Oxidative Stress and Vascular HO in Diabetes

糖尿病中的氧化应激和血管 H2O2

• 批准号: 7276681
• 负责人: Nader G. Abraham
• 金额: $ 31.47万
• 依托单位: NEW YORK MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-15 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7276681
• 关键词: Adenovirus Vector; Animals; Apoptosis; Apoptotic; Attenuated; Bilirubin; Biliverdine; Blood Vessels; Carbon Monoxide; Cardiovascular Diseases; Cardiovascular system; Cell Adhesion Molecules; Cell Death; Cells; Complication; Cytoplasm; Cytoprotection; DNA; Depth; Development; Diabetes Mellitus; Diabetic Angiopathies; Diabetic mouse; Elevated Bilirubin; Endothelial Cells; Experimental Diabetes Mellitus; Free Radicals; Functional disorder; Gene Expression; Gene Targeting; Gene Transfer; Genes; Genetic; Germ Lines; Goals; Heme; Human; Hyperglycemia; In Vitro; Inbred NOD Mice; Inflammation; Injury; Insulin; Insulin-Dependent Diabetes Mellitus; Isoenzymes; Laboratories; Lead; Lentivirus Vector; Mediating; Mitochondria; Morbidity - disease rate; Mus; Oxidative Stress; Oxygenases; Pathway interactions; Patients; Pharmaceutical Preparations; Physiological; Protein Overexpression; Proteins; Publishing; Rattus; Relative (related person); Research Proposals; Resistance; Retroviral Vector; Role; System; Testing; Time; Transgenic Mice; Up-Regulation; Vascular Diseases; Vascular Endothelium; Work; base; case control; caspase-9; cobaltiprotoporphyrin; cytochrome c; diabetic; diabetic rat; gain of function; gene therapy; heme oxygenase-1; heme oxygenase-2; improved; in vivo; innovation; loss of function; mortality; mouse model; novel; prevent; promoter; response; stem; type I and type II diabetes; type I diabetic; vascular endothelial dysfunction

DESCRIPTION (provided by applicant): Macro and microvascular diseases are the principal causes of morbidity and mortality in patients with type 1 and type 2 diabetes. Endothelial dysfunction, as evidenced by the increased release of free radicals, increases adhesion molecules and apoptosis. Previous work by us and others has shown that a decrease in HO-1 gene expression enhances apoptosis and vascular injury which can be prevented by pharmacological mediated upregulation of HO-1. Our preliminary results indicate that upregulation of HO-1 prevents endothelial apoptosis by decreasing cellular heme content, increasing CO and bilirubin levels and decreasing EC-SOD and DNA degradation. The goal of this proposal is to elucidate the mechanism by which HO-1 prevents diabetes-mediated endothelial dysfunction and to explore the use of genetic and/or pharmacological approaches to achieve long-term vascular protection. Our hypothesis is that HO-1 gene transfer will provide powerful vascular protection by increasing heme degradation and subsequently increasing CO and bilirubin synthesis. The increased levels of CO and bilirubin will result in an increase in EC-SOD, and eNOS, a decrease in O2- and improved vascular response. In addition, increase in mitochondrial HO-1 levels will increase mitochondrial transport carrier, decrease mitochondrial ROS and prevent release of cytochrome c and activation of caspase 9. We plan to test this hypothesis using genetic probes (retroviral LXSN lentiviral, and adenoviral) and in genetically spontaneously diabetic mice, (NOD) mice. We will use the loss-of-function HO-2 (-/-) and gain-of-function, HO-1 and HO-2 gene transfer to HO-2 (-/-) to decipher the role of each gene in vascular protection. The following specific aims will test the hypothesis. (1)To determine whether genetic intervention, using retroviral vectors, to selectively increases HO-1 provides vascular protection in diabetic rats and NOD mice, and whether this effect is due to CO, bilirubin or both. We will examine the effect HO-1 derived CO and bilirubin on EC-SOD, eNOS and O2- levels. (2) HO-2 (-/-) and NOD mice will be used to test the hypothesis that HO-1 gene transfer can provide vascular protection and that CO, bilirubin or both are obligatory for the vascular protection via increase in EC-SOD, eNOS, decrease in O2- and iNOS. (3)To determine the mechanism whereby HO-1 gene expression (CO and bilirubin) act on the extrinsic and intrinsic pathway of pro-apoptosis and anti-apoptotic proteins. This will examine the role of CO and bilirubin produced by HO-1 in both the mitochondria and cytoplasm. (4) To evaluate whether lentiviral vectors targeting endothelial cells using cell specific promoter (VE-CAD) to overexpress HO-1 gene is sufficient to offset diabetes-induced vascular injury. This proposal is novel in its approach. It will allow for the first time, an in-depth analysis of the function of HO-1 in vascular protection and for the development of innovative gene-targeting therapies for the treatment of type I diabetes.

描述（由申请人提供）：大血管和微血管疾病是 1 型和 2 型糖尿病患者发病和死亡的主要原因。内皮功能障碍，如自由基释放增加所证明，会增加粘附分子和细胞凋亡。我们和其他人之前的工作表明，HO-1 基因表达的减少会增强细胞凋亡和血管损伤，而这可以通过药理学介导的 HO-1 上调来预防。我们的初步结果表明，HO-1 的上调通过降低细胞血红素含量、增加 CO 和胆红素水平以及减少 EC-SOD 和 DNA 降解来防止内皮细胞凋亡。该提案的目标是阐明 HO-1 预防糖尿病介导的内皮功能障碍的机制，并探索使用遗传和/或药理学方法来实现长期血管保护。我们的假设是 HO-1 基因转移将通过增加血红素降解并随后增加 CO 和胆红素合成来提供强大的血管保护。 CO 和胆红素水平的增加将导致 EC-SOD 和 eNOS 增加，O2- 减少并改善血管反应。此外，线粒体 HO-1 水平的增加将增加线粒体转运载体，减少线粒体 ROS，并阻止细胞色素 c 的释放和 caspase 9 的激活。我们计划使用基因探针（逆转录病毒 LXSN 慢病毒和腺病毒）和遗传性自发性糖尿病小鼠（NOD）小鼠。我们将使用功能丧失的 HO-2 (-/-) 和功能获得、HO-1 和 HO-2 基因转移至 HO-2 (-/-) 来破译每个基因在血管保护。以下具体目标将检验该假设。 (1)确定使用逆转录病毒载体选择性增加HO-1的基因干预是否可以为糖尿病大鼠和NOD小鼠提供血管保护，以及这种作用是否是由CO、胆红素或两者引起的。我们将研究 HO-1 衍生的 CO 和胆红素对 EC-SOD、eNOS 和 O2- 水平的影响。 (2) HO-2 (-/-) 和 NOD 小鼠将用于测试以下假设：HO-1 基因转移可以提供血管保护，并且 CO、胆红素或两者对于通过增加 EC-SOD 的血管保护是必需的、 eNOS、O2- 和 iNOS 减少。 (3)确定HO-1基因表达(CO和胆红素)作用于促凋亡和抗凋亡蛋白的外在和内在途径的机制。这将检查 HO-1 产生的 CO 和胆红素在线粒体和细胞质中的作用。 （4）评估使用细胞特异性启动子（VE-CAD）靶向内皮细胞的慢病毒载体过表达HO-1基因是否足以抵消糖尿病引起的血管损伤。该提案的方法新颖。它将首次深入分析 HO-1 在血管保护中的功能，并开发用于治疗 I 型糖尿病的创新基因靶向疗法。