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Generation and esterification of electrophilic fatty acid nitroalkenes in triacylglycerides.

ABSTRACT: Electrophilic fatty acid nitroalkenes (NO(2)-FA) are products of nitric oxide and nitrite-mediated unsaturated fatty acid nitration. These electrophilic products induce pleiotropic signaling actions that modulate metabolic and inflammatory responses in cell and animal models. The metabolism of NO(2)-FA includes reduction of the vinyl nitro moiety by prostaglandin reductase-1, mitochondrial ?-oxidation, and Michael addition with low molecular weight nucleophilic amino acids. Complex lipid reactions of fatty acid nitroalkenes are not well defined. Herein we report the detection and characterization of NO(2)-FA-containing triacylglycerides (NO(2)-FA-TAG) via mass spectrometry-based methods. In this regard, unsaturated fatty acids of dietary triacylglycerides are targets for nitration reactions during gastric acidification, where NO(2)-FA-TAG can be detected in rat plasma after oral administration of nitro-oleic acid (NO(2)-OA). Furthermore, the characterization and profiling of these species, including the generation of beta oxidation and dehydrogenation products, could be detected in NO(2)-OA-supplemented adipocytes. These data revealed that NO(2)-FA-TAG, formed by either the direct nitration of esterified unsaturated fatty acids or the incorporation of nitrated free fatty acids into triacylglycerides, contribute to the systemic distribution of these reactive electrophilic mediators and may serve as a depot for subsequent mobilization by lipases to in turn impact adipocyte homeostasis and tissue signaling events.

SUBMITTER: Fazzari M

PROVIDER: S-EPMC4615386 | biostudies-literature | 2015 Oct

REPOSITORIES: biostudies-literature

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Generation and esterification of electrophilic fatty acid nitroalkenes in triacylglycerides.

Fazzari Marco M Khoo Nicholas N Woodcock Steven R SR Li Lihua L Freeman Bruce A BA Schopfer Francisco J FJ

Free radical biology & medicine 20150609

Electrophilic fatty acid nitroalkenes (NO(2)-FA) are products of nitric oxide and nitrite-mediated unsaturated fatty acid nitration. These electrophilic products induce pleiotropic signaling actions that modulate metabolic and inflammatory responses in cell and animal models. The metabolism of NO(2)-FA includes reduction of the vinyl nitro moiety by prostaglandin reductase-1, mitochondrial β-oxidation, and Michael addition with low molecular weight nucleophilic amino acids. Complex lipid reactio ...[more]

PMID: 26066303

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Project description:UnlabelledDietary ω-3 polyunsaturated fatty acids (PUFAs) decrease cardiovascular risk via suppression of inflammation. The generation of electrophilic α,β-unsaturated ketone derivatives of the ω-3 PUFAs docosahexaenoic acid (DHA) and docosapentaenoic acid (DPA) in activated human macrophages is catalyzed by cyclooxygenase-2 (Cox-2). These derivatives are potent pleiotropic anti-inflammatory signaling mediators that act via mechanisms including the activation of Nrf2-dependent phase 2 gene expression and suppression of pro-inflammatory NF-κB-driven gene expression. Herein, the endogenous generation of ω-3 PUFAs electrophilic ketone derivatives and their hydroxy precursors was evaluated in human neutrophils. In addition, their dietary modulation was assessed through a randomized clinical trial.MethodsEndogenous generation of electrophilic omega-3 PUFAs and their hydroxy precursors was evaluated by mass spectrometry in neutrophils isolated from healthy subjects, both at baseline and upon stimulation with calcium ionophore. For the clinical trial, participants were healthy adults 30-55 years of age with a reported EPA+DHA consumption of ≤300 mg/day randomly assigned to parallel groups receiving daily oil capsule supplements for a period of 4 months containing either 1.4 g of EPA+DHA (active condition, n = 24) or identical appearing soybean oil (control condition, n = 21). Participants and laboratory technicians remained blinded to treatment assignments.Results5-lypoxygenase-dependent endogenous generation of 7-oxo-DHA, 7-oxo-DPA and 5-oxo-EPA and their hydroxy precursors is reported in human neutrophils stimulated with calcium ionophore and phorbol 12-myristate 13-acetate (PMA). Dietary EPA+DHA supplementation significantly increased the formation of 7-oxo-DHA and 5-oxo-EPA, with no significant modulation of arachidonic acid (AA) metabolite levels.ConclusionsThe endogenous detection of these electrophilic ω-3 fatty acid ketone derivatives supports the precept that the benefit of ω-3 PUFA-rich diets can be attributed to the generation of electrophilic oxygenated metabolites that transduce anti-inflammatory actions rather than the suppression of pro-inflammatory AA metabolites.Trial registrationClinicalTrials.gov NCT00663871.

Project description:Fatty acid nitroalkenes are reversibly-reactive electrophiles, endogenously detectable at nM concentrations, displaying anti-inflammatory actions. Nitroalkenes like 9- or 10-nitro-octadec-9-enoic acid (e.g. nitro-oleic acid, OA-NO2) pleiotropically suppress cardiovascular inflammatory responses, with pulmonary responses less well defined. C57BL/6 J male mice were intratracheally administered bleomycin (3 U/kg, ITB), to induce pulmonary inflammation and acute injury, or saline and were treated with 50 μL OA-NO2 (50 μg) or vehicle in the same instillation and 72 h post-exposure to assess anti-inflammatory properties. Bronchoalveolar lavage (BAL) and lung tissue were collected 7d later. ITB mice lost body weight, with OA-NO2 mitigating this loss (-2.3 ± 0.94 vs -0.4 ± 0.83 g). Histology revealed ITB induced cellular infiltration, proteinaceous debris deposition, and tissue injury, all significantly reduced by OA-NO2. Flow cytometry analysis of BAL demonstrated loss of Siglec F+/F4/80+/CD45+ alveolar macrophages with ITB (89 ± 3.5 vs 30 ± 3.7%). Analysis of CD11b/CD11c expressing cells showed ITB-induced non-resident macrophage infiltration (4 ± 2.3 vs 43 ± 2.4%) was decreased by OA-NO2 (24 ± 2.4%). Additionally, OA-NO2 attenuated increases in mature, activated interstitial macrophages (23 ± 4.8 vs. 43 ± 5.4%) in lung tissue digests. Flow analysis of CD31-/CD45-/Sca-1+ mesenchymal cells revealed ITB increased CD44+ populations (1 ± 0.4 vs 4 ± 0.4MFI), significantly reduced by OA-NO2 (3 ± 0.4MFI). Single cell analysis of mesenchymal cells by western blotting showed profibrotic ZEB1 protein expression induced by ITB. Lung digest CD45+ cells revealed ITB increased HMGB1+ cells, with OA-NO2 suppressing this response. Inhibition of HMGB1 expression correlated with increased basal phospholipid production and SP-B expression in the lung lining. These findings indicate OA-NO2 inhibits ITB-induced pro-inflammatory responses by modulating resident cell function.

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Generation and esterification of electrophilic fatty acid nitroalkenes in triacylglycerides.

Publications

Generation and esterification of electrophilic fatty acid nitroalkenes in triacylglycerides.

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