NEURONAL MEMBRANE LIPID OXIDATION IN PARKINSONS DISEASE

帕金森病中的神经元膜脂质氧化

• 批准号: 2051757
• 负责人: GOVIND N.T. VATASSERY
• 金额: $ 14.28万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-08-01 至 1998-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2051757
• 关键词: Parkinson's disease; age difference; cellular pathology; cholesterol; human subject; human tissue; laboratory rat; lipid metabolism; lipid peroxides; mitochondrial membrane; oxidation; oxidative stress; platelets; synaptosomes; tocopherols

This project is designed to test the hypothesis that specific areas of the brain such as substantia nigra and basal ganglia are especially vulnerable to oxidative stress and that their response to biochemical oxidative reactions (or oxidative stress) is defective in Parkinsons disease (PD) as well as in normal aging. Mitochondrial and synaptic membranes may lose their function due to inadequate response to oxidation an-or from direct damage induced by cytotoxic products of oxidation (e.g., cholesterol oxidation products). Levels of oxidation products and loss of function could be small under steady state conditions. Therefore, we will perturb the system by inducing oxidation of membranes in vitro (using primarily endogenous oxidants) in order to amplify existing differences. Our preliminary studies show that mitochondrial cholesterol is more oxidizable in PD than in controls. Mitochondrial and synaptosomal fractions will be oxidized by incubation with oxidizing agents (e.g., arachidonic and linoleic acid hydroperoxide plus metal ions like iron, mixtures of iron and ascorbic acid, nitric oxide with and without superoxide etc.). Different levels of oxidation of membranes can be defined by the extent of oxidation of two membrane lipids - tocopherol and cholesterol. Since the profile of cholesterol and tocopherol oxidation products would be characteristic for various types of oxidations, information on the mechanism of oxidation will be obtained. Synaptosomes and mitochondria from young and old rats, and PD and control human autopsy brain samples will be compared in all studies. Similar studies on oxidation of platelets (as an accessible model of neuronal membranes in humans) from PD and control subjects will be conducted. The extent of oxidation of membranes will be monitored and quantitated by determining the concentrations of tocopherol and cholesterol and their oxidation products as well as by estimating thiobarbituric acid reactive substances and the decline in total suIfhydryl concentrations. Oxidizing conditions which yield maximal differences between the control and experimental groups will be established first and then the cholesterol and tocopherol oxidation products will be isolated, identified, and quantitated by GC and GC-MS. A study of the effect of monoamine oxidase(MAO) on mitochondrial oxidation will be done by incubating mitochondria with dopamine, norepinephrine or serotonin (substrates for MAO) which releases in situ hydrogen peroxide and/or other oxidants derived from it. The oxidation will be followed and oxidation products characterized as with brain fraction& The effect of MAO inhibition by deprenyl upon the oxidation will also be studied. Number of similarities exist between the uptake of serotonin by platelets and brain. Serotonin uptake by platelets and synaptosomes at various levels of oxidation will be studied to examine the effect of oxidation upon membrane function. The data will be used to determine whether membrane damage induced by oxidative stress exists in PD and/or in normal aging. The data from the platelet studies may also provide a laboratory tool for the preclinical diagnosis of PD.

该项目旨在检验以下假设： 黑质和基底神经节等大脑尤其脆弱 氧化应激及其对生化氧化的反应 帕金森病 (PD) 反应（或氧化应激）存在缺陷，如 以及正常衰老过程中的情况。线粒体和突触膜可能会丢失 它们的功能是由于对氧化反应不充分或直接 细胞毒性氧化产物（例如胆固醇）引起的损伤 氧化产物）。氧化产物的水平和功能丧失 在稳态条件下可能很小。因此，我们会扰乱 该系统通过体外诱导膜氧化（主要使用 内源性氧化剂）以放大现有的差异。我们的 初步研究表明线粒体胆固醇更易氧化 PD 中的值高于对照组。 线粒体和突触体部分将通过孵育被氧化 与氧化剂（例如花生四烯酸和亚油酸氢过氧化物 加上金属离子，如铁、铁和抗坏血酸的混合物、一氧化氮 有或没有超氧化物等）。不同程度的氧化 膜可以通过两种膜脂的氧化程度来定义 - 生育酚和胆固醇。由于胆固醇和 生育酚氧化产物是各种类型的特征 氧化，将获得有关氧化机制的信息。 来自年轻和年老大鼠的突触体和线粒体，以及 PD 和对照 所有研究都将比较人体尸检大脑样本。相似的 血小板氧化的研究（作为神经元的可访问模型） 将对来自PD和对照受试者的人体膜进行研究。这 膜的氧化程度将通过以下方式进行监测和定量 测定生育酚和胆固醇的浓度及其 氧化产物以及通过估计硫代巴比妥酸反应性 物质和总巯基浓度的下降。氧化性 控制和控制之间产生最大差异的条件 首先建立实验组，然后建立胆固醇和 生育酚氧化产物将被分离、鉴定和定量 通过 GC 和 GC-MS。单胺氧化酶（MAO）对细胞的影响研究 线粒体氧化将通过将线粒体与 多巴胺、去甲肾上腺素或血清素（MAO 的底物）会释放 原位过氧化氢和/或由其衍生的其他氧化剂。这 随后将进行氧化，氧化产物的特征为 脑部分&丙炔苯丙胺抑制MAO对氧化的影响 也将被研究。的吸收之间存在许多相似之处 血小板和大脑产生血清素。血小板摄取血清素和 将研究不同氧化水平的突触体以检查 氧化对膜功能的影响。该数据将用于 确定PD中是否存在氧化应激诱导的膜损伤 和/或在正常老化中。血小板研究的数据也可以提供 用于 PD 临床前诊断的实验室工具。