GLYCEROLIPIDS AND PROSTAGLANDIN BIOSYNTHESIS

甘油脂和前列腺素生物合成

• 批准号: 6178638
• 负责人: ATA A ABDEL-LATIF
• 金额: $ 19.92万
• 依托单位: AUGUSTA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1981
• 资助国家: 美国
• 起止时间: 1981-12-01 至 2002-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6178638
• 关键词: adrenergic receptor; arachidonate; cats; eicosanoid metabolism; enzyme activity; eye pharmacology; fatty acid biosynthesis; human tissue; iris; laboratory rabbit; muscarinic receptor; neuroregulation; phospholipase A2; phospholipase C; phospholipase D; prostaglandins; protein kinase C; protein purification; pupil; radiotracer; receptor sensitivity; second messengers; smooth muscle; stimulant /agonist; uvea ciliary body

DESCRIPTION (Investigator's Abstract): The long-term goal of this research is to elucidate the regulatory mechanisms involved in arachidonic acid (AA) liberation and eicosanoid biosynthesis in the iris-ciliary body (ICB) and other ocular tissues. The central hypothesis to be tested is that phospholipase A2 (PLA2) activation is a major source for AA release in the ICB. This represents a logical pursuit of the findings obtained during the current budget period. ICB is the major source of eicosanoid synthesis in the eye. The investigators and others have reported that activation of the ICB with various agonists, including carbachol, PGE2alpha, substance P, endothelin-1, and PAF results in the rapid hydrolysis of polyphosphoinositides into IP3 and DAG, the two second messengers, and the concomitant liberation from membrane phospholipids of AA for eicosanoid biosynthesis. The investigators have found that the generation of these phospholipid-derived second messengers and the formation of cAMP and their physiological consequences to be species dependent. While the involvement of phospholipases (A2,C,D) in AA release has been documented in several tissues, including ICB, the precise mechanism underlying the stimulated release of this polyunsaturated fatty acid remains undefined. This is also true of the phospholipid source of AA. There is strong evidence from the investigators own laboratory on the ICB and from other investigators working with different tissues that PLA2, which acts on phospholipids to liberate AA and lysophospholipid, plays a central role in the control of AA required for eicosanoid biosynthesis. Based on these findings the investigators now propose to address the following specific aims: (1) Investigate the AA content of the major membrane phospholipids, phospholipases (A2,C,D) activities, and the effects of various agonists on AA release and eicosanoid biosynthesis in ICB and trabecular meshwork of different mammalian species. (2) Investigate the mechanisms involved in agonist-induced AA release and phospholipid source of AA in the iris sphincter, and AA release from phospholipids in membranes isolated from iris sphincters. Endogenous release of AA will be monitored with GLC. (3) Investigate the regulatory role of protein kinase C in AA release and PG synthesis in the iris sphincter. Both PKC inhibitors and down-regulation of PKC with PDBu will be employed. (4) Purify and characterize phospholipase(s) A2 from rabbit ICB. Activators and inhibitors of PLA2 will be employed. An attempt will be made to reconstitute the purified PLA2 with G-proteins. (5) Investigate the mechanism of the effects of in vitro and in vivo PG receptor desensitization on basal and on agonist-induced AA release and PG synthesis, IP3 production, cAMP formation, and on contraction in the iris sphincter. The proposed studies will yield important basic information about the physiological regulation of AA release from membrane phospholipids, the link between receptor activation and AA release, the profile of PGs produced in response to each agonist, the mechanism of PG receptor desensitization and its cross desensitization of other receptors, and the role of phospholipases (A2,C,D) in stimulus- dependent release of AA in ocular tissues. Understanding the biochemical basis for the regulation of AA liberation and PG synthesis will make their pharmacological manipulation more plausible, and will lead to the development of more effective anti-glaucoma and anti-inflammatory drugs.

描述（研究者摘要）：本研究的长期目标 旨在阐明花生四烯酸（AA）的调节机制 虹膜睫状体 (ICB) 中的释放和类二十烷酸生物合成 其他眼组织。 要检验的中心假设是 磷脂酶 A2 (PLA2) 的激活是 AA 释放的主要来源 国际商业银行。 这代表了对研究期间获得的发现的逻辑追求 当前预算期。 ICB是类二十烷酸合成的主要来源 眼睛。 研究人员和其他人报告说，激活 ICB 与各种激动剂，包括卡巴胆碱、PGE2α、P 物质、 内皮素-1和PAF导致快速水解 多磷酸肌醇转化为 IP3 和 DAG（两个第二信使） AA 的膜磷脂同时释放类二十烷酸 生物合成。 调查人员发现，这些物质的产生 磷脂衍生的第二信使和cAMP的形成及其 生理后果取决于物种。 在参与的同时 AA 释放中磷脂酶 (A2、C、D) 的变化已在多个文献中得到记录 组织，包括 ICB，刺激背后的精确机制 这种多不饱和脂肪酸的释放仍不确定。 这也是 AA 的磷脂来源也是如此。有强有力的证据来自 研究人员在 ICB 上拥有自己的实验室以及其他研究人员的实验室 PLA2 与不同的组织一起工作，PLA2 作用于磷脂， 释放 AA 和溶血磷脂，在 AA 的控制中发挥核心作用 类二十烷酸生物合成所需。基于这些发现 研究人员现在建议解决以下具体目标：（1） 研究主要膜磷脂的 AA 含量， 磷脂酶（A2、C、D）活性以及各种激动剂对 ICB 和小梁网中的 AA 释放和类二十烷酸生物合成 不同的哺乳动物物种。 (2) 研究相关机制 虹膜中激动剂诱导的 AA 释放和 AA 的磷脂来源 括约肌和 AA 从分离的膜中的磷脂中释放 虹膜括约肌。 AA 的内源释放将通过 GLC 进行监测。 (3) 研究蛋白激酶 C 在 AA 释放和 PG 中的调节作用 在虹膜括约肌中合成。 PKC 抑制剂和下调 将采用带有 PDBu 的 PKC。 (4) 纯化与表征 来自兔 ICB 的磷脂酶 A2。 PLA2 的激活剂和抑制剂 将被雇用。将尝试重构纯化的 PLA2 与G蛋白。 (5) 研究体外作用机制 以及体内 PG 受体对基础和激动剂诱导的 AA 脱敏 释放和 PG 合成、IP3 产生、c​​AMP 形成等 虹膜括约肌收缩。 拟议的研究将产生 关于AA生理调节的重要基础信息 膜磷脂的释放，受体激活之间的联系 和 AA 释放，响应每种激动剂产生的 PG 概况， PG受体脱敏及其交叉脱敏机制 其他受体的作用，以及磷脂酶（A2、C、D）在刺激中的作用 眼组织中 AA 的依赖性释放。 了解生化 AA 释放和 PG 合成调节的基础将使其 药物操作更合理，并且会导致 开发更有效的抗青光眼和抗炎药物。