ENDOTHELIN AND RECEPTOR GENE EXPRESSION IN HYPOXIA

缺氧时内皮素和受体基因表达

• 批准号: 3369158
• 负责人: Yiu-Fai Chen
• 金额: $ 20.69万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-05-15 至 1997-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3369158
• 关键词: aorta; cellular pathology; chronic disease /disorder; clearance rate; computer program /software; densitometry; endocrine pharmacology; endothelin; gene expression; hormone inhibitor; hormone receptor; hypoxia; laboratory rat; metalloendopeptidases; nucleic acid hybridization; polymerase chain reaction; protease inhibitor; pulmonary artery; pulmonary hypertension; radioimmunoassay; respiratory pharmacology; solution hybridization; tissue /cell culture; transcription factor; vascular endothelium; vascular smooth muscle; vasoconstriction

Hypoxia stimulates endothelium dependent vasoconstriction in the pulmonary artery and enhances expression of the gene for endothelin-1 (ET-1) in human umbilical vein endothelial cells in culture and in rat lung, but not in organs of the rat that are supplied by the systemic arterial bed. The general hypothesis of the current proposal is that ET-1 gene expression and release of big ET-1 from endothelial cells of the pulmonary vasculature are enhanced by exposure to hypoxia, and that the ET-1 so generated plays an etiologic role in hypoxia induced pulmonary vasoconstriction and pulmonary hypertension. A subhypothesis is that inhibitors of endothelin converting enzyme (ECE) such as phosphoramidon and/or ET-1 receptor antagonists such as [Dpr-1 Asp-15I ET-1 inhibit hypoxic pulmonary vasoconstriction and protect against the development of hypoxic pulmonary hypertension. The Specific Aims are: 1) To test the hypothesis that ET-1 gene expression, ET-1 receptor mechanisms and ET-1 clearance are altered in the rat in response to acute (24, 48 hrs) and chronic (7, 14, 28 das) normobaric hypoxia and relate these alterations to hypoxia induced increases in pulmonary artery pressure. 2) To examine the cellular basis of selectivity of the hypoxic response of ET-1 for lung. 3) To define the cis-regulatory element(s) involved in amplifying transcription of the ET-1 gene in endothelial cells in response to hypoxia and to identify and clone the hypoxic transcription activator protein(s). 4) To test the hypothesis that enhanced ET-1 gene expression and, thereby, increased endogenous ET-1 levels mediate hypoxic pulmonary hypertension in the rat. The effect of lowering endogenous ET-1 levels by inhibiting ECE with the neutral metalloendopeptidase inhibitor phosphoramidon and of blocking ET-1 receptors with the ET-1 antagonist [Dpr-1, Asp15] ET-1 will be examined. The effects of phosphoramidon and [Dpr-1, Aspl5] ET-1 treatment on acute hypoxic pulmonary vasoconstriction and on chronic hypoxia-induced pulmonary hypertension will be tested. Sprague Dawley rats exposed to normobaric hypoxia (10% O2 at 1 atmosphere) will be studied in Specific Aims 1 and 4. Endothelial and smooth muscle cells in low passage derived from porcine main pulmonary artery and aorta and from precapillary arterioles of porcine lung will be studied in Specific Aim 2. Porcine aortic endothelial cells transfected with fragments of human ET-1 genomic DNA containing putative hypoxia response element(s) will be used in Specific Aim 3.

缺氧刺激内皮依赖性血管收缩， 增强内皮素-1基因的表达 人脐静脉内皮细胞和大鼠脐静脉内皮细胞内皮素-1（ET-1）的变化 肺，但不是在大鼠的器官，是由全身 动脉床 目前提案的一般假设是， 血管内皮细胞ET-1基因表达及大ET-1释放的研究 肺血管通过暴露于缺氧而增强，并且 如此产生的ET-1在缺氧诱导的脑缺血中起病因作用， 肺血管收缩和肺动脉高压。 一个子假设 内皮素转化酶（ECE）抑制剂， 磷酰胺和/或ET-1受体拮抗剂，例如[Dpr-1 Asp-15 I ET-1抑制缺氧性肺血管收缩， 缺氧性肺动脉高压的发展。 本研究的具体目的是：1）验证ET-1基因与人外周血淋巴细胞增殖有关的假说， 表达、ET-1受体机制和ET-1清除率改变， 大鼠对急性（24，48小时）和慢性（7，14，28天） 常压缺氧，并将这些变化与缺氧诱导的 肺动脉压升高。2)为了检查细胞基础 ET-1对肺缺氧反应的选择性。3)来定义 参与扩增转录的顺式调节元件 ET-1基因在内皮细胞缺氧反应中的表达及鉴定 克隆低氧转录激活蛋白。4)测试 假设增强ET-1基因表达，从而增加 内源性ET-1水平介导大鼠缺氧性肺动脉高压。 用内皮素抑制内皮素释放酶降低内源性内皮素-1水平的作用 中性金属内肽酶抑制剂phosphoramidon及其阻断 ET-1受体与ET-1拮抗剂[Dpr-1，Asp 15] ET-1的作用， 考察 磷酰胺和[Dpr-1，Asp 15] ET-1对大鼠心肌细胞凋亡的影响 治疗急性缺氧性肺血管收缩和慢性缺氧性肺血管收缩 将测试缺氧诱导的肺动脉高压。 将Sprague道利大鼠暴露于常压缺氧（10%O2，1 将在具体目标1和4中研究大气中的化学物质。内皮和 猪主肺动脉平滑肌细胞低代培养 动脉和主动脉以及来自猪肺的毛细血管前小动脉， 在具体目标2中研究。转染猪主动脉内皮细胞 用含有假定缺氧的人ET-1基因组DNA片段 具体目标3中将使用响应元素。