ETHANOL EFFECTS ON PROTEOLYTIC SYSTEMS IN THE LIVER

乙醇对肝脏蛋白水解系统的影响

• 批准号: 2894043
• 负责人: Terrence M. Donohue
• 金额: $ 10.89万
• 依托单位: UNIVERSITY OF NEBRASKA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-08-01 至 2000-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2894043
• 关键词: alcoholic beverage consumption; carbohydrate receptor; detoxification; endopeptidases; ethanol; gene expression; immunocytochemistry; intracellular transport; laboratory rat; liver cells; liver metabolism; lysosomes; mannose 6 phosphate; membrane biogenesis; nuclear factor kappa beta; proteasome; protein metabolism; protein transport; proteolysis; ubiquitin

DESCRIPTION: (Adapted from the Investigator's Abstract) The hypotheses of this proposal are: 1) chronic ethanol consumption impairs lysosome biogenesis by preventing processing and trafficking of lysosomal hydrolases, causing their placement at other intracellular or extracellular sites; 2) ethanol administration alters the ubiquitin-proteasome pathway by inactivating the proteasome which could lead to accumulation of modified proteins. Ethanol may differentially influence the proteasome in Kupffer cells. In Specific Aim 1, lysosome biogenesis will be measured by examining the processing, and compartmentalization of the protease, cathepsin L in hepatocytes isolated from control and ethanol-fed rats. This enzyme follows a specific pathway through vesicular compartments en route to the lysosome and the investigator's Aim 1 is to determine the step(s) at which this process is impaired by ethanol, because misrouting of cathepsin L and other hydrolases could potentially cause cell damage due to their potent hydrolytic capacities. In Specific Aim 1b they will examine whether ethanol influences the distribution of the mannose-6-phosphate receptor by subcellular fractionation immunocytochemistry and functional assay studies. This receptor mediates lysosome assembly by targeting cathepsin L and other hydrolases to the lysosome. The investigators postulate that ethanol may change its intracellular distribution as well as its ligand affinity for procathepsin L. In Specific Aim 2, the components of the ubiquitin-proteasome pathway will be examined in whole livers as well as parenchymal and Kupffer cells of control and ethanol-fed rats subjected to both ad lib and intra gastric feeding regimens. This nonlysosomal proteolytic pathway has a crucial role in: 1) the degradation of altered proteins; and 2) the activation of the transcription factor NFkappaB which initiates transcription of genes involved in the inflammatory and immune responses. The investigators postulate that while ethanol may down-regulate the proteasome in liver parenchymal cells, its activity may be regulated differentially in Kupffer cells, since they play a paracrine role in the pathogenesis of alcoholic liver disease.

描述：（改编自研究者的摘要）假设 该建议是：1）长期乙醇消耗会损害溶酶体 通过阻止溶酶体水解酶的加工和运输来进行生物发生， 导致它们放置在其他细胞内或细胞外位点； 2） 乙醇给药通过以下方式改变泛素-蛋白酶体途径 使蛋白酶体失活，这可能导致修饰物的积累 蛋白质。 乙醇可能对库普弗蛋白酶体产生不同的影响 细胞。 在具体目标 1 中，溶酶体生物发生将通过检查来测量 蛋白酶、组织蛋白酶 L 的加工和区室化 从对照大鼠和乙醇喂养大鼠中分离出肝细胞。 这种酶如下 通过囊泡区室到达溶酶体的特定途径 研究者的目标 1 是确定该步骤 该过程受到乙醇的损害，因为组织蛋白酶 L 和其他酶的路径错误 水解酶可能由于其强大的作用而导致细胞损伤 水解能力。 在具体目标 1b 中，他们将检查乙醇是否 通过影响甘露糖-6-磷酸受体的分布 亚细胞分级分离免疫细胞化学和功能测定研究。 该受体通过靶向组织蛋白酶 L 和其他物质介导溶酶体组装 水解酶进入溶酶体。 研究人员推测乙醇可能 改变其细胞内分布及其配体亲和力 组织蛋白酶 L 前体。在具体目标 2 中，组织蛋白酶 L 的成分 泛素-蛋白酶体途径将在整个肝脏以及 对照组和乙醇喂养大鼠的实质细胞和库普弗细胞 随意喂养和胃内喂养方案。 这种非溶酶体 蛋白水解途径在以下方面起着至关重要的作用：1）改变的降解 蛋白质； 2) 转录因子 NFkappaB 的激活 启动参与炎症和免疫的基因转录 回应。 研究人员假设，虽然乙醇可能会下调 肝实质细胞中的蛋白酶体，其活性可能受到调节 在库普弗细胞中存在差异，因为它们在 酒精性肝病的发病机制。