Activation of NADPH oxidase required for macrophage-mediated oxidation of low-density lipoprotein.
Low-density lipoprotein (LDL) oxidation by arterial wall cells, a key event during early atherogenesis, was suggested to involve the activation of 15-lipoxygenase and/or nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. We sought to analyze the role of these oxygenases in macrophage-mediated oxidation of LDL under oxidative stress. Upon incubation of LDL with the J-774 A.1 macrophage-like cell line or with human monocyte-derived macrophages (HMDM) in the presence of 1 micromol/L CuSO4, the release of superoxide anions to the medium was demonstrated. Under these conditions, the cytosolic protein components of the NADPH oxidase complex, P-47 and P-67, translocated to the plasma membrane, indicating LDL-mediated activation of the NADPH oxidase complex. Under the above-mentioned experimental conditions, the macrophage 15-lipoxygenase was also activated, as determined by the release of 15-hydroxy-5,8,11,13-eicosatetraenoic acid (15-HETE) and 13-hydroxyoctadecadienoic acid (13-HODE) to the medium. Inhibition of the macrophage NADPH oxidase with apocynin or dismutation of superoxide anions, the product of NADPH oxidase activation, with superoxide dismutase (SOD) significantly inhibited macrophage-mediated oxidation of LDL (by 61% to 89%) under these conditions. Phorbol myristate acetate (PMA), which causes NADPH oxidase activation in J-774 A.1 macrophages, had no significant effect on 15-lipoxygenase activity, but still resulted in cell-mediated oxidation of LDL. Finally, HMDM from two patients with chronic granulomatous disease (CGD) that were shown to lack active NADPH oxidase, but to possess almost normal 15-lipoxygenase activity failed to oxidize LDL. We thus conclude that LDL-induced NADPH oxidase activation (under oxidative stress) is required for macrophage-mediated oxidation of LDL, whereas activation of 15-lipoxygenase may not be sufficient for LDL oxidation under these conditions.
Aviram
M
M
Lipid Research Laboratory, Rambam Medical Center, Haifa, Israel.
Rosenblat
M
M
Etzioni
A
A
Levy
R
R
D000293:Adolescent; D001093:Arachidonate 15-Lipoxygenase / Q000378:metabolism; D002648:Child; D004789:Enzyme Activation; D006105:Granulomatous Disease, Chronic / Q000097:blood; D006801:Humans; D008077:Lipoproteins, LDL / Q000378:metabolism*; D008264:Macrophages / Q000201:enzymology / Q000378:metabolism*; D008297:Male; D009000:Monocytes / Q000201:enzymology / Q000378:metabolism; D009247:NADH, NADPH Oxidoreductases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019255:NADPH Oxidases; D010084:Oxidation-Reduction
D008077:Lipoproteins, LDL; D001093:Arachidonate 15-Lipoxygenase; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases