Vascular Biochemistry and Molecular Biology

血管生物化学和分子生物学

• 批准号: 7140025
• 负责人: Elmer M Price
• 金额: $ 24.42万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-12-01 至 2010-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7140025
• 关键词: antibody; cardiovascular disorder; cardiovascular disorder prevention; dietary lipid; exercise; gene expression; genetic library; genetically modified animals; hyperlipidemia; immunoprecipitation; in situ hybridization; laser capture microdissection; miniature swine; molecular biology; nutrition related tag; polymerase chain reaction; protein protein interaction; protein quantitation /detection; small interfering RNA; transfection /expression vector; vascular endothelium; vascular smooth muscle; western blottings

The Vascular Biochemistry and Molecular Biology Core is designed to provide investigators with access to routine as well as state-of-the-art molecular and biochemical tools, expertise, and techniques. The central theme of this Program Project is that exercise induces beneficial phenotypic changes in vascular cells. Our long-term objectives are to advance the understanding of the molecular, biochemical, cellular, and integrative mechanisms that underlie these training effects. The application of molecular and biochemical techniques will be integral in the exploration of the mechanisms underlying the physiological observations made by this PPG group. The Core's activity reflects a centralization and expansion of services that the Core Leader has successfully provided to PPG investigators for the past decade. The Core laboratory is fully equipped and dedicated to address the stated goals and is directed and staffed by well-qualified personnel. The following Aims are based on the needs of the PPG investigators and reflect a compilation of procedures and techniques that will ensure the efficient completion of each PPG project. The long-term goal of this Core is to track the expression and modulation of specific proteins throughout the coronary and peripheral vasculature and to demonstrate an effect of exercise and/or high-fat-cholesterol on these features. Aim 1 is to perform reverse transcriptase-PCR (RT-PCR) to detect specific mRNAs expressed throughout the vasculature. Aim 2 is to perform real-time PCR in order to more accurately quantitate mRNA expression in select samples from the vasculature of control and experimental animals. Aim 3 is to utilize immunoblots and co-immunoprecipitations to detect and relatively quantitate specific proteins and interactions expressed throughout the circulation. Aim 4a is to use in situ PCR to demonstrate tissue-specific expression of select mRNA species. Aim 4b is to use laser-capture microdissection to selectively isolate and recover select cells from sections of vascular tissue. Real-time PCR will then be used to assess mRNA levels in the recovered cells. Aim 5 is to generate two commercial libraries (one from conduit arteries and one from resistance arterioles) and screen for cDNAs encoding select proteins from the coronary vasculature. Aim 6 is to prepare adeno-associated viral constructs for the transduction of various proteins and siRNA into vascular cells. Aim 7 is to coordinate the commercial preparation of select porcine-specific antibodies. Aim 8 is to prepare and analyze peripheral blood cells from transgenic and non-transgenic pigs. Aim 9 is to provide additional standard techniques as well as pioneer new methods as a resource for PPG investigators.

血管生物化学和分子生物学核心旨在为研究人员提供 常规以及最先进的分子和生化工具、专业知识和技术。中心主题 这项计划项目的目的是运动诱导血管细胞中有益的表型变化。我们的长期合作 目的是促进对分子、生化、细胞和综合机制的理解。 这是这些训练效果的基础。分子和生化技术的应用将在 对这个PPG小组所做的生理观察背后的机制的探索。这个 Core的活动反映了Core Leader成功提供的服务的集中化和扩展 PPG调查人员在过去的十年里。核心实验室设备齐全，致力于解决 规定的目标，并由合格的人员指导和配备人员。以下目标是基于需求的 并反映了程序和技术的汇编，以确保有效地 完成每个PPG项目。这个核心的长期目标是跟踪基因的表达和调节 冠状动脉和外周血管系统中的特定蛋白质以及运动的效果 和/或这些特征上的高脂肪胆固醇。目的1用逆转录酶-聚合酶链式反应(RT-PCR)检测 特定的mRNAs在整个血管系统中表达。目标2是进行实时聚合酶链式反应，以获得更多 准确定量对照和实验血管系统中精选样本的mRNA表达 动物。目标3是利用免疫印迹和免疫共沉淀来检测和相对定量特定的 蛋白质和相互作用在整个循环中表达。目的4a是用原位聚合酶链式反应来证明 组织特异性表达选定的信使核糖核酸物种。目标4b是使用激光捕获显微解剖来选择性地 从血管组织切片中分离和回收精选细胞。然后将使用实时荧光聚合酶链式反应来评估 回收细胞中的mRNA水平。目标5是生成两个商业文库(一个来自导管动脉和 一个来自阻力小动脉)，并筛选编码来自冠状动脉血管系统的特定蛋白的cDNA。 目标6是制备与腺相关的病毒载体，用于将各种蛋白质和siRNA转导到 血管细胞。目标7是协调精选猪特异性抗体的商业制备。目标8是 制备和分析转基因和非转基因猪的外周血细胞。目标9是为了提供 其他标准技术以及开创性的新方法，作为PPG调查人员的资源。