Nitro-fatty Acid Reaction with Glutathione and Cysteine

Self Cite

105

127

Nitrated derivatives of fatty acids (NO 2 -FA) are pluripotent cell-signaling mediators that display anti-inflammatory properties. Current understanding of NO 2 -FA signal transduction lacks insight into how or if NO 2 -FA are modified or metabolized upon formation or administration in vivo. Here the disposition and metabolism of nitro-9-cis-octadecenoic (18:1-NO 2 ) acid was investigated in plasma and liver after intravenous injection in mice. High performance liquid chromatography-tandem mass spectrometry analysis showed that no 18:1-NO 2 or metabolites were detected under basal conditions, whereas administered 18:1-NO 2 is rapidly adducted to plasma thiol-containing proteins and glutathione. NO 2 -FA are also metabolized via ␤-oxidation, with high performance liquid chromatographytandem mass spectrometry analysis of liver lipid extracts of treated mice revealing nitro-7-cis-hexadecenoic acid, nitro-5-cis-tetradecenoic acid, and nitro-3-cis-dodecenoic acid and corresponding coenzyme A derivatives of 18:1-NO 2 as metabolites. Additionally, a significant proportion of 18:1-NO 2 and its metabolites are converted to nitroalkane derivatives by saturation of the double bond, and to a lesser extent are desaturated to diene derivatives. There was no evidence of the formation of nitrohydroxyl or conjugated ketone derivatives in organs of interest, metabolites expected upon 18:1-NO 2 hydration or nitric oxide ( ⅐ NO) release. Plasma samples from treated mice had significant extents of protein-adducted 18:1-NO 2 detected by exchange to added ␤-mercaptoethanol. This, coupled with the observation of 18:1-NO 2 release from glutathione-18:1-NO 2 adducts, supports that reversible and exchangeable NO 2 -FAthiol adducts occur under biological conditions. After administration of [ 3 H]18:1-NO 2 , 64% of net radiolabel was recovered 90 min later in plasma (0.2%), liver (18%), kidney (2%), adipose tissue (2%), muscle (31%), urine (6%), and other tissue compartments, and may include metabolites not yet identified. In aggregate, these findings show that electrophilic FA nitroalkene derivatives (a) acquire an extended half-life by undergoing reversible and exchangeable electrophilic reactions with nucleophilic targets and (b) are metabolized predominantly via saturation of the double bond and ␤-oxidation reactions that terminate at the site of acyl-chain nitration.

mentioning

confidence: 99%

Nitro-fatty Acid Metabolome: Saturation, Desaturation, β-Oxidation, and Protein Adduction

Rudolph

Schöpfer

Khoo

et al. 2009

Self Cite

105

127

“…For example, adduction of OA-NO 2 with the catalytic Cys-149 of glyceraldehyde-3-phosphate dehydrogenase inhibits enzymatic activity and promotes membrane localization due to enhanced hydrophobicity of the nitroalkene-modified enzyme (15). Compared with other biological electrophilic lipids, nitro-fatty acids react with thiols with a high rate constant (20).…”

mentioning

confidence: 99%

Nitro-oleic Acid, a Novel and Irreversible Inhibitor of Xanthine Oxidoreductase

Kelley

Batthyány

Hundley

et al. 2008

Self Cite

Xanthine oxidoreductase (XOR) generates proinflammatory oxidants and secondary nitrating species, with inhibition of XOR proving beneficial in a variety of disorders. Electrophilic nitrated fatty acid derivatives, such as nitro-oleic acid (OA-NO 2 ), display anti-inflammatory effects with pleiotropic properties. Nitro-oleic acid inhibits XOR activity in a concentrationdependent manner with an IC 50 of 0.6 M, limiting both purine oxidation and formation of superoxide (O 2 . ). Enzyme inhibition by OA-NO 2 is not reversed by thiol reagents, including glutathione, ␤-mercaptoethanol, and dithiothreitol. Structure-function studies indicate that the carboxylic acid moiety, nitration at the 9 or 10 olefinic carbon, and unsaturation is required for XOR inhibition. Enzyme turnover and competitive reactivation studies reveal inhibition of electron transfer reactions at the

“…The signaling actions of NO 2 -FA are primarily ascribed to the electrophilic olefinic carbon situated ␤ to the electron-withdrawing NO 2 substituent, facilitating kinetically rapid and reversible Michael addition with nucleophilic amino acids (i.e. Cys and His) (14). NO 2 -FA adduction of proteins and GSH occurs in model systems and clinically, with this reaction influencing apparent blood and tissue concentrations (15).…”

mentioning

confidence: 99%

Covalent Peroxisome Proliferator-activated Receptor γ Adduction by Nitro-fatty Acids

Schöpfer

Cole

Groeger

et al. 2010

Self Cite

153

155

The peroxisome proliferator-activated receptor-␥ (PPAR␥) binds diverse ligands to transcriptionally regulate metabolism and inflammation. Activators of PPAR␥ include lipids and antihyperglycemic drugs such as thiazolidinediones (TZDsThe rapidly expanding global burden of type II diabetes mellitus (DM) 3 and the concomitant increased risk for cardiovascular disease (1, 2) have motivated better understanding of relevant cell signaling pathways and potential therapeutic strategies. One major characteristic of metabolic syndrome and DM is insulin resistance, leading to hyperglycemia and dyslipidemia. Following initial clinical use of TZDs as anti-hyperglycemic agents to treat DM in the late 1990s, the nuclear receptor PPAR␥ was discovered as their molecular target. This receptor is expressed primarily in adipose tissue, muscle, and macrophages, where it regulates glucose uptake, lipid metabolism/ storage, and cell proliferation/differentiation (3-5). Thus, PPAR␥ ligands and allied downstream signaling events play a pivotal role in both the development and treatment of DM (6, 7). This is underscored by the observation that mutations in the C-terminal helix 12 of the ligand-binding domain (LBD) of PPAR␥ (e.g. P467L or V290M) are linked with severe insulin resistance and the onset of juvenile DM (8).The oxidizing inflammatory milieu contributing to the pathogenesis of obesity, diabetes, and cardiovascular disease also promotes diverse biomolecule oxidation, nitrosation, and nitration reactions by O 2 and ⅐ NO-derived species. Although oxidized fatty acids typically propagate proinflammatory conditions, the recently detected class of NO 2 -FA act as anti-inflammatory mediators. Nitroalkene derivatives of oleic acid (OA-NO 2 ) and linoleic acid (LNO 2 ) have been detected in healthy human blood and murine cardiac tissue. The levels of free/unesterified OA-NO 2 are ϳ1-3 nM in human plasma (9, 10), with OA-NO 2 produced at increased rates and present at higher concentrations during inflammatory and metabolic stress (11-13). The signaling actions of NO 2 -FA are primarily ascribed to the electrophilic olefinic carbon situated ␤ to the electron-withdrawing NO 2 substituent, facilitating kinetically rapid and reversible Michael addition with nucleophilic amino acids (i.e. Cys and His) (14). NO 2 -FA adduction of proteins and GSH occurs in model systems and clinically, with this reaction