Reduced GM1 ganglioside in CFTR-deficient human airway cells results in decreased β1-integrin signaling and delayed wound repair

Yutaka Itokazu, Richard E. Pagano, Andreas S. Schroeder, Scott M. O'Grady, Andrew H. Limper, David L. Marks

Research output: Contribution to journalArticle

22 Citations (Scopus)

Abstract

Loss of cystic fibrosis transmembrane conductance regulator (CFTR function reduces chloride secretion and increases sodium uptake, but it is not clear why CFTR mutation also results in progressive lung inflammation and infection. We previously demonstrated that CFTR-silenced airway cells migrate more slowly during wound repair than CFTR-expressing controls. In addition, CFTR-deficient cells and mouse models have been reported to have altered sphingolipid levels. Here, we investigated the hypothesis that reduced migration in CFTR-deficient airway epithelial cells results from altered sphingolipid composition. We used cell lines derived from a human airway epithelial cell line (Calu-3 stably transfected with CFTR short hairpin RNA (CFTR-silenced or nontargeting short hairpin RNA (controls. Cell migration was measured by electric cell substrate impedance sensing (ECIS. Lipid analyses, addition of exogenous glycosphingolipids, and immunoblotting were performed. We found that levels of the glycosphingolipid, GM1 ganglioside, were ~60% lower in CFTR-silenced cells than in controls. CFTR-silenced cells exhibited reduced levels of activated β1-integrin, phosphorylated tyrosine 576 of focal adhesion kinase (pFAK, and phosphorylation of Crk-associated substrate (pCAS. Addition of GM1 (but not GM3 ganglioside to CFTR-silenced cells restored activated β1-integrin, pFAK, and pCAS to near control levels and partially restored (~40% cell migration. Our results suggest that decreased GM1 in CFTR-silenced cells depresses β1-integrin signaling, which contributes to the delayed wound repair observed in these cells. These findings have implications for the pathology of cystic fibrosis, where altered sphingolipid levels in airway epithelial cells could result in a diminished capacity for wound repair after injury.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Cell Physiology
Volume306
Issue number9
DOIs
StatePublished - May 1 2014

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G(M1) Ganglioside
Integrins
Wounds and Injuries
Sphingolipids
Glycosphingolipids
Epithelial Cells
Small Interfering RNA
Cell Movement
Crk-Associated Substrate Protein
G(M3) Ganglioside
Cell Line
Focal Adhesion Protein-Tyrosine Kinases
Cystic Fibrosis Transmembrane Conductance Regulator
Electric Impedance
Immunoblotting
Cystic Fibrosis
Tyrosine
Chlorides
Pneumonia
Sodium

Keywords

  • Cell attachment
  • Chloride channels
  • Pulmonary epithelial cells
  • Sialo-lipids

ASJC Scopus subject areas

  • Physiology
  • Cell Biology

Cite this

Reduced GM1 ganglioside in CFTR-deficient human airway cells results in decreased β1-integrin signaling and delayed wound repair. / Itokazu, Yutaka; Pagano, Richard E.; Schroeder, Andreas S.; O'Grady, Scott M.; Limper, Andrew H.; Marks, David L.

In: American Journal of Physiology - Cell Physiology, Vol. 306, No. 9, 01.05.2014.

Research output: Contribution to journalArticle

Itokazu, Yutaka ; Pagano, Richard E. ; Schroeder, Andreas S. ; O'Grady, Scott M. ; Limper, Andrew H. ; Marks, David L. / Reduced GM1 ganglioside in CFTR-deficient human airway cells results in decreased β1-integrin signaling and delayed wound repair. In: American Journal of Physiology - Cell Physiology. 2014 ; Vol. 306, No. 9.
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AU - Marks, David L.

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