ACETAMINOPHEN BINDING/HEPATOCYTE HOMEOSTASIS-TOXICITY

对乙酰氨基酚(APAP)是一种应用广泛、安全的退热药 过量服用会导致严重疼痛的止痛药 肝脏坏死。肝毒性是由微粒体引起的 APAP对首先耗尽的电泳体的激活 谷胱甘肽保留，然后与关键细胞共价结合 组件。APAP或其代谢物的机制 启动和延续病理序列，从而导致 细胞死亡仍不清楚。最近对APAP的研究表明 展示了选择性的早期共价结合到单个 微体蛋白条带。APAP还导致了超微结构 内质网、线粒体的生化变化， 溶酶体和质膜。两国之间的关系 蛋白质的选择性芳基化及其对细胞的影响 APAP尚未建立。拟议的研究将 关注共价结合的性质和重要性 APAP肝毒性中的特异性蛋白。这将是 通过研究1)修饰之间的关系来解决 通过选择性共价结合和/或 氧化，2)特定细胞器功能的改变，3) 肝毒性的发生和发展。设计 将涉及协调对小鼠肝细胞的研究，在 在小鼠体内进行培养研究。我们已经开发出一种 与共价结合特异的亲和纯化抗体 APAP。它将在拟议的研究中广泛使用，以 允许鉴定APA结合的细胞器蛋白质 和APAP特异性之间关系的测定 结合和动态平衡过程中的变化以及 这些改变与肝毒性的关系。肝 损伤将通过生化、组织病理学和 在超微结构上。毒性的程度将通过 也要仔细选择APAP的剂量和时间组合 如通过实验增强和对抗。这将是 允许评估共价结合的重要性和 改变蛋白质的巯基水平以使特定的 细胞器在APAP肝毒性中的作用。澄清 APAP引起的改变的性质和识别 对毒物至关重要的同位平衡功能改变 过程将允许更清楚地了解生物化学 APAP及其类似物肝毒性机制的研究 代理肝毒素。这将促进最终的 为预防该病开发合理的替代疗法。

Acetaminophen (APAP) is a widely used, safe, antipyretic analgesic which, when taken in excessive doses, causes severe hepatic necrosis. Hepatotoxicity is the result of microsomal activation of APAP to an electrophile which first depletes glutathione reserves and then covalently binds to critical cell components. The mechanism whereby APAP or its metabolite initiate and perpetuate the pathological sequence which results in cell death remains unknown. Recent studies with APAP have demonstrated selective early covalent binding to a single microsomal protein band. APAP also resulted in ultrastructural and biochemical changes in endoplasmic reticulum, mitochondria, lysosomes and plasma membrane. The relationship between the selective arylation of proteins and the observed cellular effects of APAP has not yet been established. The proposed studies will focus upon the nature and importance of covalent binding to specific proteins in APAP-induced hepatotoxicity. This will be addressed by studying the relationships among 1) the modification of specific proteins by selective covalent binding and/or oxidation, 2) the alterations in specific organelle functions, and 3) the initiation and progression of the hepatotoxicity. The design will involve coordination of studies of mouse hepatocytes, in culture with in vivo studies in mice. We have developed an affinity purified antibody which is specific for covalently bound APAP. It will be used extensively in the proposed studies to permit identification of organelle proteins to which APA binds and determination of the relationship between APAP's specific binding and the alterations in homeostatic processes as well as the relationship of the alterations to the hepatotoxicity. Liver damage will be verified biochemically, histopathologically and ultrastructurally. The degree of toxicity will be varied through careful selection of APAP dosage and time combinations as well as through experimental enhancement and antagonism. This will permit assessment of the importance of covalent binding and altered protein sulfhydryl levels to the derangement of specific organelle functions in APAP hepatotoxicity. Clarification of the nature of the APAP induced changes and identification of the altered homcostatic functions which are critical to the toxic process will permit a clearer understanding of the biochemical mechanisms underlying the hepatotoxicity of APAP and similarly acting hepatotoxins. This will facilitate the eventual development of rational alternative therapies for its prevention.