CHARACTERIZATION OF RENAL NONCYCLOOXYGENASE ARACHIDONATE METABOLISM

肾脏细胞色素P450依赖的花生四烯酸(AA)单加氧酶 现在被认为是生物医学上重要的AA的第三个分支 卡斯卡德。P450衍生的最广泛的特征 代谢物，环氧基和19/20-羟基氨基酸，已被认为与 控制全身和肾脏血管反应性等过程， 离子通量、激素信号与实验性心肌梗死的病理生理学 高血压。因此，我们和其他人的研究表明， 这一酶系统在肾脏和身体控制中的作用 动态平衡。然而，由于缺乏适当的分子工具和 技术、肾脏P450的生理意义及其部位和 其代谢物的作用方式仍有待明确定义。 项目2，结合细胞和器官生理学部分 这一计划项目的一部分，建议利用分子方法来 细胞和/或动物模型的发展和应用 P450基因特异的功能表型。我们将应用以下组合 重组DNA和生化技术对设计和施工的影响 高周转率，自给自足，区域和对映选择性AA 单加氧酶，这些酶在培养细胞中的表达，以及 依赖插入/cDNAs基因的鉴定及其机制研究 细胞表型。有针对性的基因破坏将被用于 全动物综合功能和生化分析 肾脏特异性AA环氧合酶和AFP的意义及作用(S) Omega/omega-1水解酶。这个项目的长期目标是 提供对肾脏P450二十烷类化合物生物学的分子理解 行动的意义和方式。这些重要问题的答案 是制定有意义的办法所必需的：a) 明确定义他们的生理和/或病理生理学 意义，以及b)随后的药理学和/或临床 干预措施。

The kidney cytochrome P450-dependent arachidonic acid (AA) monooxygenase is now recognized as the third branch of the biomedically important AA cascade. The most extensively characterized of the P450-derived metabolites, the epoxy- and the 19/20-hydroxy AA, have been implicated in processes such as the control of systemic and renal vascular reactivity, ion fluxes, hormonal signaling and the pathophysiology of experimental hypertension. Thus, our studies and that of others indicate significant roles for this enzyme system in the control of kidney and body homeostasis. However, due to paucity of appropriate molecular tools and techniques, the physiological significance of renal P450 and the site and mode of action of its metabolites remains to be unequivocally defined. Project #2, in conjunction with the cell and organ physiology components of this Program Project, proposes to utilize molecular approaches to the development and use of cellular and/or animal models for the study of P450-gene specific functional phenotypes. We will apply a combination of recombinant DNA and biochemical techniques to the design and construction of high turnover, self contained, regio- and enantioselective AA monooxygenases, the expression of these enzymes in cultured cells, and the identification and mechanistic characterization of insert/cDNA-dependent cellular phenotypes. Targeted gene disruption will be employed for the whole animal integrated functional and biochemical analysis of the significance and the role(s) of specific kidney AA epoxygenase and omega/omega-1 hydrolases. The long term goals of this project are to provide a molecular understanding of renal P450 eicosanoid biological significance and mode of action. The answer to these important questions are needed for the development of meaningful approaches to: a) the unequivocal definition of their physiological and/or pathophysiological significance, and b) subsequent pharmacological and/or clinical interventions.