DEHP-INDUCIBLE CYTOCHROME P450 FORMS IN KIDNEY AND LIVER

DEHP 诱导的细胞色素 P450 在肾脏和肝脏中的形式

• 批准号: 2414948
• 负责人: Richard T. Okita
• 金额: $ 16.63万
• 依托单位: WASHINGTON STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-08-01 至 1999-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2414948
• 关键词: FK506; RNase protection assay; calcium channel blockers; cytochrome P450; diethylhexylphthalate; drug adverse effect; enzyme induction /repression; fatty acid metabolism; gene expression; high performance liquid chromatography; hydroxylation; immunosuppressive; kidney metabolism; kidney pharmacology; laboratory rabbit; laboratory rat; liver metabolism; liver pharmacology; membrane lipids; northern blottings; nuclear runoff assay; peroxidation; phospholipids; scintillation counter; western blottings

Members of the cytochrome P450 4A (CYP4A) subfamily catalyze the omega - or penultimate -hydroxylation of saturated and unsaturated fatty acids and prostaglandins. P450 4A forms are induced by peroxisome proliferators and their induction represents one of the earliest cellular events following exposure to these agents. It is thought that P450 4A induction and formation of oxidized fatty acids may be essential for induction of peroxisomal enzymes. There is considerable interest in the cellular effects of peroxisome proliferators, because these chemicals have been shown to cause hepatic tumors and testicular atrophy in rodents. In addition, 4A forms synthesize products which mediate vasoconstriction of renal vessels and an increase in P450-mediated fatty acid omega- hydroxylase activity is reported prior to the elevation in blood pressure in genetically-bred hypertensive rats. P450 4A forms have been identified in many species and one human 4A form has been sequenced. In rats, three members of the 4A family, 4A1, 4A2, and 4A3 have been identified and form specific oligonucleotide probes and an antibody which recognizes the three forms on western blots have been developed. Certain P450 4A forms exhibit organ specific expression and are regulated by sex hormones. The objectives of this competitive grant renewal are: 1) To localize specific 4A forms in distinct segments of the rat nephron by western blot analysis, because there is considerable uncertainty over the identity of renal forms. The localization of renal 4A forms will be compared in male and female rats because expression of renal 4A forms is sex hormone-dependent; 2) To identify 4A forms which are induced by two immunosuppressive drugs, cyclosporine and FK 506. Cyclosporine is an important therapeutic agent used in transplantation surgery and in treatment of autoimmune diseases, but nephrotoxicity is a major side effect. Studies indicate there is a correlation between induction of fatty acid omega-hydroxylase activity by cyclosporine and "the onset of nephrotoxicity; 3) To examine the mechanisms by which calcium channel blockers suppress induction of hepatic enzymes in DEHP (diethylhexyl phthalate), MEHP (monoethylhexyl phthalate) or clofibrate treated rats or isolated hepatocytes and to examine whether 20-hydroxyeicosatetraenoic acid (20-HETE) and other omega- or penultimate- oxidized fatty acids regulate intracellular calcium concentrations in isolated rat hepatocytes; and 4) To examine the interferon-induced suppression of hepatic 4A forms in phthalate or clofibrate treated rats or isolated hepatocytes. Interferon has been shown to inhibit the induction of P450 4A forms and its suppressive effects on peroxisome fatty acyl CoA oxidase will be examined. The long-term goals of this proposal are to determine the function of P450-mediated fatty acid omega-hydroxylases, its role in peroxisome proliferation, and its physiological function in the kidney.

细胞色素P450 4A（CYP 4A）亚家族的成员催化ω- 或倒数第二-饱和和不饱和脂肪酸的羟基化， 兰丁 P450 4A形式由过氧化物酶体增殖物诱导， 它们的诱导代表了最早的细胞事件之一， 暴露在这些特工之下据认为，P450 4A诱导和 氧化脂肪酸的形成可能是诱导 过氧化物酶体 人们对细胞生物学有相当大的兴趣， 过氧化物酶体增殖剂的影响，因为这些化学物质已经 在啮齿类动物中可导致肝肿瘤和睾丸萎缩。在 此外，4A型合成的产物介导血管收缩， 肾血管和P450介导的脂肪酸ω- 在血压升高之前报告羟化酶活性 高血压大鼠的实验结果。P450 4A表格已确定 在许多物种和一个人类4A形式已被测序。在大鼠中，3 4A家族的成员4A 1、4A 2和4A 3已经被鉴定并形成 特异性寡核苷酸探针和识别这三种寡核苷酸的抗体。 已经开发了Western印迹的形式。某些P450 4A形式显示 器官特异性表达并受性激素调节。的 这一竞争性赠款更新的目标是：1）本地化的具体 通过蛋白质印迹分析，4A在大鼠肾单位的不同节段中形成， 因为对于肾脏的身份存在相当大的不确定性， forms.肾4A型的定位将在男性和女性中进行比较。 雌性大鼠，因为肾脏4A型的表达具有性别依赖性; 2)为了鉴定由两种免疫抑制药物诱导的4A型， 环孢素和FK 506。环孢霉素是一种重要的治疗药物 用于移植手术和治疗自身免疫性疾病， 但肾毒性是主要的副作用研究表明， 脂肪酸ω-羟化酶活性的诱导与 环孢素和“肾毒性的发生; 3）检查 钙通道阻滞剂抑制肝细胞凋亡的机制 DEHP（邻苯二甲酸二乙基己酯）、MEHP（邻苯二甲酸单乙基己酯）中的酶 或氯贝特处理的大鼠或分离的肝细胞，并检查是否 20-羟基二十碳四烯酸（20-HETE）和其它ω-或倒数第二- 氧化脂肪酸调节细胞内钙浓度 分离的大鼠肝细胞;和4）检测干扰素诱导的肝细胞凋亡。 抑制邻苯二甲酸酯或氯贝特处理大鼠的肝脏4A型，或 分离的肝细胞。干扰素已被证明可以抑制诱导 P450 4A型的表达及其对过氧化物酶体脂酰辅酶A的抑制作用 将检查氧化酶。该提案的长期目标是 确定P450介导的脂肪酸ω-羟化酶的功能， 在过氧化物酶体增殖中的作用，以及其在 肾