ETHNIC VARIABILITY IN AN ETHANOL INDUCIBLE P450 ENZYME

乙醇诱导型 P450 酶的种族变异性

• 批准号: 2045014
• 负责人: JUDY L RAUCY
• 金额: $ 9.78万
• 依托单位: AGOURON INSTITUTE
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-08-01 至 1998-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2045014
• 关键词: Mexican Americans; Native Americans; alcoholism /alcohol abuse; caucasian American; cytochrome P450; disease /disorder proneness /risk; enzyme activity; enzyme mechanism; genetic markers; genetic polymorphism; human subject; lymphocyte; messenger RNA; northern blottings; racial /ethnic difference; restriction fragment length polymorphism; western blottings

The incidence of alcoholism is much higher in Native Americans and Mexican Americans that in Caucasians. The reasons may be sociological, psychological, biochemical, or a combination of these factors. In addition to a higher incidence of alcoholism, there is also an increased susceptibility of alcohol related liver disease in these two populations which may, in part, be related to metabolism of alcohol. P4502E1 is known to metabolize several compounds including ethanol to hepatotoxic metabolites. Moreover, chronic exposure to alcohol causes an increase in the expression of this particular P450 enzyme. This increase in expression may play a role in the development of occult liver disease from alcohol and other hepatotoxins metabolized by this enzyme. A focus of this proposal is to determine whether P4502E1 plays a role in the susceptibility of certain individuals to alcohol-related liver injury. this will be investigated by screening lymphocytes from 3 separate ethnic populations, Native Americans, Mexican Americans and Caucasians both alcoholic and non-alcoholic, for differences in the expression of the enzyme. To determine enzyme expression, lymphocytes will be isolated from whole blood collected from alcoholic and nonalcoholic subjects from all three subpopulations. P4502E1 will be monitored by immunoblot analysis of microsomes isolated from the lymphocytes using a mono- specific antibody prepared to human liver P4502E1. Detection of an elevation in this enzyme may serve as a predictor that chemical or alcohol-induced hepatotoxicity may result. This proposal focuses on different ethnic populations to determine whether this enzyme can be used as a biochemical marker of alcohol-dependent liver disease in subpopulations which are considered to be at high risk. Identification of specific RFLPs in the regulatory region of CYP2E1 is also proposed which will be correlated with phenotypic expression, ethnicity and alcoholism. Further characterization of P4502E1 in lymphocytes will involve examining whether ethanol causes an increase in P4502E1 mRNA with a subsequent increase in the enzyme. This will be accomplished by Northern blot analysis of mRNA isolated from alcoholic and non-alcoholic subjects and results may provide a mechanism in which ethanol increases P4502E1 in lymphocytes. Preliminary evidence from our laboratory suggests that the lymphocyte enzyme exhibits a molecular weight (MW) difference from that of the liver P450. Thus, the structure of the lymphocyte protein will also be examined and compared to liver P4502E1 in order to assess whether the altered structure modifies the activity of the enzyme. Whether the expression and regulation of P4502E1 in lymphocytes parallels that which is in liver will be determined in a clinical study using the drug chlorozoxazone as a marker of liver P4502E1 function. If the enzyme is inducible by alcohol in lymphocytes and its regulation and expression mirrors that of the liver, human blood may be a readily accessible tissue to phenotype and study regulation of liver P4502E1 in humans.

酗酒的发生率在美洲原住民中要高得多， 墨西哥裔美国人在高加索人。 原因可能是社会学的， 心理的，生物化学的，或这些因素的组合。 在 除了酗酒的发病率更高外， 这两个人群中酒精相关性肝病的易感性 这可能部分与酒精代谢有关。 P4502E1是 已知代谢几种化合物，包括乙醇至肝毒性 代谢物。 此外，长期接触酒精会导致 这种特殊的P450酶的表达。 的这种增加 表达可能在隐匿性肝病的发展中起作用 酒精和其他由这种酶代谢的肝毒素。 重点 这项建议的目的是确定P4502E1是否在 某些个体对酒精相关肝损伤的易感性。 这将通过筛选来自3个不同种族的淋巴细胞进行研究。 人口，美洲原住民，墨西哥裔美国人和高加索人都 酒精和非酒精，在表达的差异， 酵素 为了确定酶表达，将分离淋巴细胞 从酒精和非酒精受试者采集的全血中， 所有三个亚群。 将通过免疫印迹法监测P4502E1 使用单克隆抗体分析从淋巴细胞分离的微粒体， 制备人肝P4502E1特异性抗体。 检测到 这种酶的升高可以作为一种预测， 可能导致酒精诱导的肝毒性。 该提案的重点是 不同种族的人群，以确定这种酶是否可以使用 作为酒精依赖性肝病的生化标志物， 被认为是高风险的亚群。 识别 并提出了CYP2E1调控区的特异性RFLPs 这将与表型表达、种族和 酒精中毒 淋巴细胞中P4502E1的进一步表征将 涉及检测乙醇是否引起P4502E1 mRNA的增加， 随后的酶的增加。 这将通过 从酒精性和非酒精性细胞中分离的mRNA的北方印迹分析 主题和结果可以提供一种机制，其中乙醇增加 淋巴细胞中P4502E1的表达。 我们实验室的初步证据 表明淋巴细胞酶表现出分子量（MW） 与肝脏P450的差异。 因此， 还将检查淋巴细胞蛋白并与肝脏P4502E1进行比较 为了评估改变的结构是否改变了活性， 的酶。 P4502E1的表达和调控是否与细胞凋亡有关？ 淋巴细胞平行，这是在肝脏将确定在一个 使用药物氯唑沙宗作为肝脏P4502E1标记物的临床研究 功能 如果淋巴细胞中的这种酶可以被酒精诱导， 调节和表达反映了肝脏，人类血液可能是 一个容易获得的组织表型和研究肝脏的调节 P4502E1在人体内