RESUMEN
Oxidized low-density lipoprotein (LDL) is a major component in the pathophysiology of atherosclerosis and plays a role in the changes of vascular reactivity observed in this disease. Herein the authors investigate the potential involvement of platelet-activating factor (PAF)-like phospholipid components of oxidized LDL in rabbit aorta reactivity. Aortic rings were precontracted with noradrenaline (0.5 microM) and relaxation was induced by subsequent stimulation with sequential additions of acetylcholine (1 nM to 3 microM). High-performance liquid chromatography (HPLC) fractions (6- and 7-min) obtained from phospholipids extracted from oxidized LDL inhibited relaxation evoked by acetylcholine, but not the relaxation induced by sodium nitroprusside. This effect was not antagonized either by incubation of the fractions with PAF acetylhydrolase or by incubation of the aortic rings with a PAF receptor antagonist. Authentic PAF or C4-PAF, a PAF mimetic previously found in fractions 6 and 7 did not inhibit acetylcholine-induced relaxation. In contrast, lyso-PAF inhibited acetylcholine, but not sodium nitroprusside-induced relaxation. The authors conclude that phospholipids of oxidized LDL impair vascular reactivity to endothelium-dependent agonists. This effect is not due to oxidatively generated proinflammatory PAF mimetics, but rather to a metabolite of these phospholipids, lysoPAF.
Asunto(s)
Aorta/fisiología , Células Endoteliales/metabolismo , Lipoproteínas LDL/metabolismo , Músculo Liso Vascular/metabolismo , Fosfolípidos/metabolismo , Vasodilatación/fisiología , 1-Alquil-2-acetilglicerofosfocolina Esterasa/farmacología , Acetilcolina/farmacología , Animales , Aorta/efectos de los fármacos , Aterosclerosis/metabolismo , Aterosclerosis/fisiopatología , Células Endoteliales/efectos de los fármacos , Femenino , Humanos , Lipoproteínas LDL/química , Masculino , Músculo Liso Vascular/efectos de los fármacos , Nitroprusiato/farmacología , Norepinefrina/farmacología , Técnicas de Cultivo de Órganos , Fosfolípidos/farmacología , Factor de Activación Plaquetaria/análogos & derivados , Factor de Activación Plaquetaria/metabolismo , Factor de Activación Plaquetaria/farmacología , Glicoproteínas de Membrana Plaquetaria/antagonistas & inhibidores , Glicoproteínas de Membrana Plaquetaria/metabolismo , Conejos , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Receptores Acoplados a Proteínas G/metabolismo , Vasoconstrictores/farmacología , Vasodilatación/efectos de los fármacos , Vasodilatadores/farmacologíaRESUMEN
Oxidized low density lipoprotein (LDL) has an important proinflammatory role in atherogenesis. In this study, we investigated the ability of oxidized LDL (oxLDL) and its phospholipid components to induce lipid body formation in leukocytes. Incubation of mouse peritoneal macrophages with oxidized, but not with native LDL led to lipid body formation within 1 h. This was blocked by platelet-activating factor (PAF) receptor antagonists or by preincubation of oxLDL with rPAF acetylhydrolase. HPLC fractions of phospholipids purified from oxLDL induced calcium flux in neutrophils as well as lipid body formation in macrophages. Injection of the bioactive phospholipid fractions or butanoyl and butenoyl PAF, a phospholipid previously shown to be present in oxLDL, into the pleural cavity of mice induced lipid body formation in leukocytes recovered after 3 h. The 5-lipoxygenase and cyclooxygenase-2 colocalized within lipid bodies formed after stimulation with oxLDL, bioactive phospholipid fractions, or butanoyl and butenoyl PAF. Lipid body formation was inhibited by 5-lipoxygenase antagonists, but not by cyclooxygenase-2 inhibitors. Azelaoyl-phosphatidylcholine, a peroxisome proliferator-activated receptor-gamma agonist in oxLDL phospholipid fractions, induced formation of lipid bodies at late time points (6 h) and synergized with suboptimal concentrations of oxLDL. We conclude that lipid body formation is an important proinflammatory effect of oxLDL and that PAF-like phospholipids and peroxisome proliferator-activated receptor-gamma agonists generated during LDL oxidation are important mediators in this phenomenon.
Asunto(s)
Cuerpos de Inclusión/metabolismo , Leucocitos/metabolismo , Lipoproteínas LDL/metabolismo , Peroxisomas/metabolismo , Factor de Activación Plaquetaria/fisiología , Receptores Citoplasmáticos y Nucleares/agonistas , Receptores Citoplasmáticos y Nucleares/fisiología , Factores de Transcripción/agonistas , Factores de Transcripción/fisiología , Animales , Araquidonato 5-Lipooxigenasa/metabolismo , Araquidonato 5-Lipooxigenasa/fisiología , Ciclooxigenasa 2 , Sinergismo Farmacológico , Femenino , Humanos , Isoenzimas/metabolismo , Leucocitos/fisiología , Lipoproteínas LDL/administración & dosificación , Lipoproteínas LDL/farmacología , Macrófagos Peritoneales/metabolismo , Masculino , Proteínas de la Membrana , Ratones , Oxidación-Reducción , Peroxisomas/enzimología , Peroxisomas/fisiología , Fosfolípidos/metabolismo , Factor de Activación Plaquetaria/administración & dosificación , Factor de Activación Plaquetaria/metabolismo , Cavidad Pleural/citología , Prostaglandina-Endoperóxido Sintasas/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Cavidad Torácica , Factores de Transcripción/metabolismoRESUMEN
Oxidized low-density lipoprotein (LDL) contains inflammatory agents, including oxidatively fragmented phospholipids that activate the platelet-activating factor (PAF) receptor, but in vivo events caused by these pathologically generated agents are not well defined. Injection of PAF-like lipids derived from oxidized LDL, or C(4)-PAF that is a major PAF-like lipid in these particles, into the pleural cavity of mice resulted in rapid monocyte, neutrophil, and eosinophil accumulation. Increased numbers of intracellular lipid bodies in these cells show they were in an inflammatory environment. Leukocyte recruitment was abolished by a PAF receptor antagonist, as expected. PAF-like lipids induced 5-lipoxygenase expression in leukocytes, mRNA expression for monocyte chemoattractant protein-1 (MCP-1) and other chemokines, synthesis of MCP-1, and leukotriene B(4). The 5-lipoxygenase inhibitor zileuton impaired neutrophil influx, while MCP-1 had a more global role, as determined with MCP-1(-/-) mice. The lack of MCP-1 abrogated leukocyte accumulation and lipid body formation both in vivo and in vitro and chemokine transcription in vivo, and reduced in vivo leukotriene B(4) production. Thus, PAF-like phospholipids in oxidized LDL induce an inflammatory infiltrate through the PAF receptor, chemokine transcription, lipid body formation, and 5-lipoxygenase expression in leukocytes. MCP-1 has a key role in this inflammatory response, and 5-lipoxygenase products are essential for neutrophil recruitment into the inflamed pleural cavity.