MODULATION OF NO WITH PULMONARY GENE TRANSFER

通过肺部基因转移调节 NO

• 批准号: 6397761
• 负责人: Loretta G Que
• 金额: $ 12.49万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-04-01 至 2005-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6397761
• 关键词: apoptosis; arginase; biotechnology; cell proliferation; chemokine; enzyme activity; free radical oxygen; gene therapy; histology; hyperoxia; immunocytochemistry; inflammation; laboratory rabbit; lung injury; nitric oxide; nitric oxide synthase; oxidative stress; superoxide dismutase; transfection; transfection /expression vector; western blottings

Acute lung injury in the intensive care unit is treated with administration of high inspired oxygen tensions which can be toxic to the lung. Pulmonary oxygen toxicity is generally attributed to the excessive production of reactive oxygen intermediates (RO1), e.g., superoxide (O2), hydrogen peroxide (H2O2), and hydroxyl radicals. Recently nitric oxide (NO) is being used to treat hyperoxic lung disease, but its effect is variable and its role remains controversial. We hypothesize that selective modulation of NO/RO1 in different cellular compartments regulates tissue responses to hyperoxia. In this proposal, we will employ state of the art adenoviral gene transfer technique in vitro and in vivo. The transgenes we have selected for these studies are neuronal nitric oxide synthase (nNOS), extracellular superoxide dismutase (ECSOD), and arginases I and II (AI and AII). Overexpression of these four genes are expected to affect NO and reactive oxygen intermediates (RO1) production in different cell compartments: nNOS (yields intracellular NO/yields 02-), ECSOD (yields extracellular NO/yields 02-), and AI and AII (yields intracellular NO/yields O2-). We will determine how these transgenes modulate, oxidative/nitrosative stress, and NO/ROI mediated inflammation, cellular proliferation, and apoptosis. The specific aims of this proposal are 1) characterize the transgene expression of nNOS, ECSOD, AI, and AII in vitro 2) characterize intratracheal adenoviral gene transfer in normal rabbit lung and 3) evaluate how adenoviral mediated transgenes nNOS, ECSOD, AI, and AII modulate effect of NO in hyperoxia. These studies should not only better unravedl the basic mechanisms of hyperoxic lung injury, but also provide a rationale basis for therapeutic gene transfer using NO. Ultimately we are hopeful that these results may also extend to other oxidant injuries in the lung.

重症监护室中的急性肺损伤通过施用可能对肺有毒的高吸入氧张力来治疗。 肺氧中毒通常归因于活性氧中间体（RO 1）的过度产生，例如，超氧化物（O2）、过氧化氢（H2 O2）和羟基自由基。 近年来，一氧化氮（NO）被用于治疗高氧肺疾病，但其作用是可变的，其作用仍然存在争议。 我们推测，选择性调制NO/RO 1在不同的细胞室调节组织反应高氧。在这个提议中，我们将采用最先进的腺病毒基因转移技术在体外和体内。 我们为这些研究选择的转基因是神经元型一氧化氮合酶（nNOS），细胞外超氧化物歧化酶（ECSOD），和脱氢酶I和II（AI和AII）。 预期这四种基因的过表达影响不同细胞区室中的NO和活性氧中间体（RO 1）的产生：nNOS（产生细胞内NO/产生O2-）、ECSOD（产生细胞外NO/产生O2-）以及AI和AII（产生细胞内NO/产生O2-）。 我们将确定这些转基因如何调节氧化/亚硝化应激和NO/ROI介导的炎症、细胞增殖和凋亡。 本研究的具体目的是：1）体外表征nNOS、ECSOD、AI和AII的转基因表达; 2）表征正常兔肺中腺病毒基因转移; 3）评估腺病毒介导的转基因nNOS、ECSOD、AI和AII如何调节NO在高氧中的作用。这些研究不仅可以更好地揭示高氧肺损伤的基本机制，而且可以为利用NO进行治疗性基因转移提供理论基础。最终，我们希望这些结果也可以扩展到其他氧化剂肺损伤。