Abstract

In addition to its functions in thrombosis and hemostasis, thrombin also plays an important role in lung inflammation. Our previous report showed that thrombin activates the protein kinase C (PKC)α/c-Src and Gβγ/Rac1/PI3K/Akt signaling pathways to induce IκB kinase α/β (IKKα/β) activation, NF-κB transactivation, and IL-8/CXCL8 expressions in human lung epithelial cells (ECs). In this study, we further investigated the mechanism of c-Src-dependent Shc, Raf-1, and extracellular signal-regulated kinase (ERK) signaling pathways involved in thrombin-induced NF-κB activation and IL-8/CXCL8 release. Thrombin-induced increases in IL-8/CXCL8 release and κB-luciferase activity were inhibited by the Shc small interfering RNA (siRNA), p66Shc siRNA, GW 5074 (a Raf-1 inhibitor), and PD98059 (a mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitor). Treatment of A549 cells with thrombin increased p66Shc and p46/p52Shc phosphorylation at Tyr239/240 and Tyr317, which was inhibited by cell transfection with the dominant negative mutant of c-Src (c-Src DN). Thrombin caused time-dependent phosphorylation of Raf-1 and ERK, which was attenuated by the c-Src DN. Thrombin-induced IKKα/β phosphorylation was inhibited by GW 5074 and PD98059. Treatment of cells with thrombin induced Gβγ, c-Src, and p66Shc complex formation in a time-dependent manner. Taken together, these results show for the first time that thrombin activates Shc, Raf-1, and ERK through Gβγ, c-Src, and Shc complex formation to induce IKKα/β phosphorylation, NF-κB activation, and IL-8/CXCL8 release in human lung ECs.

Original languageEnglish
Pages (from-to)1166-1175
Number of pages10
JournalCellular Signalling
Volume25
Issue number5
DOIs
Publication statusPublished - May 2013

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Mitogen-Activated Protein Kinase 3
Interleukin-8
Thrombin
Epithelial Cells
Lung
Phosphorylation
Thrombin Time
Mitogen-Activated Protein Kinase Kinases
Small Interfering RNA
Hemostasis
Luciferases
Phosphatidylinositol 3-Kinases
Protein Kinase C
Transcriptional Activation
Transfection
Pneumonia
Thrombosis
Phosphotransferases

Keywords

  • Epithelial cells
  • IL-8/CXCL8
  • Lung
  • NF-κB
  • Shc
  • Thrombin

ASJC Scopus subject areas

  • Cell Biology

Cite this

@article{2cd5067d701e4c1caa2a6e1bec36764e,
title = "Thrombin-induced NF-κB activation and IL-8/CXCL8 release is mediated by c-Src-dependent Shc, Raf-1, and ERK pathways in lung epithelial cells",
abstract = "In addition to its functions in thrombosis and hemostasis, thrombin also plays an important role in lung inflammation. Our previous report showed that thrombin activates the protein kinase C (PKC)α/c-Src and Gβγ/Rac1/PI3K/Akt signaling pathways to induce IκB kinase α/β (IKKα/β) activation, NF-κB transactivation, and IL-8/CXCL8 expressions in human lung epithelial cells (ECs). In this study, we further investigated the mechanism of c-Src-dependent Shc, Raf-1, and extracellular signal-regulated kinase (ERK) signaling pathways involved in thrombin-induced NF-κB activation and IL-8/CXCL8 release. Thrombin-induced increases in IL-8/CXCL8 release and κB-luciferase activity were inhibited by the Shc small interfering RNA (siRNA), p66Shc siRNA, GW 5074 (a Raf-1 inhibitor), and PD98059 (a mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitor). Treatment of A549 cells with thrombin increased p66Shc and p46/p52Shc phosphorylation at Tyr239/240 and Tyr317, which was inhibited by cell transfection with the dominant negative mutant of c-Src (c-Src DN). Thrombin caused time-dependent phosphorylation of Raf-1 and ERK, which was attenuated by the c-Src DN. Thrombin-induced IKKα/β phosphorylation was inhibited by GW 5074 and PD98059. Treatment of cells with thrombin induced Gβγ, c-Src, and p66Shc complex formation in a time-dependent manner. Taken together, these results show for the first time that thrombin activates Shc, Raf-1, and ERK through Gβγ, c-Src, and Shc complex formation to induce IKKα/β phosphorylation, NF-κB activation, and IL-8/CXCL8 release in human lung ECs.",
keywords = "Epithelial cells, IL-8/CXCL8, Lung, NF-κB, Shc, Thrombin",
author = "Lin, {Chien Huang} and Yu, {Ming Chih} and Chiang, {Chia Chieh} and Bien, {Mauo Ying} and Chien, {Ming Hsien} and Chen, {Bing Chang}",
year = "2013",
month = "5",
doi = "10.1016/j.cellsig.2013.01.018",
language = "English",
volume = "25",
pages = "1166--1175",
journal = "Cellular Signalling",
issn = "0898-6568",
publisher = "Elsevier Inc.",
number = "5",

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TY - JOUR

T1 - Thrombin-induced NF-κB activation and IL-8/CXCL8 release is mediated by c-Src-dependent Shc, Raf-1, and ERK pathways in lung epithelial cells

AU - Lin, Chien Huang

AU - Yu, Ming Chih

AU - Chiang, Chia Chieh

AU - Bien, Mauo Ying

AU - Chien, Ming Hsien

AU - Chen, Bing Chang

PY - 2013/5

Y1 - 2013/5

N2 - In addition to its functions in thrombosis and hemostasis, thrombin also plays an important role in lung inflammation. Our previous report showed that thrombin activates the protein kinase C (PKC)α/c-Src and Gβγ/Rac1/PI3K/Akt signaling pathways to induce IκB kinase α/β (IKKα/β) activation, NF-κB transactivation, and IL-8/CXCL8 expressions in human lung epithelial cells (ECs). In this study, we further investigated the mechanism of c-Src-dependent Shc, Raf-1, and extracellular signal-regulated kinase (ERK) signaling pathways involved in thrombin-induced NF-κB activation and IL-8/CXCL8 release. Thrombin-induced increases in IL-8/CXCL8 release and κB-luciferase activity were inhibited by the Shc small interfering RNA (siRNA), p66Shc siRNA, GW 5074 (a Raf-1 inhibitor), and PD98059 (a mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitor). Treatment of A549 cells with thrombin increased p66Shc and p46/p52Shc phosphorylation at Tyr239/240 and Tyr317, which was inhibited by cell transfection with the dominant negative mutant of c-Src (c-Src DN). Thrombin caused time-dependent phosphorylation of Raf-1 and ERK, which was attenuated by the c-Src DN. Thrombin-induced IKKα/β phosphorylation was inhibited by GW 5074 and PD98059. Treatment of cells with thrombin induced Gβγ, c-Src, and p66Shc complex formation in a time-dependent manner. Taken together, these results show for the first time that thrombin activates Shc, Raf-1, and ERK through Gβγ, c-Src, and Shc complex formation to induce IKKα/β phosphorylation, NF-κB activation, and IL-8/CXCL8 release in human lung ECs.

AB - In addition to its functions in thrombosis and hemostasis, thrombin also plays an important role in lung inflammation. Our previous report showed that thrombin activates the protein kinase C (PKC)α/c-Src and Gβγ/Rac1/PI3K/Akt signaling pathways to induce IκB kinase α/β (IKKα/β) activation, NF-κB transactivation, and IL-8/CXCL8 expressions in human lung epithelial cells (ECs). In this study, we further investigated the mechanism of c-Src-dependent Shc, Raf-1, and extracellular signal-regulated kinase (ERK) signaling pathways involved in thrombin-induced NF-κB activation and IL-8/CXCL8 release. Thrombin-induced increases in IL-8/CXCL8 release and κB-luciferase activity were inhibited by the Shc small interfering RNA (siRNA), p66Shc siRNA, GW 5074 (a Raf-1 inhibitor), and PD98059 (a mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitor). Treatment of A549 cells with thrombin increased p66Shc and p46/p52Shc phosphorylation at Tyr239/240 and Tyr317, which was inhibited by cell transfection with the dominant negative mutant of c-Src (c-Src DN). Thrombin caused time-dependent phosphorylation of Raf-1 and ERK, which was attenuated by the c-Src DN. Thrombin-induced IKKα/β phosphorylation was inhibited by GW 5074 and PD98059. Treatment of cells with thrombin induced Gβγ, c-Src, and p66Shc complex formation in a time-dependent manner. Taken together, these results show for the first time that thrombin activates Shc, Raf-1, and ERK through Gβγ, c-Src, and Shc complex formation to induce IKKα/β phosphorylation, NF-κB activation, and IL-8/CXCL8 release in human lung ECs.

KW - Epithelial cells

KW - IL-8/CXCL8

KW - Lung

KW - NF-κB

KW - Shc

KW - Thrombin

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U2 - 10.1016/j.cellsig.2013.01.018

DO - 10.1016/j.cellsig.2013.01.018

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VL - 25

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