Apolipoprotein A-I is a selective target for myeloperoxidase-catalyzed oxidation and functional impairment in subjects with cardiovascular disease

Abstract

In recent studies we demonstrated that systemic levels of protein-bound nitrotyrosine (NO 2 Tyr) and myeloperoxidase (MPO), a protein that catalyzes generation of nitrating oxidants, serve as independent predictors of atherosclerotic risk, burden, and incident cardiac events. We now show both that apolipoprotein A-I (apoA-I), the primary protein constituent of HDL, is a selective target for MPO-catalyzed nitration and chlorination in vivo and that MPO-catalyzed oxidation of HDL and apoA-I results in selective inhibition in ABCA1-dependent cholesterol efflux from macrophages. Dramatic selective enrichment in NO 2 Tyr and chlorotyrosine (ClTyr) content within apoA-I recovered from serum and human atherosclerotic lesions is noted, and analysis of serum from sequential subjects demonstrates that the NO 2 Tyr and ClTyr contents of apoA-I are markedly higher in individuals with cardiovascular disease (CVD). Analysis of circulating HDL further reveals that higher NO 2 Tyr and ClTyr contents of the lipoprotein are each significantly associated with diminished ABCA1-dependent cholesterol efflux capacity of the lipoprotein. MPO as a likely mechanism for oxidative modification of apoA-I in vivo is apparently facilitated by MPO binding to apoA-I, as revealed by cross-immunoprecipitation studies in plasma, recovery of MPO within HDL-like particles isolated from human atheroma, and identification of a probable contact site between the apoA-I moiety of HDL and MPO. To our knowledge, the present results provide the first direct evidence for apoA-I as a selective target for MPO-catalyzed oxidative modification in human atheroma. They also suggest a potential mechanism for MPO-dependent generation of a proatherogenic dysfunctional form of HDL in vivo.that promote oxidative damage, cell injury, and conversion of LDL, the major carrier of cholesterol in plasma, into an atherogenic form (9, 14). Protein-bound nitrotyrosine (NO 2 Tyr), a posttranslational modification specific for protein oxidation by , is markedly enriched within human atheroma (8, 21). Further, recent clinical studies demonstrate that systemic levels of protein-bound NO 2 Tyr serve as an independent predictor of atherosclerotic risk and burden in subjects and are modulated by known CVD risk-reducing therapies such as statins (10,22). Few studies to date have focused on defining the molecular targets of nitration in subjects with CVD, the attendant functional alterations, and the enzymatic participants in nitration.One potential enzymatic source for generation of NO-derived oxidants within human atheroma is the heme protein myeloperoxidase (MPO). MPO utilizes hydrogen peroxide (H 2 O 2 ) and a variety of low-molecular weight organic and inorganic substances as substrates to form reactive oxidant species capable of promoting protein halogenation, nitration, and oxidative cross-linking (4, 5). For example, MPO directly utilizes both NO (23) and the NO metabolite nitrite (NO 2 − ) as substrates in vitro (17-19, 24) and participates

Superoxide Dismutase Inactivation in Pathophysiology of Asthmatic Airway Remodeling and Reactivity

“…38,50 -52 Nitrotyrosine is a valid and semiquantitative marker for the presence of reactive nitrogen species, specifically peroxynitrite or peroxidase-catalyzed oxidative and nitrative events. 13,26,53 Here, evidence for reactive oxygen and nitrogen species involvement in airway epithelial cell apoptosis in asthma include the finding of increased nitrotyrosine in epithelial cells after inhibition of SOD in vitro, and in airway epithelial cells in vivo in asthma in other studies. 13,22,54 Other reports have shown that MnSOD is a target for tyrosine nitration and oxidation, 30,55 which leads to loss of enzyme function, subsequent oxidative damage of mitochondrial proteins, and ultimately cell death.…”

“…Protein-bound nitrotyrosine, chlorotyrosine, bromotyrosine, di-tyrosine, o-tyrosine, and m-tyrosine were quantified by stable isotope dilution liquid chromatographytandem mass spectrometry on a triple quadruple mass spectrometer (API 4000; Applied Biosystems, Foster City, CA) interfaced to an Aria LX Series HPLC multiplexing system (Cohesive Technologies, Franklin, MA) using methods as previously described. 26 Briefly, synthetic [ 13 C 6 ]-labeled standards for each analyte were prepared and added to purified MnSOD samples and used as internal standards for quantification of natural abundance analytes. Simultaneously, universally labeled precursor amino acids, [ 13 C 9 , 15 N 1 ]tyrosine and [ 13 C 9 , 15 N 1 ]phenylalanine (Cambridge Isotopes Inc., Andover, MA), were added.…”

ApoA1 and ApoA1-specific self-antibodies in cardiovascular disease

“…75 ApoA1 oxidation and chlotorination is mainly mediated by neutrophil myeloperoxidase (MPO). 76 MPO-mediated oxidation of Trp 72 leads to the formation of the oxTrp72-ApoA1 variant that is abundantly present (20% of total apoA1 in atherosclerotic vessels) in the plaque and lack an ability to activate ABCA1. 77 MPO-dependent nitration and chlorination of ApoA1 at Tyr 192 along with methionine oxidation results in a loss of ability to activate RCT.…”

LDL protein nitration: Implication for LDL protein unfolding

“…The LDL particle is internalized by cells via the ubiquitously expressed LDL receptor (LDL-R). Rather than binding to the LDL-R commonly present in cells, protein unfolding in modified LDL promotes binding to the scavenger receptors (LDL-SR) in vascular endothelial cells [21] and to CD36 in macrophages [11,22]. ONOO − -modified LDL is recognized by macrophages, thus gaining further relevance in endothelial dysfunction and initiation of atherosclerosis [15,23,24].…”