Stretch-induced mitogen-activated protein kinase activation in lung fibroblasts is independent of receptor tyrosine kinases

Francis Boudreault, Daniel J. Tschumperlin

Research output: Contribution to journalArticlepeer-review

19 Scopus citations

Abstract

Lung growth and remodeling are modulated by mechanical stress, with fibroblasts thought to play a leading role. Little mechanistic information is available about how lung fibroblasts respond to mechanical stress. We exposed cultured lung fibroblasts to tonic stretch and measured changes in phosphorylation status of mitogen-activated protein kinases (MAPKs), selected receptor tyrosine kinases (RTKs), and phospholipase Cγ1 (PLCγ1) and activation of the small G-protein Ras. Human lung fibroblasts (LFs) were seeded on matrix-coated silicone membranes and exposed to equibiaxial 10 to 40% static stretch or 20% contraction. LFs were stimulated with EGF, FGF2, or PDGF-BB or exposed to stretch in the presence of inhibitors of EGFR (AG1478), FGFR (PD173074), and PDGFR (AG1296). Phospho-MAPK, phospho-RTK, and phospho-PLCg1 levels were measured by Western blotting. Active GTP-Ras was quantified by immunoblotting after pulldown with a glutathione S-transferase-Raf-RBD construct. Normalized p-ERK1/2, p-JNK, and p-p38 levels increased after stretch but not contraction. Ligands to RTKs broadly stimulated MAPKs, with the responses to EGF and PDGF most similar to stretch in terms of magnitude and rank order of MAPK responses. Stretching cells failed to elicit measurable activation of EGFR, FGFR (FRS2α phosphorylation), or PDGFR. Potent inhibitors of the kinase activity of each receptor failed to attenuate stretch-induced MAPK activation. PLCg1 and Ras,prominent effectors downstream of RTKs, were not activated by stretch. Our findings demonstrate that MAPKs are potently activated by stretch in lung fibroblasts, but, in contrast to stress responses observed in other cell types, RTKs are not necessary for stretch-induced MAPK activation in LFs.

Original languageEnglish (US)
Pages (from-to)64-73
Number of pages10
JournalAmerican journal of respiratory cell and molecular biology
Volume43
Issue number1
DOIs
StatePublished - Jul 1 2010

Keywords

  • EGFR
  • FGFR
  • MAPK
  • Mechanotransduction
  • PDGFR

ASJC Scopus subject areas

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

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