Effect of ethanol on retinoid metabolism and signaling in zebrafish embryos

乙醇对斑马鱼胚胎中类维生素A代谢和信号传导的影响

• 批准号: 7142426
• 负责人: WILLIAM F BOSRON
• 金额: $ 17.99万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-30 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7142426
• 关键词: biological signal transduction; bone development; craniofacial; developmental genetics; developmental neurobiology; embryo /fetus toxicology; ethanol; fetal alcohol syndrome; fluorescent dye /probe; gene expression; genetic regulation; in situ hybridization; labyrinth; morphometry; nonmammalian vertebrate embryology; phenotype; recombinant proteins; retinoate; retinoid binding proteins; teratogens; vitamin metabolism; vitamin receptor; zebrafish

DESCRIPTION (provided by applicant): This is an R21 exploratory/development grant application to develop methods to quantify the content of alcohol metabolizing enzymes and variants (initially alcohol dehydrogenase and eventually aldehyde dehydrogenase and microsomal ethanol oxidizing CYP isoenzymes) in human tissues (liver and esophagus). We believe that the application fits the R21 guidelines because we will use newly developed mass spectrometry proteomics methods to address the specific aims. The application is also in response to PA-05-074, Mechanisms of Alcohol-Induced Tissue Injury, because it provides important protein expression information to support the investigation of alcohol metabolizing enzyme genotypes that confer susceptibility or resistance to ethanol-induced cell/tissue damage. Our central hypothesis is that the expression/content of alcohol metabolizing enzymes and variants determine the pharmacokinetics and cellular toxicology of ethanol and acetaldehyde in tissues. In Aim #1, we will develop proteomics methods to quantify the expression of seven alcohol dehydrogenase (ADH) isoenzymes in tissues by triple quadrupole linear ion trap LC/MS/MS and multiple reactions monitoring analysis with appropriate peptide/protein internal standards. We will assess the feasibility of developing methods to look at the whole set of ADH, ALDH and microsomal ethanol oxidizing CYP isoenzymes involved in alcohol metabolism simultaneously. In Aim #2, we will collect and genotype human liver samples and matched normal esophagus and squamous cell carcinoma tissue samples from a local population in Indianapolis for the pilot proteomics study. The tissue samples will be genotyped at ADH and other alcohol metabolizing enzyme loci. In Aim #3, we will perform a preliminary proteomics study to look at the variability of expression/content of the seven ADH isoenzymes in the liver and esophagus tissue samples using the proteomics methods developed in Aim #1. We will identify whether there is a correlation between ADH polymorphisms and protein expression in tissues. We will estimate the individual contribution of the seven ADHs to alcohol metabolic capacity in tissue samples and the contribution of polymorphism to differences in alcohol metabolic capacity. We will assess the feasibility of performing targeted proteomics studies with tissues from other populations that express different SNPs in alcohol metabolizing enzymes.

描述（由申请人提供）：这是一项R21探索性/开发资助申请，旨在开发用于定量人体组织（肝脏和食管）中乙醇代谢酶和变体（最初为乙醇脱氢酶，最终为乙醛脱氢酶和微粒体乙醇氧化脱氢酶同工酶）含量的方法。我们认为该应用程序符合R21指南，因为我们将使用新开发的质谱蛋白质组学方法来解决特定目标。本申请也是对PA-05-074（酒精诱导的组织损伤机制）的响应，因为它提供了重要的蛋白质表达信息，以支持对酒精诱导的细胞/组织损伤易感性或抗性的酒精代谢酶基因型的研究。我们的中心假设是酒精代谢酶和变体的表达/含量决定了组织中乙醇和乙醛的药代动力学和细胞毒理学。在目标#1中，我们将开发蛋白质组学方法，通过三重四极线性离子阱定量组织中七种乙醇脱氢酶（ADH）同工酶的表达。 LC/MS/MS和多反应监测分析，使用适当的肽/蛋白质内标物。我们将评估开发方法的可行性，看看整套ADH，ALDH和微粒体乙醇氧化脱氢酶同工酶参与酒精代谢的同时。在目标#2中，我们将从印第安纳波利斯的当地人群中收集人类肝脏样本和匹配的正常食管和鳞状细胞癌组织样本并进行基因分型，用于初步蛋白质组学研究。组织样本将在ADH和其他酒精代谢酶位点进行基因分型。在目标#3中，我们将使用目标#1中开发的蛋白质组学方法进行初步蛋白质组学研究，以观察肝脏和食管组织样本中7种ADH同工酶表达/含量的变异性。我们将确定ADH多态性和组织中蛋白质表达之间是否存在相关性。我们将估计七种ADH对组织样本中酒精代谢能力的个体贡献以及多态性对酒精代谢能力差异的贡献。我们将评估对来自其他人群的在酒精代谢酶中表达不同SNP的组织进行靶向蛋白质组学研究的可行性。