title: Endothelial 12(S)-HETE vasorelaxation is mediated by thromboxane receptor inhibition in mouse mesenteric arteries.
abstract: Arachidonic acid (AA) metabolites mediate endothelium-dependent relaxation in many vascular beds. Previously, we identified the major AA 12/15-lipoxygenase (12/15-LO) metabolite of mouse arteries as 12-hydroxyeicosatetraenoic acid (12-HETE). The goal was to determine the stereospecific configuration of mouse vascular 12-HETE and characterize the role of 12-HETE stereoisomers in the regulation of vascular tone. Using normal, reverse phase, and chiral HPLC, the stereospecific configuration was identified as 12(S)-HETE. 12(S)-HETE relaxed U46619-, carbocyclic thromboxane A(2)-, PGF(2α)-, and 8-iso PGF(2α)-preconstricted mesenteric arteries, but not phenylephrine-preconstricted arteries. 12(R)-HETE was more potent than 12(S)-HETE in relaxing U46619-preconstricted mouse arteries (maximum relaxations = 91.4 ± 2.7% and 71.8 ± 5.9%, respectively). Neither 12-HETE isomer caused constriction. Pretreatment with 12(S)- or 12(R)-HETE (1 μM) inhibited constrictions to U46619 but not phenylephrine. To investigate the role of thromboxane A(2) (TP) receptors in 12-HETE vascular actions, [(3)H]SQ29548 radioligand binding studies were performed in mouse platelets. U46619, 12(R)-HETE, and 12(S)-HETE displaced [(3)H]SQ29548 binding with IC(50)s of 0.07, 0.32, and 1.73 μM, respectively. Both 12(S)- and 12(R)-HETE inhibited intracellular calcium increases induced by U46619 (10 nM) in HEK293 cells overexpressing TP(α) receptor (65.5% and 45.1%, respectively) and coexpressing prostacyclin (IP) and TP(α) receptors (58.0% and 27.1%, respectively). The LO inhibitor NDGA (10 μM) reduced AA relaxations in arteries preconstricted with U46619 but not phenylephrine. These results indicate that exogenous and endogenous 12(S)-HETE relax mouse mesenteric arteries that are preconstricted with thromboxane agonists. These 12(S)-HETE relaxations are mediated by TP receptor competitive inhibition and inhibition of TP agonist-induced increases in intracellular calcium.
authors.affiliation: Department of Pharmacology and Toxicology, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA.
authors.forename: Lawan
authors.initials: L
authors.lastname: Siangjong
authors.forename: Kathryn M
authors.initials: KM
authors.lastname: Gauthier
authors.forename: Sandra L
authors.initials: SL
authors.lastname: Pfister
authors.forename: Emer M
authors.initials: EM
authors.lastname: Smyth
authors.forename: William B
authors.initials: WB
authors.lastname: Campbell
mesh_terms: D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid / Q000494:pharmacology*; D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid / Q000494:pharmacology; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D001792:Blood Platelets / Q000187:drug effects; D015153:Blotting, Western; D019086:Bridged Bicyclo Compounds, Heterocyclic; D020013:Calcium Signaling / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D004730:Endothelium, Vascular / Q000187:drug effects*; D005231:Fatty Acids, Unsaturated; D057809:HEK293 Cells; D006801:Humans; D006834:Hydrazines / Q000494:pharmacology; D007537:Isometric Contraction / Q000187:drug effects; D008297:Male; D008638:Mesenteric Arteries / Q000187:drug effects*; D051379:Mice; D008810:Mice, Inbred C57BL; D043562:Receptors, G-Protein-Coupled / Q000187:drug effects; D018102:Receptors, Leukotriene B4 / Q000187:drug effects; D017482:Receptors, Thromboxane / Q000037:antagonists & inhibitors*; D044244:Receptors, Thromboxane A2, Prostaglandin H2 / Q000187:drug effects; D014662:Vasoconstrictor Agents / Q000494:pharmacology; D014665:Vasodilator Agents / Q000494:pharmacology*
chemical_list: D019086:Bridged Bicyclo Compounds, Heterocyclic; D005231:Fatty Acids, Unsaturated; C501954:Gpr31 protein, mouse; D006834:Hydrazines; D043562:Receptors, G-Protein-Coupled; D018102:Receptors, Leukotriene B4; D017482:Receptors, Thromboxane; D044244:Receptors, Thromboxane A2, Prostaglandin H2; D014662:Vasoconstrictor Agents; D014665:Vasodilator Agents; D016718:Arachidonic Acid; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; C045749:SQ 29548