MECHANISMS OF RESPIRATORY ENDOTHELIAL INJURY BY XO

XO 损伤呼吸内皮细胞的机制

• 批准号: 2222274
• 负责人: JOHN E REPINE
• 金额: $ 32.63万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-02-10 至 1996-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2222274
• 关键词: MECHANISMS; RESPIRATORY; ENDOTHELIAL; INJURY; XO

Xanthine oxidase (XO) appears to participate in a variety of vascular injury syndromes including pulmonary, cardiac, renal, cerebral, and intestinal ischemia-reperfusion or hypoxia/reoxygenation. This possibility is consistent with (a) the high concentrations of XO in capillary endothelial cells, (b) the capacity of XO to generate toxic O2 metabolites and chemotaxins for neutrophils in vitro, and (c) the ability of XO inhibitors to decrease injury in animals. The problem is that the role of XO in human vascular disease is still unproven because XO has been implicated only by using non-specific or incomplete inhibitors of XO activity in animals rather than more accurate modern approaches in relevant human systems. Indeed, the mechanisms which govern regulation of expression of XO and its precursor, xanthine dehydrogenase (XD), the effects of this regulation on pathologic conditions, and the molecular basis for formation of XO and from XD following ischemia and other vascular insults are unclear. Our preliminary data indicate involvement of XO in many diseases. To address definitively the contribution of XO to inflammation and vascular damage, we chose to clone and sequence human XD/XO. Since our prior submission, we have 1) cloned and partially sequenced two candidate DNA probes specific for XO, 2) developed a monoclonal antibody specific for mammalian XO, 3) determined that our human liver cDNA library contains the human XO gene, 4) developed a yeast expression system for human XD, and 5) analyzed new sequence data which supports our hypothesis for XD to XO conversion. Our goal is to test definitively the premise that XO contributes to human disease and to explore the structural basis for XD to XO conversion. Our major objectives are: (a) to construct and test an XD/XO null mutant to produce definitive evidence for the hypothesis that XO dependent mechanisms contribute to cell injury and neutrophil adherence following exposure to hypoxia and reoxygenation; (b) to determine the structural difference between XD and XO activities and the basis for conversion of XD to XO in vitro; and (c) to determine if parameters of ischemia/reperfusion regulate XD/XO gene expression. These critical objectives require cloning and sequencing a human XD/XO cDNA clone. The significance of this project is that it will precisely define the role in ischemic vascular disease of XO in light of its native regulatory mechanisms. This approach will generate new understanding of fundamental mechanisms of tissue injury and inflammation.

黄嘌呤氧化酶（XO）似乎参与了多种血管 损伤综合征，包括肺，心脏，肾脏，大脑和 肠缺血再灌注或缺氧/再氧。 这 可能性与（a）高浓度的XO一致 毛细管内皮细胞，（b）XO产生有毒O2的能力 用于体外嗜中性粒细胞的代谢物和趋化素，（c） XO抑制剂减少动物损伤的能力。 问题是XO在人血管疾病中的作用仍然是 未经证实，因为仅通过使用非特异性或 动物中XO活性的不完全抑制剂，而不是更多 相关人类系统中的准确现代方法。 确实， 控制XO表达及其表达的机制及 前体，黄嘌呤脱氢酶（XD），该调节的作用 在病理条件下以及XO形成的分子基础上 从XD出发，缺血和其他血管损伤还不清楚。 我们的初步数据表明XO参与了许多疾病。 到 确切地解决XO对炎症和血管的贡献 损坏，我们选择克隆并序列人XD/XO。 自我们之前 提交，我们有1）克隆并部分测序了两个候选DNA XO的探针，2）开发了一种特异性的单克隆抗体 哺乳动物XO，3）确定我们的人肝cDNA库包含 人XO基因，4）开发了人类XD的酵母表达系统， 5）分析了支持我们XD假设的新序列数据 进行XO转换。 我们的目标是确切地测试XO的前提 人类疾病并探索XD到XO的结构基础 转换。 我们的主要目标是：（a）构建和测试 XD/XO NULL突变体，为假设提供明确的证据 XO依赖机制有助于细胞损伤和中性粒细胞 暴露于缺氧和重氧后的依从性； （b）到 确定XD和XO活动之间的结构差异以及 在体外转化为XO的基础； （c）确定是否 缺血/再灌注的参数调节XD/XO基因表达。 这些关键目标需要克隆并测序人XD/XO cDNA克隆。 该项目的重要性是它将精确定义 根据其本地的XO缺血性血管疾病的作用 监管机制。 这种方法将产生对 组织损伤和炎症的基本机制。