmacological rescue of mutant CFTR increases the ASF to the same extent as ENaC silencing. Our results indicate that CFTR rescue and ENaC silencing both produce a significant and long-lasting increase of airway hydration in vitro.

Original languageEnglish
Pages (from-to)445-452
Number of pages8
JournalAmerican Journal of Respiratory Cell and Molecular Biology
Volume49
Issue number3
DOIs
Publication statusPublished - Sep 2013

Keywords

  • Airway surface fluid
  • CFTR
  • Cystic fibrosis
  • ENaC
  • siRNA

ASJC Scopus subject areas

  • Cell Biology
  • Pulmonary and Respiratory Medicine
  • Molecular Biology
  • Clinical Biochemistry

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