Nitro-fatty acids in cardiovascular regulation and diseases: characteristics and molecular mechanisms

Abstract

Electrophilic nitro-fatty acids (NO2-FAs) are endogenously formed by redox reactions of nitric oxide ((.)NO)- and nitrite ((.)NO2)- derived nitrogen dioxide with unsaturated fatty acids. Nitration preferentially occurs on polyunsaturated fatty acids with conjugated dienes under physiological or pathophysiological conditions such as during digestion, metabolism and as adaptive inflammatory processes. Nitro-fatty acids are present in free and esterified forms achieving broad biodistribution in humans and experimental models. Structural, functional and biological characterization of NO2-FAs has revealed clinically relevant protection from inflammatory injury in a number of cardiovascular, renal and metabolic experimental models. NO2-FAs are engaged in posttranslational modifications (PTMs) of a selective redox sensitive pool of proteins and regulate key adaptive signaling pathways involved in cellular homeostasis and inflammatory response. Here, we review and update the biosynthesis, metabolism and signaling actions of NO2-FAs, highlighting their diverse protective roles relevant to the cardiovascular system.

Figure 1

Endogenous NO₂-FA formation and metabolism. (A) Experimental models and human studies have determined that unsaturated fatty acid nitration is readily bioavailable upon oral delivery of PUFA along with inorganic nitrate (NO₃). Oral and gut microbiome mediates NO₃ reduction to nitrite (NO₂) which facilitates formation of NO₂-FAs (19) (B) Nitration derivatives of unsaturated fatty acids are generated in monocytes and macrophages as products of inflammatory-derived reactive species reaction with lipids (11, 12). Oxidative inflammatory conditions lead to nitric oxide (NO) and nitrite (NO₂)-dependent unsaturated fatty acid nitration. Fatty acid nitration products are also generated in vivo in the mitochondria of cardiomyocytes following ischemic precondition (8, 22). (C) Post-translational protein modifications induced by NO₂-FAs constitute the primary mechanism of cell signaling via Michael addition of cysteine and histidine residues regulating key metabolic and inflammatory processes. NO₂-FAs are the further endogenously metabolized by saturation via PtGR-1 and glutathione conjugation and subsequent b-oxidation and ultimately excreted via urine (20).

Figure 2

Nitroalkene mediated posttranslational modification (Michael addition reactions). Nitroalkenes react with the thiolate anion of glutathione (GSH) and cysteines (Cys) via Michael addition with a reaction constant for OA-NO₂ with glutathione is 183 M⁻¹s⁻¹. The reaction with thiols is reversible and in the case of cysteine displays a K_D of 7.5. × 10⁻⁶ M (20).

Figure 3

Key posttranslational modifications and signaling pathways regulated by NO₂-FAs. NO₂-FAs mediate posttranslational modification by regulating either their enzymatic activity, membrane receptor interaction with agonists, mitochondrial uncoupling or transcriptional regulation and signaling pathways centrally involved in cellular homeostasis. At the transcriptional level, NO₂-FAs are endogenous PPARγ ligands and regulate PPAR transcriptional activity by selective corepressor displacement and transcriptional activation of peroxisome proliferator activating elements (PPRE) (40). NO₂-FAs regulates the heat shock response by activation of heat shock factors (HSFs), such as HSF1, translocate to the nucleus, trimerize and bind to heat shock elements (HSE) driving the expression of heat shock proteins (45,60). NO₂-FAs bind Keap1 and stabilize Nrf2 transcription factor, which binds to antioxidant response elements (ARE), and regulates antioxidant and detoxifying gene expression (43, 44). NO₂-FAs inhibits the formation of pro-inflammatory lipid mediators (e.g. PGE₂, LTB4, 5- 12-HETE) and promotes anti-inflammatory eicosanoids (EETs) by inhibiting the activity of 5-lipoxygenase (80) and soluble epoxide hydrolase (75), respectively. At the cellular membrane, NO₂-FAs interfere with angiotensin II receptor (9), TGFβ receptor (87) or Toll-like receptor 4 (31) by preventing G-protein coupled receptor second messenger signaling, receptor dimerization or membrane recruitment, respectively.

Figure 4

Anti-inflammatory actions of NO₂-FAs in the vasculature. NO₂-FAs act locally in the vasculature by multiple anti-inflammatory processes during the course of atherosclerotic inflammatory responses. These include inhibition of platelet aggregation by limiting the production of pro-thrombotic mediators (71,72), preventing endothelial dysfunction leading to vascular permeability, and reducing leukocyte chemotaxis by inhibiting the formation of pro-inflammatory cytokines (42). NO₂-FAs inhibits monocyte recruitment to the vascular wall during the formation of the atherosclerotic plaque and prevents macrophage activation and lipid accumulation leading to foam cell formation, which further promotes excessive leukocyte recruitment and plaque instability (10). NO₂-FAs reduces smooth muscle cell migration and proliferation (43,58) and inhibits metalloproteinase activity (50) from activated macrophages and migrating smooth muscle cells to the intimal lesion promoting the stability of the plaque’s fibrous cap (10) allowing vascular homeostasis to be restored.