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PHAGOCYTES
From the Department of Cell Biology, the Department of
Cardiology, and the Center for Cardiovascular Diagnostics, Preventive
Cardiology Section, Cleveland Clinic Foundation, OH; the Chemistry
Department, Cleveland State University, OH; and the Inflammation
Program and Department of Medicine, University of Iowa and Veterans
Administration Medical Center, Iowa City.
More than a decade ago it was demonstrated that neutrophil
activation in plasma results in the time-dependent formation of lipid
hydroperoxides through an unknown, ascorbate-sensitive pathway. It is
now shown that the mechanism involves myeloperoxidase
(MPO)-dependent use of multiple low-molecular-weight substrates
in plasma, generating diffusible oxidant species. Addition of activated
human neutrophils (from healthy subjects) to plasma (50%, vol/vol)
resulted in the peroxidation of endogenous plasma lipids by catalase-,
heme poison-, and ascorbate-sensitive pathways, as assessed
by high-performance liquid chromatography (HPLC) with on-line
electrospray ionization tandem mass spectrometric analysis of
free and lipid-bound 9-HETE and 9-HODE. In marked contrast, neutrophils
isolated from multiple subjects with MPO deficiency failed to initiate
peroxidation of plasma lipids, but they did so after
supplementation with isolated human MPO. MPO-dependent use of a
low-molecular-weight substrate(s) in plasma for initiating lipid
peroxidation was illustrated by demonstrating that the filtrate of
plasma (10-kd MWt cutoff) could supply components required for
low-density lipoprotein lipid peroxidation in the presence of MPO and
H2O2. Subsequent HPLC fractionation of plasma
filtrate (10-kd MWt cutoff) by sequential column chromatography identified nitrite, tyrosine, and thiocyanate as major endogenous substrates and 17 The peroxidation of lipids and the consequent
generation of bioactive lipid oxidation products are believed to play
important roles in the pathogenesis of atherosclerosis and other
inflammatory processes.1-5 Lipoxygenase, cyclooxygenase,
and cytochrome P450 are considered the primary enzymatic participants
in these events.6-10 These enzymes are expressed in
leukocytes and catalyze the direct insertion of molecular oxygen
(O2) into polyenoic fatty acids, forming hydroperoxides and
other advanced oxidation products.8 Whether alternative
chemical pathways contribute to the oxidation of lipoproteins and
lipids in complex biologic matrices, such as plasma, has not yet been
fully defined.
We and others have proposed that another potential pathway for
initiating lipid peroxidation in vivo may involve myeloperoxidase (MPO), a heme protein present in neutrophils, monocytes, and certain subpopulations of tissue macrophages.11-13 On phagocyte
activation in peripheral tissues and fluids, MPO is secreted into the
extracellular milieu and into the phagolysosome, where it uses hydrogen
peroxide (H2O2) generated during a respiratory
burst as a cosubstrate. Activated intermediates, Compounds 1 and 2, are
sequentially formed that generate reactive oxidants and diffusible
radical species through 2- and 1-electron oxidation reactions,
respectively.5,14 At plasma levels of halides, chloride
(Cl In the current study we sought to definitively establish whether human
leukocytes use MPO for catalyzing the oxidation of lipids in complex
biologic matrices, such as in plasma, where numerous competing
cosubstrates for the enzyme are present. We also sought to identify
chemically the component(s) in plasma that serve as preferred
substrates for the MPO-H2O2 system of leukocytes for the initiation of lipid peroxidation. In studies using
leukocytes isolated from healthy and MPO-deficient subjects, in
combination with HPLC with on-line electrospray ionization tandem mass spectrometry (LC/ESI/MS/MS), we now show that human neutrophils use MPO to initiate lipid peroxidation in whole plasma through multiple distinct diffusible substrates.
Chemicals and reagents
General procedures
Data are presented as mean ± SD. Comparisons between control and tested groups were made using nonparametric analysis, and P < .05 was considered significant between 2 groups. MPO and lipoprotein isolation MPO (donor: hydrogen peroxide, oxidoreductase, EC 1.11.1.7) was isolated and characterized as described.28,51 Purity of isolated MPO was established by demonstrating R/Z 0.85
(A430/A280), sodium dodecyl
sulfate-polyacrylamide gel electrophoresis analysis with Coomassie
blue staining, and in-gel tetramethylbenzidine peroxidase staining to
confirm no eosinophil peroxidase contamination.51 Purified
MPO was stored in 50% glycerol at  20°C. Enzyme concentration was
determined spectrophotometrically ( 430 = 170 000
M 1cm1).52 LDL was isolated
from fresh plasma by sequential ultracentrifugation as a
1.019 < d < 1.063 g/mL fraction, and dialysis was performed in
sealed jars under argon atmosphere.53 Final preparations were kept in 50 mM sodium phosphate (pH 7.0), 100 µM DTPA, and were
stored under N2 until use. LDL concentrations are expressed per milligram LDL protein.
Human neutrophil preparations Human neutrophils were isolated from whole blood obtained from healthy and MPO-deficient subjects, as described.54 Neutrophil preparations were suspended in HBSS (Mg2+-, Ca2+-, phenol-, and bicarbonate-free, pH 7.0) and were used immediately for experiments. Molecular characterization of 2 of the MPO-deficient subjects have been reported in previous publications.55,56Lipid peroxidation reaction Isolated human neutrophils (106/mL) were incubated at 37°C with either 50% (vol/vol) normal human plasma or isolated human LDL (0.2 mg/mL) under air in HBSS supplemented with 100 µM DTPA. Neutrophils were activated by adding 200 nM phorbol myristate acetate (PMA) and were maintained in suspension by gentle mixing every 5 minutes. After 2 hours, reactions were stopped by immersion in ice or water bath, centrifugation at 4°C, and immediate addition of 50 µM butylated hydroxytoluene (BHT) and 300 nM catalase to the supernatant. Lipid peroxidation products in the supernatant were then rapidly assayed as described below.Reactions with isolated MPO were typically performed at 37°C in sodium phosphate buffer (20 mM, pH 7.0) supplemented with 100 µM DTPA using 30 nM MPO, 1 mM glucose (G), and 20 ng/mL GO. Under this condition, a constant flux of H2O2 (0.18 µM/min) was generated by the glucose-glucose oxidase (G/GO) system. Unless otherwise stated, reactions were terminated by immersion in ice or water bath and the addition of 50 µM BHT and 300 nM catalase to the reaction mixture. Lipid extraction and sample preparation Lipids were extracted and prepared for mass spectrometry analysis under argon or nitrogen atmosphere at all steps. First, hydroperoxides in the reaction mixture were reduced to their corresponding hydroxides by adding SnCl2 (1 mM final). A known amount of deuterated internal standard, 12-HETE-d8 (Cayman Chemical), was added to the sample, and plasma lipids were extracted by adding a mixture of 1 M acetic acid-2-isopropanol-hexane (2/20/30, vol/vol/vol) at a ratio of 5 mL organic solvent mix:1 mL plasma. After vortexing and centrifugation, lipids were extracted to the hexane layer. Plasma was re-extracted by the addition of an equal volume of hexane, followed by vortexing and centrifugation. Cholesteryl ester hydroperoxides (CE-H(P)ODEs) were analyzed as their stable SnCl2- reduced hydroxide forms by drying of the combined hexane extracts under N2, reconstituting samples with 200 µL 2-isopropanol-acetonitrile-water (44/54/2, vol/vol/vol) and storage at80°C under argon until analysis. For
the assay of free fatty acids and their oxidation products, total
lipids (phospholipids, cholesterol esters, triglycerides) were dried
under N2 and resuspended in 1.5 mL 2-isopropanol, and fatty
acids were released by base hydrolysis with 1.5 mL 1 M NaOH at 60°C
for 30 minutes under argon. Hydrolyzed samples were acidified to pH 3.0 with 2 M HCl, and fatty acids were extracted twice with 5 mL hexane.
The combined hexane layers were dried under N2, resuspended in 100 µL methanol, and stored under argon at 80°C until analysis by LC/ESI/MS/MS, as described below.
HPLC fractionation of plasma filtrate To study the role played by low-molecular-weight compounds in plasma as substrates for MPO in the promotion of lipid peroxidation, whole plasma from healthy donors was filtered through a 10-kd MWt cutoff filter (Centriprep YM-10; Millipore, Bedford, MA) by centrifugation. The filtrate of plasma was used either directly or after fractionation by HPLC. Reverse-phase HPLC fractionation was performed using a Beckman C-18 column (4.6 × 250 mm, 5 µm OD; Beckman Instruments, Fullerton, CA). Separation of low-molecular-weight compounds in plasma filtrate (0.5 mL) was carried out at a 1.0 mL/min flow rate with 100% mobile phase A (water containing 0.1% acetic acid) over 10 minutes, followed by a linear gradient generated with 100% mobile phase B (methanol containing 0.1% acetic acid) over 10 minutes, followed by 100% mobile phase B over 5 minutes. Effluent was collected as 1-mL fractions, dried under N2, and resuspended in buffer (0.1 mL) for analysis. Fractionation of plasma filtrate (0.5 mL) by strong anion exchange HPLC (SAX-HPLC) was performed on a SPHERIS HPLC column (4.6 × 250 mm, 5 µm SAX; Phase Separations, Norwalk, CT). The separation of low-molecular-weight compounds in plasma filtrate was carried out at the flow rate of 0.9 mL/min under isocratic conditions using 45 mM ammonium acetate buffer (pH 4.0) as mobile phase. Effluent was collected as 1-mL fractions, dried under N2, and resuspended in buffer (0.1 mL) for analysis.Mass spectrometry LC/ESI/MS/MS was used to quantify free radical-dependent oxidation products of arachidonic acid 9-H(P)ETE and linoleic acid 9-H(P)ODE. Immediately before analysis, 1 vol H2O was added to 5 vol methanol-suspended sample, which was then passed through a 0.22-µm filter (Millipore). Sample (20 µL) was injected onto a Prodigy C-18 column (1 × 250 mm, 5 µm OD, 100 A; Phenomenex, Rancho Palos Verdes, CA) at a flow rate of 50 µL/min. Separation was performed under isocratic conditions using 95% methanol in water as the mobile phase. In each analysis, the entirety of the HPLC column effluent was introduced onto a Quattro II triple quandrupole MS (Micromass). Analyses were performed using electrospray ionization in negative-ion mode with multiple reaction monitoring of parent and characteristic daughter ions specific for the isomers monitored. Transitions monitored were mass-to-charge ratio (m/z) 295 171 for 9-HODE, m/z 319151 for 9-HETE, and m/z 327184 for
12-HETE-d8. N2 was used as the curtain gas in the
electrospray interface. Internal standard 12-HETE-d8 was used
to calculate extraction efficiencies (greater than 80% for all
analyses). External calibration curves constructed with authentic
standards were used to quantify 9-HETE and 9-HODE.
Reverse-phase HPLC quantification of CE-H(P)ODEs Samples (100 µL) reconstituted in methanol (without base hydrolysis) were injected onto a Beckman C-18 column (4.6 × 250 mm, 5 µm OD; Beckman Instruments). Lipids were separated using an isocratic solvent system composed of 2-isopropanol-acetonitrile-water (44/54/2, vol/vol/vol) at a flow rate of 1.5 mL/min. CE-H(P)ODEs were quantified as their stable hydroxide forms by UV detection at 234 nm using CE-9-HODE (Cayman Chemical) for generation of an external calibration curve.
Neutrophils isolated from MPO-deficient subjects fail to initiate lipid peroxidation in plasma but do so after the addition of isolated MPO More than a decade ago, Frei et al57 made the seminal observation that neutrophil activation in plasma results in the time-dependent formation of lipid hydroperoxides. The precise mechanism involved was not identified but was characterized by its sensitivity to ascorbate, which had to be depleted before lipid oxidation products were formed.57 To test the hypothesis that MPO might serve as the enzymatic catalyst for leukocyte-dependent peroxidation of plasma lipids, we compared neutrophils from healthy and MPO-deficient subjects. Lack of functional MPO activity was confirmed by the absence of HOCl production after leukocyte activation (Table 1) and the absence of an MPO activity band within native gels of leukocyte detergent extracts after in-gel tetramethylbenzidine peroxidase staining (data not shown). MPO-deficient neutrophils displayed enhanced agonist-dependent O2 generation relative to comparably
treated normal neutrophils (Table 1), as previously
reported.58-60 Neutrophils isolated from healthy and
MPO-deficient subjects failed to generate detectable levels of NO or
NO
To determine the role of MPO in promoting lipid oxidation in plasma
exposed to activated neutrophils, we next incubated cells with whole
plasma (50%, vol/vol) and physiological levels of Cl
Characterization and reaction requirements for peroxidation of endogenous plasma lipids by activated human neutrophils and isolated human MPO Addition of catalase, but not heat-inactivated catalase, to cell mixtures resulted in the near complete ablation of lipid peroxidation in plasma, strongly suggesting a critical role for H2O2 in the cell-dependent reaction (Figure 2). Incubation of reaction mixtures with SOD failed to attenuate the oxidation of plasma lipids (Figure 2). In contrast, addition of heme poisons (eg, azide, cyanide) and water-soluble antioxidant ascorbate resulted in complete inhibition of neutrophil-dependent peroxidation of plasma lipids. Finally, addition of HOCl scavengers, such as dithiothreitol and the thioether methionine, failed to attenuate neutrophil-dependent peroxidation of endogenous plasma lipids, assessed by quantification of 9-H(P)ODE and 9-H(P)ETE (Figure 2).
Results thus far presented strongly suggest that neutrophils use the
MPO-H2O2 system to generate reactive species
distinct from chlorinating intermediates as the primary oxidants for
initiation of lipid peroxidation in plasma. To confirm a physiological
role for MPO, we next added purified human MPO and an
H2O2-generating system (G/GO) to plasma and
monitored the formation of specific oxidation products by LC/ESI/MS/MS
analysis. Formation of 9-H(P)ODE and 9-H(P)ETE occurred readily and had
an absolute requirement for the presence of MPO and the
H2O2-generating system (Figure 3). Lipid oxidation was again inhibited
by catalase, azide, or ascorbate but was not affected by the addition
of SOD or methionine (Figure 3). Collectively, these results strongly
support a pivotal role for the MPO-H2O2 system
of leukocytes as a primary mechanism for initiating lipid peroxidation
in complex biologic tissues and fluids such as plasma.
Endogenous low-molecular-weight substances in plasma serve as cosubstrates for the MPO-catalyzed initiation of lipid peroxidation in whole plasma The active site of MPO sits at the base of a deep and narrow heme pocket inaccessible to compounds significantly larger than a dipeptide.61 Thus, the ability of isolated MPO and of an H2O2-generating system to initiate lipid peroxidation in plasma is consistent with low-molecular-weight compounds in plasma serving as cosubstrates for MPO to generate diffusible species capable of conveying oxidizing equivalents from the heme group to distant targets, such as plasma lipoproteins. To test this hypothesis, isolated human LDL was incubated with MPO and an H2O2-generating system to generate a physiological flux of H2O2. In the absence of other cosubstrates, no significant oxidation of lipoprotein lipids was observed (Figure 4). In contrast, the addition of low-molecular-weight constituents recovered from plasma that had been filtered through a 10-kd MWt cutoff filter reconstituted the capacity of the MPO-H2O2 system to promote lipid peroxidation (Figure 4, left). In a parallel set of experiments, either plasma or dialyzed plasma was exposed to the MPO-H2O2 system, and the extent of lipid peroxidation was determined. Lipid peroxidation occurred in plasma, but not in dialyzed plasma, exposed to the MPO-H2O2 system (Figure 4, right), suggesting that MPO used low-molecular-weight substrates within plasma to initiate peroxidation of plasma lipids. Consistent with this observation, the subsequent addition of plasma filtrate to reaction mixtures using dialyzed plasma as the target for oxidation restored the ability of the MPO-H2O2 system to promote lipid peroxidation (Figure 4, right).
HPLC fractionation and identification of low-molecular-weight substrates in plasma used by MPO for initiation of lipid peroxidation To identify the component(s) within the filtrate of plasma that served as physiological cosubstrates for MPO and that promoted peroxidation of plasma lipids, plasma filtrate was fractionated on a reverse-phase HPLC column, and the ability of each fraction to provide substrates for MPO-dependent oxidation of LDL surface and core lipids was determined (Figure 5). Comparisons with the retention times for potential candidate substrates for MPO suggested that the early eluting substrate(s) in fraction 3 comigrated with low-molecular-weight organic anions and SCN,
fraction 4 with NO the m/z anticipated for the
molecular cation of tyrosine (data not shown). The high salt content of
fractions 3 and 4 prevented analysis by LC/ESI/MS/MS.
To further identify and confirm the cosubstrates of MPO in plasma that
support the initiation of lipid peroxidation by the enzyme, plasma
filtrate was fractionated by alternative column chromatographies (ion
exchange, straight phase). Under every chromatography system examined,
4 compounds (tyrosine, NO
Confirmation that MPO uses physiologically relevant levels of
nitrite, SCN (100 mM). As shown in
Figure 7, even in the presence of the
competing cosubstrate Cl, biologically relevant
concentrations of NO
Although studies have shown that SCN
MPO is the single most abundant protein in neutrophils, but the
precise substrates used and the biochemical reactions mediated by this
enzyme in vivo are still not fully defined. MPO uses Cl Although lipid peroxidation and lipid-derived signaling molecule
formation are believed to be critical in atherosclerosis and other
inflammatory disorders, the pathways responsible for these processes in
vivo are not fully established. Leukocyte activation in whole plasma
has long been appreciated as a physiological mechanism for promoting
peroxidation of endogenous plasma lipids.57 However, the
enzymatic participant(s) and the reactive intermediates involved in
leukocyte-mediated lipid oxidation within complex matrices such as
plasma have not been directly defined. Results of the current study
definitively identify MPO as a major enzymatic catalyst for promoting
lipid oxidation by activated human neutrophils in plasma. Furthermore,
the MPO-H2O2 system uses low-molecular-weight components in plasma distinct from Cl Recent studies using leukocytes isolated from wild-type and MPO knockout mice report modest differences in their capacity to promote lipoprotein lipid oxidation and inhibition in lipid oxidation by the addition of SOD.37 Two major species differences between murine and human neutrophils include the significant generation of NO by mouse but not human neutrophils and the 6- to 10-fold decrease in MPO content found in murine neutrophils compared with that observed in humans.30,42,43 Moreover, mixed leukocyte preparations rather than isolated neutrophils were used in the studies with MPO knockout mice.37 Whether these factors contributed to the limited role reported for MPO by the elicited murine leukocytes and the sensitivity to SOD remains to be established. Neutrophils from multiple unrelated MPO-deficient humans all failed to promote peroxidation of endogenous lipids in plasma unless supplemented with catalytic levels of purified MPO. Catalase, peroxidase inhibitors, and ascorbate, but not SOD, inhibited leukocyte-dependent peroxidation of plasma lipids, consistent with the MPO-H2O2 system as the responsible mechanism. The current results thus strongly support a major physiological role for MPO in initiating lipid peroxidation by activated human leukocytes and suggest that a function of the enzyme at sites of inflammation may be to generate lipid oxidation products with biologic activity. Four compounds in human plasma could support MPO-dependent peroxidation
of LDL lipids in the presence of physiological levels of
Cl Cigarette smoking is a risk factor for cardiovascular disease, and
plasma levels of SCN Identification of 17 Taken together, our results highlight the probable contribution of MPO
in promoting lipid oxidation at sites of inflammation. The development
of peroxidase inhibitors as novel anti-inflammatory agents thus merits
consideration. The sensitivity of leukocyte- and MPO-dependent
oxidation of plasma lipids to ascorbate also has implications for the
choice of anti-oxidant regimen one considers. In studies using
whole plasma, small unilamellar vesicles, or LDL as targets, ascorbate
inhibits MPO-mediated peroxidation of lipids far more effectively than
that observed with Results of the first large study evaluating nearly 100 MPO-deficient subjects compared with a control population were recently reported.79 In addition to a modestly increased frequency of infectious diseases, MPO deficiency was associated with a decreased incidence of cardiovascular events.79 We have demonstrated a strong positive correlation between the level of MPO per leukocyte and the risk for atherosclerosis in subjects with angiographically defined coronary artery disease status.80 The current results provide further evidence for a potential mechanism contributing to the many links between MPO and cardiovascular disease.
We thank Dave Schmitt for technical assistance. Mass spectrometry experiments were performed at the Cleveland Clinic Foundation Mass Spectrometry Resource within the Center for Cardiovascular Diagnostics and Prevention.
Submitted July 12, 2001; accepted October 23, 2001.
Supported by National Institutes of Health grants HL62526 and HL61878 (S.L.H.) and a Merit Review Grant from the Veterans Administration (W.M.N.). R.Z. is supported by a postdoctoral fellowship from the American Heart Association. Z.S. is the recipient of a Jane Coffin Childs Memorial Fund for Medical Research Fellowship.
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734.
Reprints: Stanley L. Hazen, Cleveland Clinic Foundation, Lerner Research Institute, Dept of Cell Biology, 9500 Euclid Ave, NC-10, Cleveland, OH 44195; e-mail: hazens{at}ccf.org.
1. Steinberg D, Parthasarathy S, Carew TE, Khoo JC, Witztum JL. Beyond cholesterol: modifications of low-density lipoprotein that increase its atherogenicity. N Engl J Med. 1989;320:915-924[Medline] [Order article via Infotrieve]. 2. Berliner JA, Heinecke JW. The role of oxidized lipoproteins in atherogenesis. Free Radic Biol Med. 1996;20:707-727[CrossRef][Medline] [Order article via Infotrieve]. 3. Yla-Herttuala S. Oxidized LDL and atherogenesis. Ann N Y Acad Sci. 1999;874:134-137[CrossRef][Medline] [Order article via Infotrieve].
4.
Chisolm GM, Hazen SL, Fox PL, Cathcart MK.
The oxidation of lipoproteins by monocytes-macrophages: biochemical and biological mechanisms.
J Biol Chem.
1999;274:25959-25962 5. Podrez EA, Abu-Soud HM, Hazen SL. Myeloperoxidase-generated oxidants and atherosclerosis. Free Radic Biol Med. 2000;28:1717-1725[CrossRef][Medline] [Order article via Infotrieve].
6.
Kuehl FA Jr, Egan RW.
Prostaglandins, arachidonic acid, and inflammation.
Science.
1980;210:978-984 7. Higgs GA, Moncada S, Vane JR. Eicosanoids in inflammation. Ann Clin Res. 1984;16:287-299[Medline] [Order article via Infotrieve]. 8. Needleman P, Turk J, Jakschik BA, Morrison AR, Lefkowith JB. Arachidonic acid metabolism. Annu Rev Biochem. 1986;55:69-102[CrossRef][Medline] [Order article via Infotrieve]. 9. Lewis RA, Austen KF, Soberman RJ. Leukotrienes and other products of the 5-lipoxygenase pathway: biochemistry and relation to pathobiology in human diseases. N Engl J Med. 1990;323:645-655[Medline] [Order article via Infotrieve]. 10. Cathcart MK, Folcik VA. Lipoxygenases and atherosclerosis: protection versus pathogenesis. Free Radic Biol Med. 2000;28:1726-1734[CrossRef][Medline] [Order article via Infotrieve]. 11. Schmitt D, Shen Z, Zhang R, et al. Leukocytes utilize myeloperoxidase-generated nitrating intermediates as physiological catalysts for the generation of biologically active oxidized lipids and sterols in serum. Biochemistry. 1999;38:16904-16915[CrossRef][Medline] [Order article via Infotrieve]. 12. Podrez EA, Schmitt D, Hoff HF, Hazen SL. Myeloperoxidase-generated reactive nitrogen species convert LDL into an atherogenic form in vitro. J Clin Invest. 1999;103:1547-1560[Medline] [Order article via Infotrieve].
13.
Savenkova ML, Mueller DM, Heinecke JW.
Tyrosyl radical generated by myeloperoxidase is a physiological catalyst for the initiation of lipid peroxidation in low-density lipoprotein.
J Biol Chem.
1994;269:20394-20400 14. Kettle AJ, Winterbourn CC. Myeloperoxidase: a key regulator of neutrophil oxidant production. Redox Rep. 1997;3:3-15.
15.
Harrison JE, Schultz J.
Studies on the chlorinating activity of myeloperoxidase.
J Biol Chem.
1976;251:1371-1374
16.
Weiss SJ, Lampert MB, Test ST.
Long-lived oxidants generated by human neutrophils: characterization and bioactivity.
Science.
1983;222:625-628
17.
Thomas EL, Fishman M.
Oxidation of chloride and thiocyanate by isolated leukocytes.
J Biol Chem.
1986;261:9694-9702 18. van Dalen CJ, Whitehouse MW, Winterbourn CC, Kettle AJ. Thiocyanate and chloride as competing substrates for myeloperoxidase. Biochem J. 1997;327:487-492.
19.
Klebanoff SJ.
Estrogen binding by leukocytes during phagocytosis.
J Exp Med.
1977;145:983-998 20. Metodiewa D, Dunford HB. The role of myeloperoxidase in the oxidation of biologically active polyhydroxyphenols (substituted catechols). Eur J Biochem. 1990;193:445-448[Medline] [Order article via Infotrieve]. 21. Heinecke JW, Li W, Francis GA, Goldstein JA. Tyrosyl radical generated by myeloperoxidase catalyzes the oxidative cross-linking of proteins. J Clin Invest. 1993;91:2866-2872.
22.
van der Vliet A, Eiserich JP, Halliwell B, Cross CE.
Formation of reactive nitrogen species during peroxidase-catalyzed oxidation of nitrite: a potential additional mechanism of nitric oxide-dependent toxicity.
J Biol Chem.
1997;272:7617-7625 23. Hsuanyu Y, Dunford HB. Oxidation of clozapine and ascorbate by myeloperoxidase. Arch Biochem Biophys. 1999;368:413-420[CrossRef][Medline] [Order article via Infotrieve]. 24. Auchere F, Capeillere-Blandin C. NADPH as a co-substrate for studies of the chlorinating activity of myeloperoxidase. Biochem J. 1999;343:603-613. 25. Dunford HB, Hsuanyu Y. Kinetics of oxidation of serotonin by myeloperoxidase compounds I and II. Biochem Cell Biol. 1999;77:449-457[CrossRef][Medline] [Order article via Infotrieve].
26.
Abu-Soud HM, Hazen SL.
Nitric oxide is a physiological substrate for mammalian peroxidases.
J Biol Chem.
2000;275:37524-37532
27.
Burner U, Furtmuller PG, Kettle AJ, Koppenol WH, Obinger C.
Mechanism of reaction of myeloperoxidase with nitrite.
J Biol Chem.
2000;275:20597-20601
28.
Heinecke JW, Li W, Daehnke HL III, Goldstein JA.
Dityrosine, a specific marker of oxidation, is synthesized by the myeloperoxidase-hydrogen peroxide system of human neutrophils and macrophages.
J Biol Chem.
1993;268:4069-4077 29. Eiserich JP, Hristova M, Cross CE, Jones AD, Freeman BA, Halliwell B. Formation of nitric oxide-derived inflammatory oxidants by myeloperoxidase in neutrophils. Nature. 1998;391:393-397[CrossRef][Medline] [Order article via Infotrieve].
30.
Hazen SL, Zhang R, Shen Z, et al.
Formation of nitric oxide-derived oxidants by myeloperoxidase in monocytes: pathways for monocyte-mediated protein nitration and lipid peroxidation in vivo.
Circ Res.
1999;85:950-958
31.
Leeuwenburgh C, Rasmussen JE, Hsu FF, Mueller DM, Pennathur S, Heinecke JW.
Mass spectrometric quantification of markers for protein oxidation by tyrosyl radical, copper, and hydroxyl radical in low density lipoprotein isolated from human atherosclerotic plaques.
J Biol Chem.
1997;272:3520-3526 32. Beckman JS, Ye YZ, Anderson PG, et al. Extensive nitration of protein tyrosines in human atherosclerosis detected by immunohistochemistry. Biol Chem Hoppe Seyler. 1993;375:81-88.
33.
Leeuwenburgh C, Hardy MM, Hazen SL, et al.
Reactive nitrogen intermediates promote low-density lipoprotein oxidation in human atherosclerotic intima.
J Biol Chem.
1997;272:1433-1436
34.
van der Vliet A, Nguyen MN, Shigenaga MK, Eiserich JP, Marelich GP, Cross CE.
Myeloperoxidase and protein oxidation in cystic fibrosis.
Am J Physiol Lung Cell Mol Physiol.
2000;279:L537-L546 35. Buss IH, Darlow BA, Winterbourn CC. Elevated protein carbonyls and lipid peroxidation products correlating with myeloperoxidase in tracheal aspirates from premature infants. Pediatr Res. 2000;47:640-645[Medline] [Order article via Infotrieve].
36.
MacPherson JC, Comhair SA, Erzurum SC, et al.
Eosinophils are a major source of nitric oxide-derived oxidants in severe asthma: characterization of pathways available to eosinophils for generating reactive nitrogen species.
J Immunol.
2001;166:5763-5772
37.
Noguchi N, Nakano K, Aratani Y, Koyama H, Kodama T, Niki E.
Role of myeloperoxidase in the neutrophil-induced oxidation of low-density lipoprotein as studied by myeloperoxidase-knockout mouse.
J Biochem.(Tokyo)
2000;127:971-976
38.
Carr AC, Frei B.
The nitric oxide congener nitrite inhibits myeloperoxidase/H2O2/ Cl 39. Santanam N, Parthasarathy S. Paradoxical actions of antioxidants in the oxidation of low-density lipoprotein by peroxidases. J Clin Invest. 1995;95:2594-2600. 40. Exner M, Alt E, Hermann M, et al. p-Hydroxyphenylacetaldehyde, the major product of tyrosine oxidation by the activated myeloperoxidase system, can act as an antioxidant in LDL. FEBS Lett. 2001;490:28-31[CrossRef][Medline] [Order article via Infotrieve]. 41. Panasenko OM. The mechanism of the hypochlorite-induced lipid peroxidation. Biofactors. 1997;6:181-190[Medline] [Order article via Infotrieve]. 42. Xie QW, Nathan C. Biological oxidants: generation and injurious consequences. San Diego, CA: Academic Press; 1992.
43.
Rausch PG, Moore TG.
Granule enzymes of polymorphonuclear neutrophils: a phylogenetic comparison.
Blood.
1975;46:913-919 44. Nauseef WM. The proper study of mankind. J Clin Invest. 2001;107:401-403[Medline] [Order article via Infotrieve]. 45. Brennan mL, Anderson MM, Shih DM, et al. Increased atherosclerosis in myeloperoxidase-deficient mice. J Clin Invest. 2001;107:419-430[Medline] [Order article via Infotrieve]. 46. Markwell MA, Haas SM, Bieber LL, Tolbert NE. A modification of the Lowry procedure to simplify protein determination in membrane and lipoprotein samples. Anal Biochem. 1978;87:206-210[CrossRef][Medline] [Order article via Infotrieve]. 47. Nelson DP, Kiesow LA. Enthalpy of decomposition of hydrogen peroxide by catalase at 25 degrees C (with molar extinction coefficients of H2O2 solutions in the UV). Anal Biochem. 1972;49:474-478[CrossRef][Medline] [Order article via Infotrieve]. 48. Capeillere-Blandin C. Oxidation of guaiacol by myeloperoxidase: a two-electron-oxidized guaiacol transient species as a mediator of NADPH oxidation. Biochem J. 1998;336:395-404. 49. Kettle AJ, Winterbourn CC. Assays for the chlorination activity of myeloperoxidase. Methods Enzymol. 1994;233:502-512[Medline] [Order article via Infotrieve]. 50. Fridovich I. In: Greenwald RA, ed. CRC Handbook of Methods: Oxygen Radical Research. Boca Raton, FL: CRC Press; 1985:213-215. 51. Wu W, Chen Y, d'Avignon A, Hazen SL. 3-Bromotyrosine and 3,5-dibromotyrosine are major products of protein oxidation by eosinophil peroxidase: potential markers for eosinophil-dependent tissue injury in vivo. Biochemistry. 1999;38:3538-3548[CrossRef][Medline] [Order article via Infotrieve]. 52. Odajima T, Yamazaki I. Myeloperoxidase of the leukocyte of normal blood, I: reaction of myeloperoxidase with hydrogen peroxide. Biochim Biophys Acta. 1970;206:71-77[Medline] [Order article via Infotrieve]. 53. Hatch FT. Practical methods for plasma lipoprotein analysis. Adv Lipid Res. 1968;6:1-68[Medline] [Order article via Infotrieve].
54.
Hazen SL, Hsu FF, Heinecke JW.
p-Hydroxyphenylacetaldehyde is the major product of L-tyrosine oxidation by activated human phagocytes: a chloride-dependent mechanism for the conversion of free amino acids into reactive aldehydes by myeloperoxidase.
J Biol Chem.
1996;271:1861-1867
55.
Nauseef WM, Cogley M, McCormick S.
Effect of the R569W missense mutation on the biosynthesis of myeloperoxidase.
J Biol Chem.
1996;271:9546-9549 56. Nauseef WM, Cogley M, Bock S, Petrides PE. Pattern of inheritance in hereditary myeloperoxidase deficiency associated with the R569W missense mutation. J Leukoc Biol. 1998;63:264-269[Abstract].
57.
Frei B, Stocker R, Ames BN.
Antioxidant defenses and lipid peroxidation in human blood plasma.
Proc Natl Acad Sci U S A.
1988;85:9748-9752
58.
Nauseef WM, Metcalf JA, Root RK.
Role of myeloperoxidase in the respiratory burst of human neutrophils.
Blood.
1983;61:483-492
59.
Locksley RM, Wilson CB, Klebanoff SJ.
Increased respiratory burst in myeloperoxidase-deficient monocytes.
Blood.
1983;62:902-909 60. Gerber CE, Kuci S, Zipfel M, Niethammer D, Bruchelt G. Phagocytic activity and oxidative burst of granulocytes in persons with myeloperoxidase deficiency. Eur J Clin Chem Clin Biochem. 1996;34:901-908[Medline] [Order article via Infotrieve]. 61. Davey CA, Fenna RE. A resolution x-ray crystal structure of the bisubstrate analogue inhibitor salicylhydroxamic acid bound to human myeloperoxidase: a model for a prereaction complex with hydrogen peroxide. Biochemistry. 1996;35:10967-10973[CrossRef][Medline] [Order article via Infotrieve]. 62. Burtis CA,Ashwood ER, eds. Tietz Fundamentals of Clinical Chemistry. Philadelphia, PA: WB Saunders; 2001. 63. Hazen SL, Heinecke JW. 3-Chlorotyrosine, a specific marker for myeloperoxidase-catalyzed halogenation, is present in human atherosclerotic aorta. J Clin Invest. 1997;99:2075-2081[Medline] [Order article via Infotrieve].
64.
Aratani Y, Koyama H, Nyui S, Suzuki K, Kura F, Maeda N.
Severe impairment in early host defense against Candida albicans in mice deficient in myeloperoxidase.
Infect Immun.
1999;67:1828-1836 65. Nauseef WM. Myeloperoxidase deficiency. Hematol Pathol. 1990;4:165-178[Medline] [Order article via Infotrieve].
66.
van Dalen CJ, Winterbourn CC, Senthilmohan R, Kettle AJ.
Nitrite as a substrate and inhibitor of myeloperoxidase: implications for nitration and hypochlorous acid production at sites of inflammation.
J Biol Chem.
2001;275:11638-11644 67. Scanlon CE, Berger B, Malcom G, Wissler RW. Evidence for more extensive deposits of epitopes of oxidized low-density lipoprotein in aortas of young people with elevated serum thiocyanate levels: PDAY Research Group. Atherosclerosis. 1996;121:23-33[CrossRef][Medline] [Order article via Infotrieve].
68.
Rehak NN, Cecco SA, Niemela JE, Elin RJ.
Thiocyanate in smokers interferes with the Nova magnesium ion-selective electrode.
Clin Chem.
1997;43:1595-1600 69. Botti TP, Amin H, Hiltscher L, Wissler RW. A comparison of the quantitation of macrophage foam cell populations and the extent of apolipoprotein E deposition in developing atherosclerotic lesions in young people: high and low serum thiocyanate groups as an indication of smoking: PDAY Research Group: pathobiological determinants of atherosclerosis in youth. Atherosclerosis. 1996;124:191-202[CrossRef][Medline] [Order article via Infotrieve].
70.
Arlandson M, Decker T, Roongta VA, et al.
Eosinophil peroxidase oxidation of thiocyanate: characterization of major reaction products and a potential sulfhydryl-targeted cytotoxicity system.
J Biol Chem.
2001;276:215-224 71. van Dalen CJ, Kettle AJ. Substrates and products of eosinophil peroxidase. Biochem J. 2001;358:233-239[CrossRef][Medline] [Order article via Infotrieve]. 72. Klaiber EL, Broverman DM, Haffajee CI, Hochman JS, Sacks GM, Dalen JE. Serum estrogen levels in men with acute myocardial infarction. Am J Med. 1982;73:872-881[CrossRef][Medline] [Order article via Infotrieve]. 73. Christeff N, Benassayag C, Carli-Vielle C, Carli A, Nunez EA. Elevated oestrogen and reduced testosterone levels in the serum of male septic shock patients. J Steroid Biochem. 1988;29:435-440[CrossRef][Medline] [Order article via Infotrieve]. 74. Wiseman H, O'Reilly J. Oestrogens as antioxidant cardioprotectants. Biochem Soc Trans. 1997;25:54-59[Medline] [Order article via Infotrieve].
75.
Hulley S, Grady D, Bush T, et al.
Randomized trial of estrogen plus progestin for secondary prevention of coronary heart disease in postmenopausal women: Heart and Estrogen/progestin Replacement Study (HERS) Research Group.
JAMA.
1998;280:605-613
76.
Herrington DM, Reboussin DM, Brosnihan KB, et al.
Effects of estrogen replacement on the progression of coronary-artery atherosclerosis.
N Engl J Med.
2000;343:522-529 77. Podrez EA, Febbraio M, Sheibani N, et al. Macrophage scavenger receptor CD36 is the major receptor for LDL modified by monocyte-generated reactive nitrogen species. J Clin Invest. 2000;105:1095-1108[Medline] [Order article via Infotrieve]. 78. Febbraio M, Podrez EA, Smith JD, et al. Targeted disruption of the class B scavenger receptor CD36 protects against atherosclerotic lesion development in mice. J Clin Invest. 2000;105:1049-1056[Medline] [Order article via Infotrieve]. 79. Kutter D, Devaquet P, Vanderstocken G, Paulus JM, Marchal V, Gothot A. Consequences of total and subtotal myeloperoxidase deficiency: risk or benefit? Acta Haematol. 2000;104:10-15[CrossRef][Medline] [Order article via Infotrieve].
80.
Zhang R, Brennan ML, Fu X, et al.
Myeloperoxidase is an independent marker of coronary artery disease.
JAMA.
2001;286:2136-2142
© 2002 by The American Society of Hematology.
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  S. J. Nicholls and S. L. Hazen Myeloperoxidase, modified lipoproteins, and atherogenesis J. Lipid Res., April 1, 2009; 50(Supplement): S346 - S351. [Abstract] [Full Text] [PDF]
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 C. Erridge, S. Kennedy, C. M. Spickett, and D. J. Webb Oxidized Phospholipid Inhibition of Toll-like Receptor (TLR) Signaling Is Restricted to TLR2 and TLR4: ROLES FOR CD14, LPS-BINDING PROTEIN, AND MD2 AS TARGETS FOR SPECIFICITY OF INHIBITION J. Biol. Chem., September 5, 2008; 283(36): 24748 - 24759. [Abstract] [Full Text] [PDF]
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 K. E. Broderick, P. Potluri, S. Zhuang, I. E. Scheffler, V. S. Sharma, R. B. Pilz, and G. R. Boss Cyanide Detoxification by the Cobalamin Precursor Cobinamide. Experimental Biology and Medicine, May 1, 2006; 231(5): 641 - 649. [Abstract] [Full Text] [PDF]
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 S. Bluml, S. Kirchberger, V. N. Bochkov, G. Kronke, K. Stuhlmeier, O. Majdic, G. J. Zlabinger, W. Knapp, B. R. Binder, J. Stockl, et al. Oxidized Phospholipids Negatively Regulate Dendritic Cell Maturation Induced by TLRs and CD40 J. Immunol., July 1, 2005; 175(1): 501 - 508. [Abstract] [Full Text] [PDF]
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 S. J. Nicholls and S. L. Hazen Myeloperoxidase and Cardiovascular Disease Arterioscler. Thromb. Vasc. Biol., June 1, 2005; 25(6): 1102 - 1111. [Abstract] [Full Text] [PDF]
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S. L. Hazen Myeloperoxidase and Plaque Vulnerability Arterioscler. Thromb. Vasc. Biol., July 1, 2004; 24(7): 1143 - 1146. [Full Text] [PDF]
|
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S. Sugiyama, K. Kugiyama, M. Aikawa, S. Nakamura, H. Ogawa, and P. Libby Hypochlorous Acid, a Macrophage Product, Induces Endothelial Apoptosis and Tissue Factor Expression: Involvement of Myeloperoxidase-Mediated Oxidant in Plaque Erosion and Thrombogenesis Arterioscler. Thromb. Vasc. Biol., July 1, 2004; 24(7): 1309 - 1314. [Abstract] [Full Text] [PDF]
|
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|
 Y. Hasuike, T. Nakanishi, R. Moriguchi, Y. Otaki, M. Nanami, Y. Hama, M. Naka, K. Miyagawa, M. Izumi, and Y. Takamitsu Accumulation of cyanide and thiocyanate in haemodialysis patients Nephrol. Dial. Transplant., June 1, 2004; 19(6): 1474 - 1479. [Abstract] [Full Text] [PDF]
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 E. POLIAKOV, M.-L. BRENNAN, J. MACPHERSON, R. ZHANG, W. SHA, L. NARINE, R. G. SALOMON, and S. L. HAZEN Isolevuglandins, a novel class of isoprostenoid derivatives, function as integrated sensors of oxidant stress and are generated by myeloperoxidase in vivo FASEB J, December 1, 2003; 17(15): 2209 - 2220. [Abstract] [Full Text] [PDF]
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 M. H. Shishehbor, M.-L. Brennan, R. J. Aviles, X. Fu, M. S. Penn, D. L. Sprecher, and S. L. Hazen Statins Promote Potent Systemic Antioxidant Effects Through Specific Inflammatory Pathways Circulation, July 29, 2003; 108(4): 426 - 431. [Abstract] [Full Text] [PDF]
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R. Zhang, M.-L. Brennan, Z. Shen, J. C. MacPherson, D. Schmitt, C. E. Molenda, and S. L. Hazen Myeloperoxidase Functions as a Major Enzymatic Catalyst for Initiation of Lipid Peroxidation at Sites of Inflammation J. Biol. Chem., November 22, 2002; 277(48): 46116 - 46122. [Abstract] [Full Text] [PDF]
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M.-L. Brennan, W. Wu, X. Fu, Z. Shen, W. Song, H. Frost, C. Vadseth, L. Narine, E. Lenkiewicz, M. T. Borchers, et al. A Tale of Two Controversies. DEFINING BOTH THE ROLE OF PEROXIDASES IN NITROTYROSINE FORMATION IN VIVO USING EOSINOPHIL PEROXIDASE AND MYELOPEROXIDASE-DEFICIENT MICE, AND THE NATURE OF PEROXIDASE-GENERATED REACTIVE NITROGEN SPECIES J. Biol. Chem., May 10, 2002; 277(20): 17415 - 17427. [Abstract] [Full Text] [PDF]
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Top
Abstract
Introduction
-estradiol as a novel minor endogenous substrate in
plasma for MPO in promoting peroxidation of plasma lipids. These
results strongly suggest that the MPO-H2O2
system of human leukocytes serves as a physiological mechanism for
initiating lipid peroxidation in vivo.
(Blood. 2002;99:1802-1810)
-tocopherol (R.Z. and S.L.H., unpublished
results, January 2000). Finally, the capacity of the
MPO-H2O2 system of leukocytes to form
bioactive lipid oxidation products in plasma suggests that MPO plays a
broad role in inflammation biology and host defenses. Indeed, we
recently demonstrated that exposure of LDL to the
MPO-H2O2 system of activated monocytes in
media containing NO