ARACHIDONATE PRODUCTS IN DIOXIN AND PCB TOXICITY

花生四烯酸产品的二恶英和多氯联苯毒性

• 批准号: 3251062
• 负责人: ARLEEN B. RIFKIND
• 金额: $ 20.23万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-12-01 至 1994-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3251062
• 关键词: arachidonate; cardiotoxin; chick embryo; cytochrome P450; dioxins; eicosanoid metabolism; environmental contamination; environmental toxicology; fatty acid metabolism; halobiphenyl /halotriphenyl compound; heart cell; high performance liquid chromatography; leukotrienes; lipoxygenase; liver cells; prostaglandin endoperoxide synthase; radioimmunoassay; receptor; thin layer chromatography; toxin metabolism; unspecific monooxygenase

Dioxin (TCDD) and PCBs are major environmental toxins, but neither the precise mechanism of their toxicity or biochemical mechanisms of defense are known. Furthermore there are large differences in species sensitivity to TCDD and PCBs which have yet to be explained. In this research we will extend our previous original observations that TCDD causes major increases in NADPH-dependent arachidonic acid (AA) metabolism. The factors regulating TCDD's effects and the role of AA metabolites in enhancing or limiting TCDD toxicity will be investigated, principally using a chick embryo model. TCDD's effects on AA metabolism in intact cells will be studied using cultured hepatocytes, Kupffer cells and endothelial cells to identify the types of liver cells in which AA metabolism is increased and in cultured cardiac myocytes. Its effects on AA metabolites formation in ovo will also be examined. TCDD's effects on AA release from membranes and on activation of phospholipases A2 and C and on diacylglycerol will be investigated using cultured hepatocytes and cardiac myocytes. Dose-response curves for TCDD's effects on AA metabolism and release and on deethylation of 7-ethoxycoumarin and 7-ethoxyresorufin by cultured cells will be compared to asses relationships between TCDD's effects on eicosanoids and mixed function oxidases. Immunoinhibition of TCDD and PB induced AA metabolism in microsomes and cell homogenates by antibodies against purified P-450 from TCDD-and PB-microsomes and cell homogenates by antibodies against purified P-450s from TCDD- and PB-microsomes and to NADPH- cytochrome P-450 reductase will be used to prove that P-450 mediates TCDD's increase of NADPH-dependent eicosanoid metabolism, to identify the isozymes involved and determine if the same or different isozymes mediate TCDD's and PB's effects. TCDD's specificity will be probed by comparing its effects to those of representative toxic and nontoxic PCBs and to PB. A role for eicosanoids in TCDD's effects will be studied by investigating if HPLC-purified eicosanoids generated by TCDD-treated livers mimic or modify TCDD's effects on protein kinase C in cultured hepatocytes and myocytes or cardiac contractile responses to beta- adrenergic agonists and if inhibitors of AA metabolism diminish the effects. Structure of bio-active eicosanoids will be characterized by GC/MS. In vivo modifications of TCDD toxicity by pharmacologic agents which alter AA metabolism will be examined to learn if inhibiting P-450 mediated eicosanoid metabolism decreases TCDD toxicity. AA metabolism and MFO activity will be assyed concomitantly so that valid conclusions can be drawn.

二恶英（TCDD）和多氯联苯（PCB）是主要的环境毒素， 其毒性或生化机制的确切机制 防御是已知的。 此外， 物种对TCDD和PCBs的敏感性， 解释道 在这项研究中，我们将扩展我们以前的原始 观察到TCDD导致NADPH依赖性 花生四烯酸（AA）代谢。 调节TCDD的因素 AA代谢物在增强或限制 TCDD毒性将主要使用小鸡进行研究 胚胎模型 TCDD对完整细胞AA代谢的影响将 使用培养的肝细胞、枯否细胞和 内皮细胞，以确定其中AA的肝细胞类型 在培养的心肌细胞中，代谢增加。 其 还将检查对卵内AA代谢物形成的影响。 TCDD对AA从细胞膜释放和活化的影响 将研究磷脂酶A2和C以及对二酰基甘油的作用 使用培养的肝细胞和心肌细胞。 剂量反应 TCDD对AA代谢和释放的影响曲线， 7-乙氧基香豆素和7-乙氧基试卤灵的脱乙基化 将比较细胞以评估TCDD之间的关系 对类花生酸和混合功能氧化酶的影响。 TCDD和PB诱导AA代谢的免疫抑制作用 和细胞匀浆，通过抗纯化的P-450的抗体， TCDD-和PB-微粒体和细胞匀浆，通过抗 从TCDD-和PB-微粒体和NADPH-纯化的P-450 细胞色素P-450还原酶将用于证明P-450 介导TCDD增加NADPH依赖性类花生酸代谢， 以鉴定所涉及的同工酶，并确定是否相同或 不同的同工酶介导TCDD和PB的作用。 TCDD's 特异性将通过比较其效果与 代表性的有毒和无毒多氯联苯和PB。 的作用 类花生酸在TCDD的影响将通过调查， TCDD处理的肝脏模拟物产生的HPLC纯化的类二十烷酸 或改变TCDD对培养的细胞中蛋白激酶C的作用。 肝细胞和肌细胞或心脏收缩反应， 肾上腺素能激动剂，如果AA代谢抑制剂减少 方面的影响. 生物活性类花生酸的结构将被表征 通过GC/MS测定TCDD的体内毒性。 将检查改变AA代谢的药物，以了解是否 抑制P-450介导的类花生酸代谢降低TCDD 毒性 AA代谢和MFO活性将被测定 以便得出有效的结论。