HYPOXIC REGULATION OF VASCULAR ENDOTHELIAL GROWTH FACTOR

血管内皮生长因子的缺氧调节

• 批准号: 2519200
• 负责人: ANDREW Peter LEVY
• 金额: $ 2.77万
• 依托单位: GEORGETOWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-01 至 1997-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2519200
• 关键词: DNA binding protein; DNA footprinting; gel mobility shift assay; gene expression; genetic regulation; genetic regulatory element; genetic transcription; growth factor; ischemia; laboratory rat; messenger RNA; myocardium; nuclear runoff assay; posttranscriptional RNA processing; protein purification; reporter genes; tissue /cell culture; vascular endothelial growth factors; vascular endothelium; vascular endothelium permeability

The primary objective of this project is to delineate the regulation of rat vascular endothelial growth factor gene expression by hypoxia in vitro and ischemia in vivo. Vascular endothelial growth factor (VEGF) is a potent angiogenic factor and potent endothelial cell specific mitogen. Preliminary data have shown that the steady state mRNA for VEGF is markedly upregulated by hypoxia in vitro. The specific aims of the studies described herein are as follows: 1) to define the contribution of new transcription versus mRNA stabilization to the increase in the steady state VEGF mRNA levels following a hypoxic stimulus; 2) to define the cis- acting regulatory sequences responsible for mediating VEGF transcriptional activation and/or mRNA stabilization by hypoxia; 3) to identify specific nucleic acid binding proteins that bind to these cis regulatory sequences; 4) to utilize reporter gene constructs to allow for a rational pharmacological approach to manipulate-endogenous VEGF gene expression. Methods to be employed include functional assays (nuclear run-off transcription assays,transient expression assays, in vitro RNA degradation analyses) and structural analyses (DNA and RNA mobility shift assays, in vitro DNAase l footprint analysis, protein purification and characterization). Acquiring a better understanding of the molecular mechanisms governing the regulation of VEGF, and hence angiogenesis, by hypoxia may permit the development of novel new strategies to treat human conditions resulting from vascular insufficiency such as coronary artery disease and peripheral vascular disease.

本项目的主要目标是描述 体外低氧诱导大鼠血管内皮生长因子基因表达 和体内缺血。血管内皮生长因子（VEGF）是一种 有效的血管生成因子和有效的内皮细胞特异性丝裂原。 初步数据表明，VEGF的稳态mRNA是 在体外缺氧时显著上调。研究的具体目标 本文所描述的是：1）定义新的贡献 转录相对于mRNA稳定的增加， 状态VEGF mRNA水平后缺氧刺激; 2）以确定顺式- 负责介导VEGF转录的作用调节序列 通过缺氧激活和/或mRNA稳定化; 3）鉴定特异性 结合这些顺式调节序列的核酸结合蛋白; 4)利用报告基因构建体， 药理学方法来操纵内源性VEGF基因表达。 所采用的方法包括功能测定（核流出 转录测定、瞬时表达测定、体外RNA降解 分析）和结构分析（DNA和RNA迁移率变动分析， 体外DNA酶I足迹分析、蛋白质纯化和 表征）。 更好地了解控制细胞凋亡的分子机制， 通过缺氧调节VEGF，从而调节血管生成，可能允许 开发新的治疗人类疾病的新策略， 血管功能不全，如冠状动脉疾病和外周 血管疾病