Mechanisms of thrombin-induced MAPK activation associated with cell proliferation in human cultured tracheal smooth muscle cells

Chih Chung Lin, Ming Hwang Shyr, Chin Sung Chien, Chuan Chwan Wang, Chi Tso Chiu, Li Der Hsiao, Chuen Mao Yang*

*Corresponding author for this work

Research output: Contribution to journalJournal Article peer-review

31 Scopus citations

Abstract

The elevated level of thrombin has been detected in the airway fluids of asthmatic patients. However, the implication of thrombin in the pathogenesis of bronchial hyperreactivity was not completely understood. Therefore, in this study we investigated the effect of thrombin on cell proliferation and p42/p44 mitogen-activated protein kinase (MAPK) activation in human tracheal smooth muscle cells (TSMCs). Thrombin stimulated [3H]thymidine incorporation and p42/p44 MAPK phosphorylation in a time- and concentration-dependent manner in TSMCs. Pretreatment of TSMCs with pertussis toxin (PTX) significantly inhibited [3H]thymidine incorporation and phosphorylation of MAPK induced by thrombin. These responses were attenuated by tyrosine kinase inhibitors genistein and herbimycin A, phosphatidyl inositide (PI)-phospholipase C (PLC) inhibitor U73122, protein kinase C (PKC) inhibitor GF109203X, removal of Ca2+ by addition of BAPTA/AM plus EGTA, and PI 3-kinase inhibitors wortmannin and LY294002. In addition, thrombin-induced [3H]-thymidine incorporation and p42/p44 MAPK phosphorylation was completely inhibited by PD98059 (an inhibitor of MEK1/2), indicating that activation of MEK1/2 was required for these responses. Furthermore, overexpression of dominant negative mutants, RasN17 and Raf-301, significantly suppressed p42/p44 MAPK activation induced by thrombin and PDGF-BB, indicating that Ras and Raf may be required for activation of these kinases. These results conclude that the mitogenic effect of thrombin was mediated through the activation of Ras/Raf/MEK/MAPK pathway. Thrombin-mediated MAPK activation was modulated by PI-PLC, Ca2+, PKC, tyrosine kinase, and PI 3-kinase associated with cell proliferation in cultured human TSMCs.

Original languageEnglish
Pages (from-to)257-267
Number of pages11
JournalCellular Signalling
Volume13
Issue number4
DOIs
StatePublished - 2001

Keywords

  • Cell proliferation
  • MEK
  • Mitogen-activated protein kinase
  • Protein kinase C
  • Thrombin
  • Tracheal smooth muscle cell
  • Tyrosine kinase

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