Watson A D, Subbanagounder G, Welsbie D S, Faull K F, Navab M, Jung M E, Fogelman A M, Berliner J A
Department of Medicine, University of California, Los Angeles, California 90095, USA.
J Biol Chem. 1999 Aug 27;274(35):24787-98. doi: 10.1074/jbc.274.35.24787.
One of the earliest steps in the development of the atherosclerotic lesion is the accumulation of monocyte/macrophages within the vessel wall. Oxidized lipids present in minimally modified-low density lipoproteins (MM-LDL) contribute to this process by activating endothelial cells to express monocyte-specific adhesion molecules and chemoattractant factors. A major focus of our group has been the isolation and characterization of the biologically active oxidized lipids in MM-LDL. We have previously characterized three oxidized phospholipids present in MM-LDL, atherosclerotic lesions of fat fed rabbits, and autoxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (Ox-PAPC) that induced human aortic endothelial cells to adhere human monocytes in vitro. We have used sequential normal and reverse phase-high performance liquid chromatography to isolate various isomers of an oxidized phospholipid from autoxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine. The fatty acid in the sn-2 position of this biologically active isomer and its dehydration product was released by phospholipase A(2) and characterized. Hydrogenation with platinum(IV) oxide/hydrogen suggested a cyclic moiety, and reduction with sodium borohydride suggested two reducible oxygen-containing groups in the molecule. The fragmentation pattern produced by electrospray ionization-collision induced dissociation-tandem mass spectrometry was consistent with a molecule resembling an E-ring prostaglandin with an epoxide at the 5,6 position. The structure of this lipid was confirmed by proton nuclear magnetic resonance spectroscopy analysis of the free fatty acid isolated from the dehydration product of m/z 828.5. Based on these studies, we arrived at the structure of the biologically active oxidized phospholipids as 1-palmitoyl-2-(5, 6-epoxyisoprostane E(2))-sn-glycero-3-phosphocholine. The identification of this molecule adds epoxyisoprostanes to the growing list of biologically active isoprostanes.
动脉粥样硬化病变发展过程中最早出现的步骤之一是单核细胞/巨噬细胞在血管壁内的积聚。存在于轻度修饰的低密度脂蛋白（MM-LDL）中的氧化脂质通过激活内皮细胞表达单核细胞特异性黏附分子和趋化因子来促进这一过程。我们团队的一个主要研究重点是MM-LDL中生物活性氧化脂质的分离和表征。我们之前已经鉴定了存在于MM-LDL、高脂喂养兔的动脉粥样硬化病变以及自氧化的1-棕榈酰-2-花生四烯酰-sn-甘油-3-磷酸胆碱（Ox-PAPC）中的三种氧化磷脂，它们在体外可诱导人主动脉内皮细胞黏附人单核细胞。我们使用了连续的正相和反相高效液相色谱法从自氧化的1-棕榈酰-2-花生四烯酰-sn-甘油-3-磷酸胆碱中分离出一种氧化磷脂的各种异构体。这种生物活性异构体及其脱水产物的sn-2位脂肪酸通过磷脂酶A(2)释放并进行了表征。用二氧化铂/氢气氢化表明存在一个环状部分，用硼氢化钠还原表明分子中有两个可还原的含氧基团。电喷雾电离-碰撞诱导解离-串联质谱产生的碎片模式与一个类似于E环前列腺素且在5,6位有一个环氧基的分子一致。通过对从m/z 828.5的脱水产物中分离出的游离脂肪酸进行质子核磁共振光谱分析，证实了这种脂质的结构。基于这些研究，我们得出生物活性氧化磷脂的结构为1-棕榈酰-2-(5,6-环氧异前列腺素E(2))-sn-甘油-3-磷酸胆碱。该分子的鉴定将环氧异前列腺素添加到了不断增加的生物活性异前列腺素列表中。
