TY - JOUR
T1 - PKC-δ/c-Src-mediated EGF receptor transactivation regulates thrombin-induced COX-2 expression and PGE2 production in rat vascular smooth muscle cells
AU - Hsieh, Hsi Lung
AU - Sun, Chi Chin
AU - Wang, Tze Shyuan
AU - Yang, Chuen Mao
PY - 2008/9
Y1 - 2008/9
N2 - The thrombin/proteinase-activated receptors (PARs) have been shown to regulate smooth muscle cell proliferation, migration, and vascular maturation. Thrombin up-regulates expression of several proteins including cyclooxygenase (COX)-2 in vascular smooth muscle cells (VSMCs) and contributes to vascular diseases. However, the mechanisms underlying thrombin-regulated COX-2 expression in VSMCs remain unclear. Western blotting, RT-PCR, and EIA kit analyses showed that thrombin induced the expression of COX-2 mRNA and protein and PGE2 release in a time-dependent manner, which was attenuated by inhibitors of PKC (GF109203X and rottlerin), c-Src (PP1), EGF receptor (EGFR; AG1478) and MEK1/2 (U0126), or transfection with dominant negative mutants of PKC-δ, c-Src or extracellular regulated kinase (ERK) and ERK1 short hairpin RNA interference (shRNA). These results suggest that transactivation of EGFR participates in COX-2 expression induced by thrombin in VSMCs. Accordingly, thrombin stimulated phosphorylation of ERK1/2 which was attenuated by GF109203X, rottlerin, PP1, GM6001, CRM197, AG1478, or U0126, respectively. Furthermore, this up-regulation of COX-2 mRNA and protein was blocked by selective inhibitors of AP-1 and NF-κB, curcumin and helenalin, respectively. Moreover, thrombin-stimulated activation of NF-κB, AP-1, and COX-2 promoter activity was blocked by the inhibitors of c-Src, PKC, EGFR, MEK1/2, AP-1 and NF-κB, suggesting that thrombin induces COX-2 promoter activity mediated through PKC(δ)/c-Src-dependent EGFR transactivation, MEK-ERK1/2, AP-1, and NF-κB. These results demonstrate that in VSMCs, activation of ERK1/2, AP-1 and NF-κB pathways was essential for thrombin-induced COX-2 gene expression. Understanding the regulation of COX-2 expression and PGE2 release by thrombin/PARs system on VSMCs may provide potential therapeutic targets of vascular inflammatory disorders including arteriosclerosis.
AB - The thrombin/proteinase-activated receptors (PARs) have been shown to regulate smooth muscle cell proliferation, migration, and vascular maturation. Thrombin up-regulates expression of several proteins including cyclooxygenase (COX)-2 in vascular smooth muscle cells (VSMCs) and contributes to vascular diseases. However, the mechanisms underlying thrombin-regulated COX-2 expression in VSMCs remain unclear. Western blotting, RT-PCR, and EIA kit analyses showed that thrombin induced the expression of COX-2 mRNA and protein and PGE2 release in a time-dependent manner, which was attenuated by inhibitors of PKC (GF109203X and rottlerin), c-Src (PP1), EGF receptor (EGFR; AG1478) and MEK1/2 (U0126), or transfection with dominant negative mutants of PKC-δ, c-Src or extracellular regulated kinase (ERK) and ERK1 short hairpin RNA interference (shRNA). These results suggest that transactivation of EGFR participates in COX-2 expression induced by thrombin in VSMCs. Accordingly, thrombin stimulated phosphorylation of ERK1/2 which was attenuated by GF109203X, rottlerin, PP1, GM6001, CRM197, AG1478, or U0126, respectively. Furthermore, this up-regulation of COX-2 mRNA and protein was blocked by selective inhibitors of AP-1 and NF-κB, curcumin and helenalin, respectively. Moreover, thrombin-stimulated activation of NF-κB, AP-1, and COX-2 promoter activity was blocked by the inhibitors of c-Src, PKC, EGFR, MEK1/2, AP-1 and NF-κB, suggesting that thrombin induces COX-2 promoter activity mediated through PKC(δ)/c-Src-dependent EGFR transactivation, MEK-ERK1/2, AP-1, and NF-κB. These results demonstrate that in VSMCs, activation of ERK1/2, AP-1 and NF-κB pathways was essential for thrombin-induced COX-2 gene expression. Understanding the regulation of COX-2 expression and PGE2 release by thrombin/PARs system on VSMCs may provide potential therapeutic targets of vascular inflammatory disorders including arteriosclerosis.
KW - COX-2
KW - EGF receptor
KW - ERK1/2
KW - Thrombin
KW - Vascular smooth muscle cell
UR - http://www.scopus.com/inward/record.url?scp=49149104318&partnerID=8YFLogxK
U2 - 10.1016/j.bbamcr.2008.03.016
DO - 10.1016/j.bbamcr.2008.03.016
M3 - 文章
C2 - 18452714
AN - SCOPUS:49149104318
SN - 0167-4889
VL - 1783
SP - 1563
EP - 1575
JO - Biochimica et Biophysica Acta - Molecular Cell Research
JF - Biochimica et Biophysica Acta - Molecular Cell Research
IS - 9
ER -