Functional coupling expression of COX-2 and cPLA2 induced by ATP in rat vascular smooth muscle cells: role of ERK1/2, p38 MAPK, and NF-kappaB.
OBJECTIVE
Vascular smooth muscle cells (VSMCs) that function as synthetic units play important roles in inflammatory diseases such as atherosclerosis and angiogenesis. As extracellular nucleotides such as ATP have been shown to act via activation of P(2) purinoceptors implicated in various inflammatory diseases, we hypothesized that extracellular nucleotides contribute to vascular diseases via upregulated expression of inflammatory proteins, such as cyclooxygenase (COX-2) and cytosolic phospholipase A2 (cPLA2) in VSMCs.


RESULTS
Western blotting, promoter assay, RT-PCR, and PGE2 immunoassay revealed that ATPgammaS induced expression of COX-2 and prostaglandin (PGE2) synthesis through the activation of p42/p44 MAPK (mitogen-activated protein kinase), p38 MAPK, and nuclear factor-kappaB (NF-kappaB). These responses were attenuated by inhibitors of MAPK/ERK kinase (MEK1/2; U0126), p38 MAPK (SB202190), and NF-kappaB (helenalin), or by tranfection with dominant negative mutants of p42, p38, IkappaB kinase (IKK)alpha, and IKKbeta. Furthermore, the ATPgammaS-stimulated translocation of NF-kappaB into the nucleus and degradation of IkappaBalpha was blocked by U0126 and helenalin. In addition, the ATPgammaS-stimulated cPLA2 expression was inhibited by U0126, SB202190, helenalin, celecoxib (a selective COX-2 inhibitor), and PGE2 receptor antagonists (AH6809, GW627368X, and SC-19220). However, the inhibitory effect of celecoxib on cPLA2 expression was reversed by addition of exogenous PGE2.


CONCLUSIONS
Our results suggest that in VSMCs, activation of p42/p44 MAPK, p38 MAPK, and NF-kappaB is essential for ATPgammaS-induced COX-2 expression and PGE2 synthesis. Newly synthesized PGE2 was observed to act as an autocrine signal contributing to cPLA2 expression, which may be implicated in inflammatory responses. Collectively, our findings provide insights into the correlation between COX-2 and cPLA2 expression in ATPgammaS-stimulated VSMCs with similar molecular mechanisms and functional coupling to amplify the occurrence of vessel disease-related vascular inflammation.
Lin
Chih-Chung
CC
Department of Anesthetics, Chang Gung University, Tao-Yuan, Taiwan, Republic of China.
Lin
Wei-Ning
WN
Wang
Wei-Jung
WJ
Sun
Chi-Chin
CC
Tung
Wei-Hsuan
WH
Wang
Hui-Hsin
HH
Yang
Chuen-Mao
CM
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism*; D000818:Animals; D019898:Autocrine Communication; D002478:Cells, Cultured; D051546:Cyclooxygenase 2 / Q000378:metabolism*; D015232:Dinoprostone / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000201:enzymology*; D032389:Myocytes, Smooth Muscle / Q000201:enzymology*; D016328:NF-kappa B / Q000378:metabolism; D054510:Phospholipases A2, Cytosolic / Q000378:metabolism*; D010766:Phosphorylation; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D016328:NF-kappa B; D012333:RNA, Messenger; C022571:adenosine 5'-O-(3-thiotriphosphate); D000255:Adenosine Triphosphate; D051546:Cyclooxygenase 2; C489691:Ptgs2 protein, rat; D048049:Extracellular Signal-Regulated MAP Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D054510:Phospholipases A2, Cytosolic; D015232:Dinoprostone