BIOLOGICAL ACTIVITY OF OXIDIZED PHOSPHOLIPIDS

氧化磷脂的生物活性

• 批准号: 2221532
• 负责人: Thomas M McIntyre
• 金额: $ 21.11万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-05-01 至 1995-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2221532
• 关键词: aldehydes; arachidonate; blood chemistry; chemical models; chemical structure function; high performance liquid chromatography; human subject; laboratory rabbit; leukocyte activation /transformation; macrophage; mass spectrometry; neutrophil; oxidation; ozone; pathologic process; phosphatidylcholines; platelet activating factor; receptor; vascular endothelium; xanthine oxidase

The overall goal of this project is to identify and characterize the members of a novel class of biologically active molecules, oxidatively- fragmented phospholipids. We have discovered that oxidation of a prevalent cellular phospholipid generates potent (sub micromolar) activators of leukocyte function. Oxidation of phosphatidylcholine that contains an arachidonic acid residue in the second position, can fragment the arachidonoyl residue at double bonds to create shorter residue. Oxidation can be initiated either enzymatically, by an enzyme (xanthine oxidase) thought to play a major role in a variety of pathological states, or by a well characterized chemical reaction, ozonolysis. We use ozonolysis to generate a single, defined oxidized phospholipid that has an sn-2 residue that is only 5 carbon atoms long terminated with an aldehyde group (5- aldehydic-phosphatidylcholine or [5-AI]PC). We have characterized this product by a variety of physical techniques, so [5-AI]PC serves as our model compound. We have observed that the oxidation of synthetic phosphatidylcholine, or even human serum, byu xanthine oxidase generates new products that are biologically active, like [5-AI]PC itself. The fragment phosphatidylcholines, but not the intact molecule, stimulated neutrophil function and induced hypotension in a rabbit. Furthermore, it appears that the activation is receptor-mediated in that the oxidatively fragmented phosphatidylcholines activated a receptor that previously been considered to be specific for Platelet-activating Factor (PAF is an ether phosphatidylcholine with a short sn-2 acetate residue). The reaction that creates these oxidized phospholipids is an unregulated one that occurs in a variety of pathologic processes such as: injury due to reperfusion of ischemic heart, gut, brain and kidney; hemorrhagic shock; adult respiratory distress syndrome; inhalation injury; and, asbestosis. Oxidative reactions play a significant role in these processes as inhibition of xanthine oxidase, or removing or preventing oxidizing free radicals ameliorates tissue damage. Finally, PAF has been proposed to be involved based on the ability of PAF receptor antagonists to ameliorate these conditions. However, authentic PAF has not been detected in these pathological processes. Since all of these conditions involve oxidative processes, we propose that oxidatively fragmented phosphatidylcholines may be the actual mediators that activate leukocytes to increase tissue damage. In support of this, we find that oxidant-treated human endothelial cells bind PMN by an unknown mechanism that does not involve the synthesis of authentic PAF. There are three specific aims. 1) To identify oxidatively fragmented phosphatidylcholines, other than [5-AI]PC, that can activate leukocytes. We will isolate and identify the products of oxidation of synthetic phospholipids that stimulate leukocytes by a variety of physical methods, especially mass spectrometry. 2) To determine if a single receptor recognizes PAF and the various oxidatively fragmented phospholipids. This will use several structurally unrelated PAF receptor antagonists, and will take advantage of homologous desensitization of the PAF receptor by its agonists. 3) To determine if bioactive oxidatively-fragmented phospholipids occur in plasma, living cells nad tissue. This will rely on HPLC separation and identification of individual species by the methods developed in Aim 1.

该项目的总体目标是识别和表征 一类新型生物活性分子的成员，氧化 碎磷脂。 我们发现普遍存在的氧化 细胞磷脂会产生有效的（亚微摩尔）活化剂 白细胞功能。 磷脂酰胆碱的氧化 蛛网膜酸残基在第二个位置可以碎片 双键的蛛网残留物可产生较短的残留物。 氧化 可以通过酶（黄嘌呤氧化酶）从酶上启动 被认为在各种病态状态中发挥了重要作用，或 化学反应良好，臭氧解析。 我们使用臭氧溶解 产生一个具有SN-2残基的单个定义的氧化磷脂 长期以来，只有5个碳原子，该原子长期以醛组终止（5-- 醛 - 磷脂酰胆碱或[5-AI] PC）。 我们已经描述了这个 产品通过各种物理技术，因此[5-ai] PC用作我们的 模型化合物。 我们已经观察到合成磷脂酰胆碱的氧化或 甚至人血清，BYU黄嘌呤氧化酶也会生成新产品 生物学活跃，例如[5-ai] PC本身。 碎片 磷脂酰胆碱，但没有完整的分子刺激中性粒细胞 兔子的功能和诱发性低血压。 此外，看来 激活是受体介导的，因为氧化碎片 磷脂酰胆碱激活了以前被认为的受体 特定于血小板激活因子（PAF是一个以太 磷脂酰胆碱具有短Sn-2乙酸盐残基）。 产生这些氧化磷脂的反应是不受管制的 发生在多种病理过程中的一种，例如： 再灌注缺血性心脏，肠道，脑和肾脏；出血性休克； 成人呼吸窘迫综合征；吸入损伤；并且，石棉病。 氧化反应在这些过程中起着重要作用 抑制黄嘌呤氧化酶，或去除或防止氧化无氧化 自由基改善组织损伤。 最后，已提议PAF为 根据PAF受体拮抗剂改善的能力参与 这些条件。 但是，在这些中尚未检测到真实的PAF 病理过程。由于所有这些条件都涉及氧化 过程，我们建议氧化碎片的磷脂酰胆碱可能 是激活白细胞以增加组织损伤的实际介体。 为此，我们发现经过氧化剂处理的人内皮细胞 通过不涉及合成的未知机制结合PMN 正宗的PAF。 有三个特定的目标。 1）识别氧化碎片的 除[5-AI] PC以外，可以激活白细胞的磷脂酰胆碱。 我们将隔离并确定合成的氧化产物 通过多种物理方法刺激白细胞的磷脂， 特别是质谱。 2）确定是否单个受体 识别PAF和各种氧化碎片的磷脂。 这 将使用几个结构无关的PAF受体拮抗剂，并将 利用PAF受体的同源脱敏 激动剂。 3）确定生物活性是否氧化碎片 磷脂发生在血浆，活细胞NAD组织中。 这将依靠 通过这些方法对单个物种的分离和鉴定 在目标1中开发。