OXIDATION-INDUCED MEMBRANE DAMAGE IN CATARACTOGENESIS

白内障发生过程中氧化引起的膜损伤

• 批准号: 2158595
• 负责人: KAILASH C BHUYAN
• 金额: $ 26.7万
• 依托单位: COLUMBIA UNIV NEW YORK MORNINGSIDE
• 依托单位国家: 美国
• 财政年份: 1978
• 资助国家: 美国
• 起止时间: 1978-09-01 至 1996-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2158595
• 关键词: aging; aminoacid analyzer; antioxidants; calcium transporting ATPase; cataract; disulfide bond; eye disorder chemotherapy; fluorimetry; free radical oxygen; gel electrophoresis; genetic strain; high performance liquid chromatography; human tissue; hydrogen peroxide; hydroxyl radical; ion transport; laboratory mouse; laboratory rabbit; lens proteins; lipid peroxides; membrane channels; membrane lipids; membrane permeability; membrane proteins; microspectrophotometry; oxidation; oxidative stress; pathogenic diet; phospholipids; polarography; radiotracer; sodium potassium exchanging ATPase; thin layer chromatography

Increase in cell membrane permeability to cations, is a common occurrence in cataracts, leading to intracellular ionic imbalance. Oxidation of protein occurs in human senile cataract, leading to crosslinking and the formation of high molecular weight crystallin aggregates. The peroxidation of membrane lipids may participate in both membrane permeability increases and protein oxidation. Lipids are more prone to oxidation than proteins, generating peroxide intermediates and free radicals that can initiate further oxidation of proteins. We hypothesize that oxygen-derived free radicals are the agents that trigger cataractogenesis by inducing peroxidation of membrane lipids, an early event in both membrane damage and protein oxidation. We will determine the sequence of appearance of relatively stable oxidants, hydrogen peroxide, lipid peroxide and the products of oxidation of cell membrane components in human donor normal lenses as a function of age, and in surgically extracted cataractous lenses by radioisotopic and enzyme-coupled spectrophotometric and spectrofluorometric techniques. Lipids and lipid peroxides will be identified by HPLC and their adducts by TLC. Adducts of lipid peroxidation products and proteins will be analysed by column chromatography and gel electrophoresis. Lipid repair mechanisms will be determined using radioactive probes. The formation of disulfide crosslinks in key plasma membrane proteins such as gap junction protein, MIP26 will be measured chromatographically and thiols spectrophotometrically. Na+, K+-activated and Ca2+-activated ATPases will be monitored biochemically for age-related loss in activity and oxidation. To test the hypothesis that oxidants will accelerate and antioxidants will retard the development of cataractous changes in the Emory mouse, a model of human senile cataract, we shall observe the effects of a low level of oxidative stress by feeding selenium in the diet, or an antioxidant, acetylsalicylate, in diet.

细胞膜对阳离子的渗透性增加是常见现象 在白内障中，导致细胞内离子失衡。氧化 人类老年性白内障中蛋白质发生交联， 高分子量晶状体蛋白聚集体的形成。 这 膜脂的过氧化可能参与两种膜 渗透性增加和蛋白质氧化。 血脂更容易发生 比蛋白质氧化，生成过氧化物中间体和游离 可以引发蛋白质进一步氧化的自由基。 我们假设 氧自由基是引发 通过诱导膜脂过氧化而发生白内障，这是一种早期的 膜损伤和蛋白质氧化的事件。 我们将确定 相对稳定的氧化剂氢出现的顺序 过氧化物、脂质过氧化物及细胞膜氧化产物 人类捐赠者正常晶状体中的成分随年龄变化，并且 通过放射性同位素手术摘除白内障晶状体 酶联分光光度和荧光分光光度技术。 脂质和脂质过氧化物将通过 HPLC 及其加合物进行鉴定 通过薄层色谱法。 脂质过氧化产物和蛋白质的加合物 通过柱色谱法和凝胶电泳进行分析。 脂质修复 将使用放射性探针确定机制。 的形成 关键质膜蛋白（例如间隙连接）中的二硫键交联 蛋白质、MIP26 和硫醇将通过色谱法进行测量 分光光度法。 Na+、K+ 激活和 Ca2+ 激活的 ATP 酶 将对与年龄相关的活动损失进行生化监测， 氧化。 检验氧化剂会加速和 抗氧化剂将延缓白内障的发展 埃默里小鼠，人类老年性白内障模型，我们将观察 通过饲喂硒来降低氧化应激水平的影响 饮食，或饮食中的抗氧化剂乙酰水杨酸盐。