ARACHIDONATE PRODUCTS IN DIOXIN AND PCB TOXICITY

二恶英(TCDD)和多氯联苯是主要的环境毒素，但 其毒性的确切机制或生化机制 都是已知的防御手段。此外，在这一点上， 物种对TCDD和多氯联苯的敏感性 解释过了。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依赖的显著增加 花生四烯酸(AA)代谢。TCDD的调控因素 AA代谢物在增强或限制中的作用和作用 将对TCDD的毒性进行研究，主要是使用雏鸡 胚胎模型。TCDD对完整细胞内AA代谢的影响 用培养的肝细胞、库普弗细胞和 内皮细胞鉴定再生障碍性贫血的肝细胞类型 在培养的心肌细胞中代谢增加。它的 对卵子中AA代谢物形成的影响也将被检测。 TCDD对膜上AA释放和激活的影响 将研究磷脂酶A2和C以及二酰甘油 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及其他 7-乙氧基香豆素和7-乙氧基间苯二酚的培养脱乙基 将比较细胞以评估TCDD之间的关系 对二十烷类化合物和混合功能氧化酶的影响。 TCDD和PB诱导的微粒体内AA代谢的免疫抑制 以及细胞匀浆中抗纯化的P-450的抗体 抗TCDD-和PB-微粒体和细胞匀浆 从TCDD-和PB-微粒体中提纯P-450并转化为NADPH- 细胞色素P-450还原酶将被用来证明P-450 介导TCDD增加NADPH依赖的二十烷类代谢， 以确定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的 我们将通过比较其效果来探讨其特异性。 具有代表性的有毒和无毒的多氯联苯和聚丁二烯。一个角色是 将通过调查以下情况来研究TCDD中的二十烷类化合物的影响 经TCDD处理的肝脏产生的高效液相纯化二十烷类化合物模拟 或改变TCDD对体外培养的蛋白激酶C的影响 肝细胞和心肌细胞或心脏对β-氨基丁酸的收缩反应 肾上腺素能激动剂和AA代谢的IF抑制剂降低 效果。生物活性二十烷类化合物的结构将被表征 GC/MS法研究TCDD的体内药理毒性 将对改变AA代谢的药物进行检查，以了解是否 抑制P-450介导的二十烷类化合物代谢可降低TCDD 毒性。AA代谢和MFO活性将会下降 因此，可以得出有效的结论。

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.