Identification of 13-hydroxy-14,15-epoxyeicosatrienoic acid as an acid-stable endothelium-derived hyperpolarizing factor in rabbit arteries

Yuttana Chawengsub, Kathryn M. Gauthier, Kasem Nithipatikom, Bruce D. Hammock, J R Falck, Dubasi Narsimhaswamy, William B. Campbell

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Abstract

Arachidonic acid (AA) is metabolized by endothelial 15-lipoxygenase (15-LO) to several vasodilatory eicosanoids such as 11,12,15-trihydroxyeicosatrienoic acid (11,12,15-THETA) and its proposed unstable precursor 15-hydroxy-11,12-epoxyeicosatrienoic acid (15-H-11,12-EETA). In the present study, the acid-stable 13-hydroxy-trans-14,15-epoxy-eicosatrienoic acid (13-H-14,15-EETA) was identified and its vascular activities characterized. Rabbit aorta, mesenteric arteries, and the combination of 15-LO and cytochrome P450 2J2 converted AAto two distinct HEETA metabolites. The HEETA metabolites were resistant to acidic hydrolysis but were hydrolyzed by recombinant sEH to a more polar metabolite identified by mass spectrometry as 13,14,15-THETA. Mass spectrometric analyses and HPLC comigration identified the HEETAs as threo- and erythro-diastereomers of 13-H-trans-14,15-EETA. Erythro- and threo-diastereomers of 13-H-trans-14,15-EETA relaxed endothelium-denuded rabbit small mesenteric arteries with maximum relaxations of 22.6±6.0% and 8.6±4.3%, respectively. Apamin (10-7 M) inhibited the relaxations to the erythro-isomer(maximumrelaxation=1.2±5.6%) and increasing [K+]o from 4.6 to 30 mM blocked relaxations to both isomers. In cell-attached patches of mesenteric arterial smooth muscle cells (SMCs), erythro-13-H-trans-14,15-EETA (1-3 × 10-6 M) increased mean open time of small conductance K+ channels (13-14 pS) from 0.0007 ± 0.0007 to 0.0053 ± 0.0042. This activation was inhibited by apamin. The erythro, but not the threo, isomer blocked angiotensin II-stimulated aortic SMC migration. These studies demonstrate that 13-H-14,15-EETAs induces vascular relaxation via K+ channel activation to cause SMC hyperpolarization. Thus, 13-H-14,15-EETA represents a new endothelial factor.

Original languageEnglish (US)
Pages (from-to)31280-31290
Number of pages11
JournalJournal of Biological Chemistry
Volume284
Issue number45
DOIs
StatePublished - Nov 6 2009

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Metabolites
Arachidonate 15-Lipoxygenase
Isomers
Smooth Muscle Myocytes
Endothelium
Apamin
Muscle
Mesenteric Arteries
Arteries
Rabbits
Acids
Blood Vessels
Chemical activation
Eicosanoids
Arachidonic Acid
Angiotensin II
Cytochrome P-450 Enzyme System
Cell Movement
Mass spectrometry
Aorta

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Identification of 13-hydroxy-14,15-epoxyeicosatrienoic acid as an acid-stable endothelium-derived hyperpolarizing factor in rabbit arteries. / Chawengsub, Yuttana; Gauthier, Kathryn M.; Nithipatikom, Kasem; Hammock, Bruce D.; Falck, J R; Narsimhaswamy, Dubasi; Campbell, William B.

In: Journal of Biological Chemistry, Vol. 284, No. 45, 06.11.2009, p. 31280-31290.

Research output: Contribution to journalArticle

Chawengsub, Yuttana ; Gauthier, Kathryn M. ; Nithipatikom, Kasem ; Hammock, Bruce D. ; Falck, J R ; Narsimhaswamy, Dubasi ; Campbell, William B. / Identification of 13-hydroxy-14,15-epoxyeicosatrienoic acid as an acid-stable endothelium-derived hyperpolarizing factor in rabbit arteries. In: Journal of Biological Chemistry. 2009 ; Vol. 284, No. 45. pp. 31280-31290.
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abstract = "Arachidonic acid (AA) is metabolized by endothelial 15-lipoxygenase (15-LO) to several vasodilatory eicosanoids such as 11,12,15-trihydroxyeicosatrienoic acid (11,12,15-THETA) and its proposed unstable precursor 15-hydroxy-11,12-epoxyeicosatrienoic acid (15-H-11,12-EETA). In the present study, the acid-stable 13-hydroxy-trans-14,15-epoxy-eicosatrienoic acid (13-H-14,15-EETA) was identified and its vascular activities characterized. Rabbit aorta, mesenteric arteries, and the combination of 15-LO and cytochrome P450 2J2 converted AAto two distinct HEETA metabolites. The HEETA metabolites were resistant to acidic hydrolysis but were hydrolyzed by recombinant sEH to a more polar metabolite identified by mass spectrometry as 13,14,15-THETA. Mass spectrometric analyses and HPLC comigration identified the HEETAs as threo- and erythro-diastereomers of 13-H-trans-14,15-EETA. Erythro- and threo-diastereomers of 13-H-trans-14,15-EETA relaxed endothelium-denuded rabbit small mesenteric arteries with maximum relaxations of 22.6±6.0{\%} and 8.6±4.3{\%}, respectively. Apamin (10-7 M) inhibited the relaxations to the erythro-isomer(maximumrelaxation=1.2±5.6{\%}) and increasing [K+]o from 4.6 to 30 mM blocked relaxations to both isomers. In cell-attached patches of mesenteric arterial smooth muscle cells (SMCs), erythro-13-H-trans-14,15-EETA (1-3 × 10-6 M) increased mean open time of small conductance K+ channels (13-14 pS) from 0.0007 ± 0.0007 to 0.0053 ± 0.0042. This activation was inhibited by apamin. The erythro, but not the threo, isomer blocked angiotensin II-stimulated aortic SMC migration. These studies demonstrate that 13-H-14,15-EETAs induces vascular relaxation via K+ channel activation to cause SMC hyperpolarization. Thus, 13-H-14,15-EETA represents a new endothelial factor.",
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AU - Gauthier, Kathryn M.

AU - Nithipatikom, Kasem

AU - Hammock, Bruce D.

AU - Falck, J R

AU - Narsimhaswamy, Dubasi

AU - Campbell, William B.

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