METHYLENE DIANILINE--A SELECTIVE BILE DUCT TOXICANT?

DAPM（4，4 '-diaminodiphenylmethane）被提议作为选择性胆管 有毒，因为DAPM导致胆管上皮细胞形态学损伤 对肝细胞无影响的肝内胆管BEC 并迅速损害胆功能，特别是葡萄糖重吸收 但不改变肝细胞转运 功能协调发展的关于DAPM和其他药物的作用机制知之甚少。 胆管破坏剂损害胆管。DAPM后3小时，BEC 显示线粒体超微结构改变， 微绒毛和高尔基池扩张。肝脏研究 在DAPM处理后3小时，线粒体显示在 酶活性、线粒体通透性特征，或 组织化学染色模式。观察到BEC线粒体 是DAPM损伤的早期部位，可以解释DAPM损伤的快速性和严重性。 DAPM引起的病变，并提供了一个独特的机会， 研究胆管毒物的作用机制。 我们的总体目标是了解DAPM如何以及为什么会导致早期， 选择性损伤胆管细胞。该应用程序的重点将 对DAPM代谢产物在胆汁中的排泄及其对BEC的破坏能力有影响。 我们的实验方法将使用25 mg/kg剂量的DAPM， BEC中中度损伤性功能/结构改变，但 不改变胆汁流量或损伤肝细胞。代谢物研究 将在麻醉大鼠中进行胆汁排泄和胆汁功能 尿道内注入牛磺胆酸盐以维持胆汁流动。要求1 将阐明DAPM对紧密连接渗透性的影响， 细胞旁标志物和细胞化学超微结构方法。目的2 将表征体内DAPM代谢物，特别是胆汁中的代谢物， 将使用分离的细胞在体外评估DAPM的细胞毒性， 肝细胞与BEC。AIM 3将确定是否抑制剂 缀合反应降低了BEC和BEC中的II相酶， 肝细胞，通过确定DAPM的可能近似毒物 在DAPM的胆汁排泄已被抑制的大鼠中的损伤程度 在体内调节，并评估可疑的近端 通过体外细胞毒性试验测定毒物。我们的新数据提供了这样的 强有力的证据表明，DAPM对BEC的选择性作用，特别是 在BEC线粒体上，一种新的AIM 4提出了生理和 BEC处理的分离的线粒体功能的生化研究 关于DAPM 拟议的研究将大大有助于我们了解 BEC损伤的机制，并提供了一个新的模型系统， 关于BEC在胆汁形成中的作用的问题。

DAPM (4,4'-diaminodiphenylmethane) is proposed as a selective bile duct toxicant because DAPM causes morphological injury to biliary epithelial cells (BEC) of the intrahepatic bile ducts without effects on hepatocytes and rapidly impairs biliary functions, particularly glucose reabsorption from the biliary traCt, but does not alter hepatocellular transport functions. Little is known about the mechanisms by which DAPM and other cholangiodestructive agents damage bile ducts. By 3 hr after DAPM, BEC show ultrastructural alterations in mitochondria, loss of lumenal microvilli and dilation of Golgi Cisternae. Studies of liver mitochondria 3 hr after DAPM treatment demonstrate no alterations in enzyme activities, mitochondrial permeability characteristics, or histochemical staining patterns. The observation that BEC mitochondria are an early site of DAPM injury may explain the rapidity and severity of lesions caused by DAPM, and provides a unique opportunity to investigate mechanisms of bile duct toxicants. Our overall objective is to understand how and why DAPM causes early, selective injury to bile duct cells. The focus of this application will be on DAPM metabolites excreted in bile and their capacity to damage BEC. Our experimental approach will use the 25 mg/kg dose of DAPM which causes moderately injurious functional/structural alterations in BEC but does not alter bile flow or damage hepatocytes. Studies of metabolite excretion in bile and biliary function will be done in anesthetized rats infused with taurocholate intraduodenally to maintain bile flow. AIM 1 will clarify the effects of DAPM on tight junction permeability using paracellular markers and cytochemical ultrastructural methods. AIM 2 will characterize DAPM metabolites in vivo, particularly in bile, and will use isolated cells to assess DAPM cytotoxicity in vitro in hepatocytes versus BEC. AIM 3 will determine if inhibitors of conjugation reactions decrease Phase Il enzymes in both BEC and hepatocytes, identify possible proximate toxicants of DAPM by determining extent of injury in rats whose biliary excretion of DAPM has been modulated in vivo, and assess relative toxicities of suspect proximate toxicants by in vitro cytotoxicity assays. Our new data provide such strong evidence of the selective effect of DAPM on BEC, and particularly on BEC mitochondria, that a new AIM 4 proposes physiological and biochemical studies of mitochondrial function in isolated BEC treated with DAPM. The proposed research will contribute significantly to our understanding of the mechanisms of BEC injury and provide a new model system to ask questions about the role of BEC in bile formation.