Human lung myofibroblast TGFβ1-dependent Smad2/3 signalling is Ca2+-dependent and regulated by KCa3.1 K+ channels

Katy M. Roach, Carol Feghali-Bostwick, Heike Wulff, Yassine Amrani, Peter Bradding

Research output: Contribution to journalArticle

27 Scopus citations

Abstract

Background: Idiopathic pulmonary fibrosis (IPF) is a common and invariably lethal interstitial lung disease with poorly effective therapy. Blockade of the K+ channel KCa3.1 reduces constitutive α-SMA and Smad2/3 nuclear translocation in IPF-derived human lung myofibroblasts (HLMFs), and inhibits several transforming growth factor beta 1 (TGFβ1)-dependent cell processes. We hypothesized that KCa3.1-dependent cell processes also regulate the TGFβ1-dependent Smad2/3 signalling pathway in HLMFs. HLMFs obtained from non-fibrotic controls (NFC) and IPF lungs were grown in vitro and examined for αSMA expression by immunofluorescence, RT-PCR, and flow cytometry. Two specific and distinct KCa3.1 blockers (TRAM-34 200 nM and ICA-17043 [Senicapoc] 100 nM) were used to determine their effects on TGFβ1-dependent signalling. Expression of phosphorylated and total Smad2/3 following TGFβ1 stimulation was determined by Western blot and Smad2/3 nuclear translocation by immunofluorescence. Results: KCa3.1 block attenuated TGFβ1-dependent Smad2/3 phosphorylation and nuclear translocation, and this was mimicked by lowering the extracellular Ca2+ concentration. KCa3.1 block also inhibited Smad2/3-dependent gene transcription (αSMA, collagen type I), inhibited KCa3.1 mRNA expression, and attenuated TGFβ1-dependent αSMA protein expression. Conclusions: KCa3.1 activity regulates TGFβ1-dependent effects in NFC- and IPF-derived primary HLMFs through the regulation of the TGFβ1/Smad signalling pathway, with promotion of downstream gene transcription and protein expression. KCa3.1 blockers may offer a novel approach to treating IPF.

Original languageEnglish (US)
Article number5
JournalFibrogenesis and Tissue Repair
Volume8
Issue number1
DOIs
StatePublished - Mar 26 2015

Keywords

  • Human lung myofibroblast
  • Idiopathic pulmonary fibrosis
  • Potassium channel KCa3.1

ASJC Scopus subject areas

  • Medicine (miscellaneous)
  • Dermatology
  • Gastroenterology
  • Hepatology
  • Rheumatology

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