CYTOCHROME P450 DEPENDENT ARACHIDONIC ACID METABOLISM

描述(逐字摘自申请者摘要)：两条主要途径 细胞色素P450(CYP)催化的花生四烯酸(AA)代谢 欧米伽-羟基化生成20-羟基二十碳四烯酸(20-HETE)和 产生区域和立体异构体环氧二十碳三烯酸的环氧化反应 (EETS)。EETs被环氧化物水解酶进一步代谢成 相应的二羟基二十碳三烯酸(DHETs)。这些由CYP派生的 二十烷类化合物令人感兴趣，因为它们是许多 组织，具有广泛的生物效应。特定的细胞 酶的表达模式、相对丰度和活性 催化这些反应将是细胞内的一个主要决定因素 这些二十烷类化合物的作用。在肾脏中，CYP衍生的二十烷类化合物具有强大的 对肾血管张力和肾小管离子转运的影响 牵涉到血压的调节。该计划的总体目标 提出的研究是为了了解控制的分子机制 CYP催化的大鼠肾脏AA代谢及肾脏CYP的重要性 二十烷类化合物在血压调节中的水平。《公约》的具体目标 建议的研究是：(1)研究 抑制AA omega-羟基酶活性的降压作用。 具体地说，我们将确定异构体的特异性和效力 基于机制的CYP AA omega-羟基酶抑制剂及其评价 对血压和血管张力的影响；(2)确定其贡献 细胞色素P4E亚型与肾脏AA代谢有关。CYP4F表达将本地化 研究了大鼠肾脏和氨基酸在细胞色素P4F异构体中的代谢； (3)分离大鼠肾脏表达的主要环氧合酶。这个 编码细胞色素P450 2J2免疫活性蛋白的c DNA在 自发性高血压大鼠肾脏将通过表达克隆进行鉴定 以及(4)研究EET的调节 大鼠肾脏中的水解物。具体地说，我们将确定生化 EET在大鼠肾微粒体内的水解性基础及可溶性的影响 环氧化物水解酶对血压和肾脏二十烷类化合物形成的抑制作用。 这些研究的结果将导致对 细胞色素P450和环氧化物水解酶的表达与调控 参与AA新陈代谢。这些研究的一个长期目标是发展 调节肾细胞胞二磷酸二十糖类化合物的新型靶向治疗药物 生产和血压。将这些原则应用于监管 预计在其他组织中也会形成CYP二十烷类化合物。

DESCRIPTION (Verbatim from Applicant's Abstract): The two major pathways of cytochrome P450 (CYP)-catalyzed arachidonic acid (AA) metabolism are omega-hydroxylation to form 20-hydroxyeicosatetraenoic acid (20-HETE) and epoxidation which produces regio- and stereoisomeric epoxyeicosatrienoic acids (EETs). The EETs are further metabolized by epoxide hydrolases to the corresponding dihydroxyeicosatrienoic acids (DHETs). These CYP-derived eicosanoids are of interest since they are endogenous constituents of numerous tissues and posses a wide array of biological effects. The cell specific pattern of expression, relative abundance, and activity of the enzymes catalyzing these reactions will be a major determinant of the intracellular effect of these eicosanoids. In the kidney, CYP-derived eicosanoids have potent effects on renal vascular tone and tubular ion transport and have been implicated in the regulation of blood pressure. The overall goal of the proposed studies is to understand the molecular mechanisms controlling CYP-catalyzed AA metabolism in the rat kidney and the importance of renal CYP eicosanoid levels in blood pressure regulation. The specific aims of the proposed studies are (1) to examine the mechanistic basis of the antihypertensive effect of inhibition of AA omega-hydroxylase activity. Specifically, we will determine the isoform-specificity and potency of mechanism-based inhibitors of CYP AA omega-hydroxylases and evaluate their effect on blood pressure and vascular tone; (2) to determine the contribution of CYP4E isoforms to renal AA metabolism. CYP4F expression will be localized in the rat kidney and AA metabolism by the CYP4F isoforms will be characterized; (3) to isolate the major CYP2J epoxygenase expressed in the rat kidney. The cDNA encoding the CYP2J2 immunoreactive protein overexpressed in the spontaneously hypertensive rat kidney will be identified by expression cloning and functionally characterized; and (4) to examine the regulation of EET hydrolysis in the rat kidney. Specifically, we will determine the biochemical basis of EET hydrolysis in rat renal microsomes and the effect of soluble epoxide hydrolase inhibition on blood pressure and renal eicosanoid formation. The findings from these studies will lead to a comprehensive understanding of the expression and regulation of the major CYP and epoxide hydrolase enzymes involved in AA metabolism. A long term goal of these studies is to develop novel targeted therapeutics for the regulation of renal CYP eicosanoid production and blood pressure. Application of these principles for regulating CYP eicosanoid formation in other tissues is anticipated.