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: 3113468
• 负责人: Terrence M. Donohue
• 金额: $ 17.8万
• 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 1996-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3113468
• 关键词: alcoholic beverage consumption; alcoholism /alcohol abuse; autophagy; ethanol; fluorimetry; immunocytochemistry; laboratory rat; liver cells; liver metabolism; liver toxic disorder; lysosomes; mitochondria; pathogenic diet; protein metabolism; proteolysis; ubiquitin; vesicle /vacuole; western blottings

The central hypothesis of the proposed investigation is that ethanol consumption inhibits hepatic proteolysis by affecting the lysosomal and extralysosomal proteolytic pathways in the liver. An ethanol-elicited inhibition of protein degradation is one of the principal factors responsible for ethanol-induced protein accumulation in the liver, a phenomenon which contributes to hepatomegaly resulting from chronic drinking. Our aim in these investigations is to examine two key hepatic proteolytic systems in rat liver and to determine to what degree they are affected by acute and,/or chronic ethanol administration. The systems to be analyzed are the hepatic autophagic/lysosomal system and the ubiquitin proteolytic pathway. These pathways are responsible for the complete degradation of most intracellular proteins. Disruption of their intracellular function(s) may lead to other disturbances in liver cell function. The specific objectives of this proposal are to examine the effects of acute and/or chronic ethanol consumption on 1) The formation and maturation of hepatic autophagic vacuoles, 2) endogenous proteolytic activities as well as HSP-70-mediated proteolytic activities of liver lysosomes 3) the content of hepatic ubiquitin as well as ubiquitin dependent conjugation and proteolytic activities in rat liver extracts, and 4) the activities of these proteolytic systems in different zones of the hepatic lobule. The latter objective will be important in determining whether any relationship exists between the ethanol-induced reduction in hepatic proteolysis and ethanol-elicited specific cell injury. Our investigation will utilize recently updated techniques for the enrichment and isolation of autophagic vacuoles from the livers of control and ethanol-fed rats. We will analyze these vacuoles morphologically and biochemically in order to determine whether ethanol ingestion affects the formation, maturation and proteolytic capacities of these organelles. We will then assess the effects of ethanol consumption, as well as that of ethanol's initial oxidation product acetaldehyde, on the formation and the degradation of ubiquitin-protein conjugates. Finally, we will analyze the protein content, the rates of protein catabolism, as well as the aforementioned proteolytic systems in isolated perivenous and periportal hepatocytes from control and ethanol-fed rats. Since the liver is responsible for the regulation of serum protein, amino acid, lipid and glucose levels, it is important to understand the effects of ethanol toxicity or hepatic protein metabolism.

拟议研究的中心假设是乙醇 消耗通过影响溶酶体和 肝脏中的溶酶体外蛋白水解途径。 乙醇引发的 抑制蛋白质降解是主要因素之一 负责乙醇诱导的肝脏蛋白质积累， 导致慢性肝肿大的现象 喝。 我们这些研究的目的是检查两个关键的肝脏 大鼠肝脏中的蛋白水解系统并确定其程度 受急性和/或慢性乙醇施用的影响。 该系统 待分析的是肝自噬/溶酶体系统和泛素 蛋白水解途径。 这些路径负责完整的 大多数细胞内蛋白质的降解。 扰乱他们的 细胞内功能可能导致肝细胞的其他紊乱 功能。 该提案的具体目标是检验以下措施的影响： 急性和/或慢性乙醇消耗对 1) 形成和成熟的影响 肝自噬泡，2) 内源性蛋白水解活性 以及 HSP-70 介导的肝溶酶体的蛋白水解活性 3) 肝泛素的含量以及泛素依赖性缀合和 大鼠肝脏提取物中的蛋白水解活性，以及​​ 4) 的活性 这些蛋白水解系统位于肝小叶的不同区域。 这 后一个目标对于确定是否存在任何关系非常重要 乙醇诱导的肝蛋白水解酶减少与 乙醇引起特异性细胞损伤。 我们的调查将利用最近更新的技术 从对照肝脏中富集和分离自噬泡 和乙醇喂养的大鼠。 我们将从形态上分析这些液泡 生化以确定乙醇摄入是否会影响 这些细胞器的形成、成熟和蛋白水解能力。 我们 然后将评估乙醇消耗的影响以及 乙醇的初始氧化产物乙醛的形成和 泛素-蛋白质缀合物的降解。 最后我们来分析一下 蛋白质含量、蛋白质分解代谢率以及 上述分离的静脉周围和门脉周围的蛋白水解系统 来自对照大鼠和乙醇喂养大鼠的肝细胞。 由于肝脏负责调节血清蛋白、氨基酸 酸、脂质和葡萄糖水平，了解其影响很重要 乙醇毒性或肝蛋白质代谢。