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CPI-17 silencing-reduced responsiveness in control and TNF-alpha-treated human bronchi

Caroline Morin1, Marco Sirois, Vincent Echave

  • 1Le Bilarium, Department of Physiology and Biophysics, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Sherbrooke, Quebec, Canada.

American Journal of Respiratory Cell and Molecular Biology
|September 2, 2008
PubMed

Insights

Protein kinase C-dependent phosphorylation of CPI-17 regulates airway smooth muscle function. Inhibiting CPI-17 in human bronchi reduced airway hyperresponsiveness (AHR) and improved relaxation responses.

Area of Science:

  • Pulmonary Medicine
  • Molecular Biology
  • Pharmacology

Background:

  • Airway hyperresponsiveness (AHR) involves altered bronchial smooth muscle (BSM) function.
  • Protein kinase C (PKC)-dependent phosphorylation of CPI-17 modulates BSM Ca2+ sensitivity and reactivity.
  • CPI-17 is a key regulator in pathophysiologic conditions affecting BSM.

Purpose of the Study:

  • To investigate the role of CPI-17 in AHR.
  • To assess the therapeutic potential of targeting CPI-17 in human bronchi.
  • To evaluate the effect of CPI-17 inhibition on BSM reactivity and relaxation.

Main Methods:

  • Utilized control and TNF-alpha-treated human bronchi.
  • Employed a siRNA duplex targeting human CPI-17 transcripts via X-TremeGene (X-TG) transfection.
  • Assessed mRNA and protein expression, Ca2+ sensitivity, and muscle reactivity to methacholine and beta2-agonist.
  • Validated transfection strategy using reversible permeabilization.
  • Corroborated findings with Western blot analysis and 14,15-EET treatment.

Main Results:

  • X-TG-siRNA targeting CPI-17 reduced CPI-17 mRNA and protein expression in human bronchi.
  • Transfection with CPI-17 siRNA significantly reduced BSM Ca2+ sensitivity.
  • Overall bronchial smooth muscle reactivity to methacholine decreased, while relaxation to beta2-agonist increased.
  • Western blot confirmed decreased CPI-17 and myosin light chain (MLC) phosphorylation.
  • Results mirrored those obtained with the anti-inflammatory eicosanoid 14,15-EET.

Conclusions:

  • CPI-17 plays a critical molecular role in airway hyperresponsiveness (AHR).
  • Targeting CPI-17 effectively reduces AHR in human bronchi without inducing bronchial wall remodeling.
  • Inhibition of CPI-17 phosphorylation presents a potential therapeutic strategy for AHR.

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