Myeloperoxidase-generated reactive nitrogen species convert LDL into an atherogenic form in vitro
Eugene A. Podrez, … , Henry F. Hoff, Stanley L. Hazen
Eugene A. Podrez, … , Henry F. Hoff, Stanley L. Hazen
Published June 1, 1999
Citation Information: J Clin Invest. 1999;103(11):1547-1560. https://doi.org/10.1172/JCI5549.
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Article

Myeloperoxidase-generated reactive nitrogen species convert LDL into an atherogenic form in vitro

Abstract

Oxidized LDL is implicated in atherosclerosis; however, the pathways that convert LDL into an atherogenic form in vivo are not established. Production of reactive nitrogen species may be one important pathway, since LDL recovered from human atherosclerotic aorta is enriched in nitrotyrosine. We now report that reactive nitrogen species generated by the MPO-H2O2-NO2– system of monocytes convert LDL into a form (NO2-LDL) that is avidly taken up and degraded by macrophages, leading to massive cholesterol deposition and foam cell formation, essential steps in lesion development. Incubation of LDL with isolated MPO, an H2O2-generating system, and nitrite (NO2–)— a major end-product of NO metabolism—resulted in nitration of apolipoprotein B 100 tyrosyl residues and initiation of LDL lipid peroxidation. The time course of LDL protein nitration and lipid peroxidation paralleled the acquisition of high-affinity, concentration-dependent, and saturable binding of NO2-LDL to human monocyte–derived macrophages and mouse peritoneal macrophages. LDL modification and conversion into a high-uptake form occurred in the absence of free metal ions, required NO2–, occurred at physiological levels of Cl–, and was inhibited by heme poisons, catalase, and BHT. Macrophage binding of NO2-LDL was specific and mediated by neither the LDL receptor nor the scavenger receptor class A type I. Exposure of macrophages to NO2-LDL promoted cholesteryl ester synthesis, intracellular cholesterol and cholesteryl ester accumulation, and foam cell formation. Collectively, these results identify MPO-generated reactive nitrogen species as a physiologically plausible pathway for converting LDL into an atherogenic form.

Authors

Eugene A. Podrez, David Schmitt, Henry F. Hoff, Stanley L. Hazen

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Figure 11

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Cholesteryl oleate synthesis (a), cholesteryl ester mass (b), and choles...
Cholesteryl oleate synthesis (a), cholesteryl ester mass (b), and cholesterol mass (c) of cells exposed to LDL modified by the MPO-H2O2-NO2– system. [125I]LDL (0.2 mg/mL) was incubated with isolated human MPO (30 nM), glucose (100 μM), glucose oxidase (20 ng/mL), and NO2– (500 μM) in sodium phosphate buffer (50 mM, pH 7.0) supplemented with DTPA (100 μM) (complete system) or the indicated additions or deletions. Lipoproteins were then incubated with thioglycollate-elicited MPMs, and the extent of cholesteryl [14C]oleate formation (a), cellular cholesteryl ester mass (b), and free cholesterol mass (c) was determined as described in Methods. Data represent the mean ± SD of triplicate determinations. Similar results were observed in 3 independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001 for comparison vs. LDL modified by MPO and an H2O2-generating system only (– NO2–). No significant cholesteryl [14C]oleate formation or increase in cholesteryl ester or free cholesterol mass above basal values was observed in LDL preparations modified by the complete system in the absence of either MPO or GGOx. Hi catalase, heat-inactivated catalase.