NADPH-dependent microsomal metabolism of 14,15-epoxyeicosatrienoic acid to diepoxides and epoxyalcohols

doi:10.1016/0003-9861(88)90111-7

Archives of Biochemistry and Biophysics

Volume 261, Issue 1, 15 February 1988, Pages 122-133

https://doi.org/10.1016/0003-9861(88)90111-7 Get rights and content

Abstract

The arachidonic acid epoxygenase metabolite 14,15-epoxyeicosatrienoic acid is further metabolized by rat liver microsomal fractions to regioisomeric diepoxides and epoxyalcohols. Diepoxides result from epoxidation at the 5,6-, 8,9-, or 11,12-olefins. Hydroxylation leading to epoxyalcohols with a cis, trans-conjugated dienol occurs at carbons 5, 8, 9, or 12. Structural assignments were established by chromatographic and mass spectral comparisons with synthetic standards. The reaction requires NADPH and is inhibited by typical cytochrome P-450 inhibitors. Analysis of the time course of product formation during arachidonic acid oxidation by rat liver microsomal fractions indicated that all four regioisomeric epoxyeicosatrienoic acids can be further metabolized by the enzyme system.

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5,14-HEDGE, a 20-HETE mimetic, reverses hypotension and improves survival in a rodent model of septic shock: Contribution of soluble epoxide hydrolase, CYP2C23, MEK1/ERK1/2/IKKβ/IκB-α/NF-κB pathway, and proinflammatory cytokine formation
2013, Prostaglandins and Other Lipid Mediators
Citation Excerpt :
Although each enzyme is able to convert AA to all four EET regioisomers, 11,12- and 14,15-EET have been reported as the main products of these epoxygenases [23]. EETs can be further metabolized by several pathways including hydration by soluble epoxide hydrolase (sEH) [24,25], esterification to glycerophospholipids [26,27], and β-oxidation by CYPs and COXs [28–31]. EETs are rapidly hydrated in vivo by epoxide hydrolases, primarily sEH in the cytosol, to their more stable and less active corresponding diols, dihydroxyeicosatrienoic acids (DHETs) [24,25].
We have previously demonstrated that a stable synthetic analog of 20-HETE, N-[20-hydroxyeicosa-5(Z),14(Z)-dienoyl]glycine (5,14-HEDGE), restores vascular reactivity, blood pressure, and heart rate in endotoxemic rats. The aim of this study was to determine whether decreased renal expression and activity of soluble epoxide hydrolase (sEH), MEK1, ERK1/2, IKKβ, IκB-α, and NF-κB as well as systemic and renal proinflammatory cytokine production associated with increased expression and activity of CYP2C23 contributes to the effect of 5,14-HEDGE to prevent hypotension, tachycardia, inflammation, and mortality in response to systemic administration of lipopolysaccharide (LPS). Blood pressure fell by 33 mmHg and heart rate rose by 57 beats/min in LPS (10 mg/kg, i.p.)-treated rats. Administration of LPS also increased mRNA and protein expression of sEH associated with a decrease in CYP2C23 mRNA and protein expression. Increased activity of sEH and p-MEK1, p-ERK1/2, p-IκB-α, NF-κB, and p-NF-κB protein levels as well as TNF-α and IL-8 production by LPS were also associated with a decreased activity of AA epoxygenases. These effects of LPS were prevented by 5,14-HEDGE (30 mg/kg, s.c.; 1 h after LPS). Treatment of endotoxemic mice with 5,14-HEDGE also raised the survival rate of animals from 84% to 98%. A competitive antagonist of vasoconstrictor effects of 20-HETE, 20-hydroxyeicosa-6(Z),15(Z)-dienoic acid, 20-HEDE (30 mg/kg, s.c.; 1 h after LPS) prevented the effects of 5,14-HEDGE on blood pressure, heart rate, expression and/or activity of sEH, CYP2C23, and ERK1/2 as well as TNF-α and IL-8 levels in rats treated with LPS. These results suggest that decreased expression and/or activity of sEH and MEK1/ERK1/2/IKKβ/IκB-α/NF-κB pathway as well as proinflammatory cytokine production associated with increased CYP2C23 expression and antiinflammatory mediator formation participate in the protective effect of 5,14-HEDGE against hypotension, tachycardia, inflammation, and mortality in the rodent model of septic shock.
Epoxygenase eicosanoids: Synthesis of tetrahydrofuran-diol metabolites and their vasoactivity
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Human CYP4F3s are the main catalysts in the oxidation of fatty acid epoxides
2004, Journal of Lipid Research
CYP4F isoforms are involved in the oxidation of important cellular mediators such as leukotriene B₄ (LTB4) and prostaglandins. The proinflammatory agent LTB4 and cytotoxic leukotoxins have been associated with several inflammatory diseases. We present evidence that the hydroxylation of Z 9(10)-epoxyoctadecanoic, Z 9(10)-epoxyoctadec-Z 12-enoic, and Z 12(13)-epoxyoctadec-Z 9-enoic acids and that of monoepoxides from arachidonic acid [epoxyeicosatrienoic acid (EET)] is important in the regulation of leukotoxin and EET activity. These three epoxidized derivatives from the C18 family (C18-epoxides) were converted to 18-hydroxy-C18-epoxides by human hepatic microsomes with apparent K_m values of between 27.6 and 175 μM. Among recombinant P450 enzymes, CYP4F2 and CYP4F3B catalyzed mainly the ω-hydroxylation of C18-epoxides with an apparent V_max of between 0.84 and 15.0 min⁻¹, whereas the apparent V_max displayed by CYP4F3A, the isoform found in leukocytes, ranged from 3.0 to 21.2 min⁻¹. The rate of ω-hydroxylation by CYP4A11 was experimentally found to be between 0.3 and 2.7 min⁻¹. CYP4F2 and CYP4F3 exhibited preferences for ω-hydroxylation of Z 8(9)-EET, whereas human liver microsomes preferred Z 11(12)-EET and, to a lesser extent, Z 8(9)-EET. Moreover, vicinal diol from both C18-epoxides and EETs were ω-hydroxylated by liver microsomes and by CYP4F2 and CYP4F3.
These data support the hypothesis that the human CYP4F subfamily is involved in the ω-hydroxylation of fatty acid epoxides. These findings demonstrate that another pathway besides conversion to vicinal diol or chain shortening by β-oxidation exists for fatty acid epoxide inactivation.
The CYP4A isoforms hydroxylate epoxyeicosatrienoic acids to form high affinity peroxisome proliferator-activated receptor ligands
2002, Journal of Biological Chemistry
Cytochromes P450 of the CYP2Cand CYP4A gene subfamilies metabolize arachidonic acid to 5,6-, 8,9-, 11,12-, and 14,15-epoxyeicosatrienoic acids (EETs) and to 19- and 20-hydroxyeicosatetraenoic acids (HETEs), respectively. Abundant functional studies indicate that EETs and HETEs display powerful and often opposing biological activities as mediators of ion channel activity and regulators of vascular tone and systemic blood pressures. Incubation of 8,9-, 11,12-, and 14,15-EETs with microsomal and purified forms of rat CYP4A isoforms led to rapid NADPH-dependent metabolism to the corresponding 19- and 20-hydroxylated EETs. Comparisons of reaction rates and catalytic efficiency with those of arachidonic and lauric acids showed that EETs are one of the best endogenous substrates so far described for rat CYP4A isoforms. CYP4A1 exhibited a preference for 8,9-EET, whereas CYP4A2, CYP4A3, and CYP4A8 preferred 11,12-EET. In general, the closer the oxido ring is to the carboxylic acid functionality, the higher the rate of EET metabolism and the lower the regiospecificity for the EET ω-carbon. Analysis of cis-parinaric acid displacement from the ligand-binding domain of the human peroxisome proliferator-activated receptor-α showed that ω-hydroxylated 14,15-EET bound to this receptor with high affinity (K _i = 3 ± 1 nm). Moreover, at 1 μm, the ω-alcohol of 14,15-EET or a 1:4 mixture of the ω-alcohols of 8,9- and 11,12-EETs activated human and mouse peroxisome proliferator-activated receptor-α in transient transfection assays, suggesting a role for them as endogenous ligands for these orphan nuclear receptors.
Stereospecific synthesis of EET metabolites via Suzuki-Miyaura coupling
2001, Tetrahedron Letters
Epoxygenase Pathways of Arachidonic Acid Metabolism
2001, Journal of Biological Chemistry
Citation Excerpt :
Whereas the rapid conversion of EETs to their corresponding diols has generally been viewed as a process whereby EETs are rendered biologically inactive, DHETs have been shown to be vasoactive in the coronary circulation and are inhibitors of the hydroosmotic effect of arginine vasopressin in the kidney (62-64). Incubations of EETs with rat or mouse liver microsomal P450 results in the production of a series of diepoxyeicosadienoic acids (diepoxides) and monohydroxyepoxyeicosatrienoic acids (epoxyalcohols) (59, 65). The diepoxides can be further metabolized by sEH to diol epoxides, which cyclize to the corresponding tetrahydrofuran-diols (THF-diols) (59).

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^☆: This work was supported by USPHS Grants GM 31278 and 33541 and the Robert A. Welch Foundation (I-782).

^☆☆: The chemical syntheses and spectral data of the oxygenated standards appear as a Miniprint Supplement.

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NADPH-dependent microsomal metabolism of 14,15-epoxyeicosatrienoic acid to diepoxides and epoxyalcohols☆,☆☆

Abstract

J. Biol. Chem

Biochem. Biophys. Res. Commun

J. Biol. Chem

Arch. Biochem. Biophys

J. Lipid Res

Biochim. Biophys. Acta

Arch. Biochem. Biophys

Arch. Biochem. Biophys

Biochem. Biophys. Res. Commun

Biochem. Biophys. Res. Commun

J. Biol. Chem

J. Biol. Chem