title: COX-2 and cytosolic PLA2 mediate IL-1beta-induced cAMP production in human vascular smooth muscle cells.
abstract: Interleukin (IL)-1 is a potent vasodilator that causes prolonged induction of prostacyclin (PGI2) and cAMP synthesis in human vascular smooth muscle cells (HVSMC). The present study investigated IL-1 induction of PG synthetic enzymes in HVSMC and tested their respective roles in PGI2 and cAMP production. Cyclooxygenase (COX)-1 mRNA was not detectable in either control or IL-1-treated HVSMC, as assessed by RT-PCR. In contrast, COX-2 mRNA was detectable in control HVSMC, increased markedly (16-fold) after 1 h of IL-1 exposure, and increased further (52-fold) after 24 h. COX-2 protein levels, assessed by Western analysis, were increased concomitantly. HVSMC contained mRNA encoding both the secreted and cytosolic forms of phospholipase A2 (sPLA2 and cPLA2, respectively). IL-1 stimulation did not affect sPLA2 mRNA levels, but cPLA2 mRNA levels increased at 8 h, after the initial induction of PG synthesis. HVSMC constitutively expressed PGI2 synthase mRNA, and its levels were not affected by IL-1. A selective COX-2 inhibitor, NS-398, reversed IL-1-induced PGI2 and cAMP production, supporting a role of COX-2 in mediating increased PG synthesis. IL-1-induced cAMP was also reversed by a selective cPLA2 inhibitor, AACOCF3, but not by thioetheramide phosphorylcholine, which inhibits sPLA2 preferentially over cPLA2, supporting a requirement for cPLA2-derived arachidonic acid in IL-1-induced PG synthesis. The delayed induction of cPLA2 mRNA was also attenuated by NS-398, suggesting that it was secondary to the initial COX-2-induced PG synthesis. Together, the results support the hypothesis that IL-1 induces intracellular PG synthesis in HVSMC via rapid upregulation of COX-2, which utilizes cPLA2-derived arachidonic acid to generate PG metabolites that regulate adenylate cyclase.
authors: Department of Medicine and Tupper Research Institute, New England Medical Center Hospitals, Tufts University School of Medicine, Boston, Massachusetts 02111, USA. debbie.beasley@es.nemc.org
authors: D
authors: D
authors: Beasley
mesh_terms: D002478:Cells, Cultured; D000242:Cyclic AMP / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D051546:Cyclooxygenase 2; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D003600:Cytosol / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology*; D019746:Intramolecular Oxidoreductases / Q000378:metabolism; D007527:Isoenzymes / Q000235:genetics / Q000502:physiology*; D008565:Membrane Proteins; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D009578:Nitrobenzenes / Q000494:pharmacology; D010741:Phospholipases A / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D054467:Phospholipases A2; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics / Q000502:physiology*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology; D013997:Time Factors
chemical_list: D004791:Enzyme Inhibitors; D007375:Interleukin-1; D007527:Isoenzymes; D008565:Membrane Proteins; D009578:Nitrobenzenes; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; D013449:Sulfonamides; C080955:N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide; D003577:Cytochrome P-450 Enzyme System; D000242:Cyclic AMP; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; D010741:Phospholipases A; D054467:Phospholipases A2; D019746:Intramolecular Oxidoreductases; C022461:prostacyclin synthetase