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(Arteriosclerosis, Thrombosis, and Vascular Biology. 1997;17:3505-3512.)
© 1997 American Heart Association, Inc.

Articles

Copper-Catalyzed Oxidation Mediates PAF Formation in Human LDL Subspecies

Protective Role of PAF:Acetylhydrolase in Dense LDL

Demokritos C. Tsoukatos; Muriel Arborati; Theodoros Liapikos; Keith L. Clay; Robert C. Murphy; M. John Chapman; ; Ewa Ninio

From the Laboratory of Biochemistry, Department of Chemistry, University of Ioannina, 45110 Ioannina, Greece (D.C.T., T.L.); Institut National de la Santé et de la Recherche Medicale (INSERM), Unité de Recherches sur les Lipoprotéines et l'Athérogénèse, U-321, Pavillon Benjamin Delessert, Hôpital de la Pitié, Paris, France (M.A., M.J.C., E.N.); and National Jewish Medical and Research Center, Denver, CO (K.L.C., R.C.M.).

Correspondence to Ewa Ninio, INSERM U-321, Pavillon Benjamin Delessert, Hôpital de la Pitié, Bd de l' Hôpital, 75651 Paris, Cedex 13, France. E-mail: eninio{at}infobiogen.fr

Abstract Free radical-mediated oxidation of cholesterol-rich LDL plays a key role in atherogenesis and involves the formation of oxidized phospholipids with proinflammatory biological activity. We evaluated the production of platelet-activating factor (PAF), a potent inflammatory mediator, in human LDL subspecies on copper-initiated oxidation (4 µmol/L CuCl₂, 80 µg/mL for 3 hours at 37°C). PAF formation was determined by biological assay of HPLC-purified lipid extracts of copper-oxidized lipoproteins; chemical identity was confirmed by gas chromatographic and mass spectrometric analyses. PAF, characterized as the C16:0 molecular species, was preferentially produced in intermediate LDL (d=1.029 to 1.039 g/mL) (8.6±5.7 pmol PAF/3 h per mg LDL protein) and light LDL (d=1.019 to 1.029 g/mL), but was absent from dense LDL particles (d=1.050 to 1.063 g/mL). As PAF:acetylhydrolase inactivates PAF and oxidized forms of phosphatidylcholine, we evaluated the relationship of lipoprotein-associated PAF:acetylhydrolase to PAF formation. We confirmed that PAF:acetylhydrolase activity was elevated in native, dense LDL (41.5±9.5 nmol/min per mg protein) but low in LDL subspecies of light and intermediate density (d 1.020 to 1.039 g/mL) (3.5±1.6 nmol/min per mg protein) [Tselepis et al, Arterioscler Thromb Vasc Biol. 1995;15:1764–1773]. On copper-mediated oxidation for 3 hours at 37°C, dense LDL particles conserved 20±14% of their initial enzymatic activity; in contrast, PAF:acetylhydrolase activity was abolished in light and intermediate LDL subspecies. Clearly, the elevated PAF: acetylhydrolase activity of dense LDL efficiently diminishes the potential inflammatory role of endogenously formed PAF; nonetheless, formation of proatherogenic lysophospholipids results. In contrast, LDL particles of the light and intermediate subclasses can accumulate PAF on oxidative modification.

Key Words: inflammation • density gradient ultracentrifugation • lipoprotein particle subspecies

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