MASS SPECTROMETRIC IDENTIFICATION OF NEW LIPID MEDIATORS

新脂质介质的质谱鉴定

• 批准号: 6231365
• 负责人: MICHAEL BALAZY
• 金额: $ 19.48万
• 依托单位: NEW YORK MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-03-01 至 2006-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6231365
• 关键词: arachidonate; bacterial disease; chemical synthesis; cyclic GMP; electrospray ionization mass spectrometry; endotoxins; free radical oxygen; high performance liquid chromatography; infrared spectrometry; laboratory rat; lipopolysaccharides; membrane; membrane lipids; nitrogen oxides; nuclear magnetic resonance spectroscopy; oxidative stress; pathology; phospholipids; plasmalogens; protein degradation; technology /technique development; tissue /cell preparation; toxicant interaction

DESCRIPTION (Applicant's abstract): The overall aim of this application is to characterize new mechanisms of biological membrane injury resulting form infection with bacterial endotoxin (LPS). The role of free radicals in LPS-induced endotoxemia is becoming well established, however, little is known about the processes involved in the damage to biomembrane lipids by nitrogen dioxide (NO2) radical (a product of nitric oxide oxidation). We have developed a new methodology based on electrospray tandem mass spectrometry, which allows identification and quantification of specific lipid products formed by the reaction of NO2 with arachidonic acid. Preliminary studies have revealed that this reaction generates a complex mixture of lipids containing characteristic products: trans isomers of arachidonic acid and lipids containing nitrogen-carbon bond (nitroeicosanoids). In addition, plasmalogen phospholipids reacted with NO2, which resulted in complete removal of this group of lipids and generation of 1 -lyso-phospholipids. We hypothesize that increased production of NO generates NO2. which is a key radical that targets arachidonic acid and phospholipids. thereby causing membrane injury. Specific aims are to: 1) study relationships between the magnitude of trans-arachidonic acid generation and other markers of free radical damage (isoprostaglandin, nitrite/nitrate, nitrotyrosine); 2) examine the nitration of arachidonic acid and the effects of nitroeicosanoids on NO and cGMP levels in tissues; 3) characterize modifications of plasmalogen phospholipids in endotoxemia. In order to address the role of NO, L-NMA, a NO synthase inhibitor will be used to prevent generation of NO. Uric acid was shown to scavenge NO2, and thus serve as a good probe to study effects of NO2 in LPS-induced injury. Thus these probes will be used to determine the role of NO and NO2 in arachidonic acid isomerization, nitration and plasmalogen degradation. We anticipate that in the long term the proposed studies will provide a foundation for a rational design of new strategies for development of new drugs (inhibitors of lipid isomexization and nitration), and therapies to treat symptoms of sepsis related to the N02-induced cytotoxicity.

描述（申请人的摘要）：本申请的总体目标是 表征生物膜损伤的新机制 细菌内毒素（LPS）感染。自由基的作用 LPS 诱发的内毒素血症已得到广泛证实，但人们知之甚少 关于氮对生物膜脂质损伤的过程 二氧化氮 (NO2) 自由基（一氧化氮氧化的产物）。我们开发了 一种基于电喷雾串联质谱法的新方法，允许 特定脂质产物的鉴定和定量 NO2 与花生四烯酸的反应。初步研究表明 该反应产生一种复杂的脂质混合物，其中含有特征 产品：花生四烯酸反式异构体及含脂类 氮-碳键（硝基二十烷酸）。此外，缩醛磷脂 与 NO2 发生反应，从而完全去除这组脂质 和1-溶血磷脂的生成。我们假设增加了 NO 的产生会产生 NO2。这是针对花生四烯酸的关键激进分子 酸和磷脂。从而造成膜损伤。具体目标是： 1）研究反式花生四烯酸大小之间的关系 自由基损伤的产生和其他标志物（异前列腺素， 亚硝酸盐/硝酸盐、硝基酪氨酸）； 2) 考察花生四烯酸的硝化情况 以及硝基二十烷酸对组织中 NO 和 cGMP 水平的影响； 3） 表征内毒素血症中缩醛磷脂的修饰。在 为了解决 NO 的作用，L-NMA 将使用 NO 合酶抑制剂 防止NO的产生。尿酸被证明可以清除 NO2，从而发挥作用 作为研究 NO2 在 LPS 诱导损伤中的作用的良好探针。因此这些 探针将用于确定 NO 和 NO2 在花生四烯酸中的作用 异构化、硝化和缩醛磷脂降解。我们预计在 从长远来看，拟议的研究将为合理设计提供基础 新药开发新策略（脂质抑制剂） 异构化和硝化），以及治疗脓毒症相关症状的疗法 NO2 诱导的细胞毒性。