title: Cyclooxygenase-2 induction by bradykinin in human pulmonary artery smooth muscle cells is mediated by the cyclic AMP response element through a novel autocrine loop involving endogenous prostaglandin E2, E-prostanoid 2 (EP2), and EP4 receptors.
abstract: Bradykinin (BK) is an important mediator in several inflammatory and vascular diseases that acts in part via induction of cyclooxygenase-2 (COX-2). The mechanisms involved in BK-mediated COX-2 induction are unclear. Here we characterized the transcriptional mechanisms involved in human pulmonary artery smooth muscle cells. BK stimulated the activity of a transiently transfected 966-bp (-917 to + 49) COX-2 promoter luciferase reporter construct. There was no reduction in BK-induced luciferase activity in cells transfected with COX-2 promoter constructs of 674, 407, 239, and 135 bp or constructs with mutated CCAAT/enhancer-binding protein- or NF-kappaB-binding sites. In contrast luciferase activity was reduced in cells transfected with a 407-bp COX-2 promoter fragment containing a mutated cAMP response element (CRE)-binding site, suggesting that the CRE binding site is critical. Electrophoretic mobility shift assays using oligonucleotides specific for the CRE-binding region of the COX-2 promoter and consensus oligonucleotides showed strong specific binding. Furthermore BK increased consensus cAMP-responsive luciferase reporter (p6CRE/luc)-mediated luciferase expression. CRE activation occurred by BK inducing cytosolic phospholipase A2-mediated arachidonic acid release and rapid prostaglandin E2 (PGE2) production, thereby increasing cAMP. Indomethacin inhibited BK-induced PGE2 production, cAMP accumulation, and CRE/luc reporter and COX-2 promoter luciferase activity. Exogenous PGE2 and EP2 (ONO-AE1 259) and EP4 (ONO-AE1 329) PGE2 receptor agonists mimicked the effect of BK. Collectively these studies indicate that COX-2 induction by BK in human pulmonary artery smooth muscle cells is mediated by the CRE through a novel autocrine loop involving endogenous PGE2.
mesh_terms: D016718:Arachidonic Acid / Q000378:metabolism; D015153:Blotting, Western; D001920:Bradykinin / Q000737:chemistry*; D002455:Cell Division; D002470:Cell Survival; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism*; D051546:Cyclooxygenase 2; D004252:DNA Mutational Analysis; D004305:Dose-Response Relationship, Drug; D017353:Gene Deletion; D017930:Genes, Reporter; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D008156:Luciferases / Q000378:metabolism; D008565:Membrane Proteins; D008954:Models, Biological; D009130:Muscle, Smooth / Q000166:cytology*; D011401:Promoter Regions, Genetic; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism*; D011651:Pulmonary Artery / Q000166:cytology*; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D018078:Receptors, Prostaglandin E / Q000737:chemistry*; D058307:Receptors, Prostaglandin E, EP2 Subtype; D058309:Receptors, Prostaglandin E, EP4 Subtype; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014158:Transcription, Genetic; D014162:Transfection
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D007527:Isoenzymes; D008565:Membrane Proteins; C545644:PTGER2 protein, human; C545650:PTGER4 protein, human; D012333:RNA, Messenger; D018078:Receptors, Prostaglandin E; D058307:Receptors, Prostaglandin E, EP2 Subtype; D058309:Receptors, Prostaglandin E, EP4 Subtype; D016718:Arachidonic Acid; D012313:RNA; D000242:Cyclic AMP; D008156:Luciferases; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; D001920:Bradykinin