INOS GENE TRANSFECTION IN PULMONARY HYPERTENSION

肺动脉高压(PH)是发病率和 死亡率影响的患者范围很广。新生儿肺 高血压是新生儿入院的第二大原因 呼吸支持的重症监护室。在成年人中，PH导致 慢性阻塞性疾病患者的显著发病率和死亡率 肺部疾病。在所有患者中，PH的特征是细胞性 肺血管床的增殖和血管反应性改变。 这项建议的目的是评估血管扩张剂的疗效。 病毒介导的诱导型一氧化氮产生的NO的毒性 将诱导型一氧化氮合酶(INOS)基因导入小鼠肺内，并测定其对小鼠肺组织的影响 病毒介导的诱导型一氧化氮合酶在PH发病机制中的作用这个 一般的假设是，病毒感染的iNOS会导致 足够的一氧化氮生成来调节肺血管收缩和 减轻与肺血管相关的肺血管改变 高血压，但NO生成不足，不足以导致毒性。 利用人诱导型一氧化氮合酶基因和大肠杆菌lac Z报告基因作为对照 编码β-半乳糖苷酶(β-GAL)的腺病毒构建我们的 本提案提出的目标是：1)优化iNOS基因传递 并在大鼠肺组织中表达，2)确定转染组的作用 INOS对肺动脉高压发展的影响，以及3)比较 诱导型一氧化氮合酶基因的血管内和气管内传递 基因表达、血管反应性和毒性。这些目标是 在以下具体目标中处理：具体目标1：评估 腺病毒介导的诱导型一氧化氮合酶基因转染人卵巢癌细胞的实验研究 减轻急性肺血管收缩反应。特定目标 第二项：评估诱导型一氧化氮合酶基因转导对血管内皮细胞的影响 慢性低氧性肺动脉高压的发生。特定的 目标3：评估血管内联合化疗的疗效和毒性 气管内给药腺病毒iNOS构建。 这些方法将涉及使用腺病毒载体来携带该基因。 对于将被血管内给药的iNOS或β-Gal； 然后将对肺进行研究以确定血管反应性，否 诱导型一氧化氮合酶基因的产生和定位。一些老鼠会成为 转基因后暴露于慢性低氧环境。最后，血管内和 气管内分娩将在基因定位方面进行比较 和毒性。

Pulmonary hypertension (PH) is a significant cause of morbidity and mortality affecting a broad range of patients. Neonatal pulmonary hypertension is the second leading cause for admission to neonatal intensive care units for respiratory support. In adults, PH causes significant morbidity and mortality in patients with chronic obstructive pulmonary disease. In all patients, PH is characterized by cellular proliferation and altered vasoreactivity in the pulmonary vascular bed. The objectives of this proposal are to evaluate the vasodilator efficacy and toxicity of NO produced by virally mediated inducible nitric oxide synthase (iNOS) gene transfection in the lung and to determine the effect of virally transfected iNOS on the pathogenesis of PH. The general hypothesis is that virally transfected iNOS will result in sufficient NO formation to modulate pulmonary vasoconstriction and attenuate pulmonary vascular changes associated with pulmonary hypertension, but insufficient NO formation to result in toxicity. Utilizing human iNOS gene and, as a control, the E. coli lac Z reporter gene coding for beta-galactosidase (beta-gal) adenovirus constructs our goals set forth in this proposal are: 1) to optimize iNOS gene delivery and expression in the rat lung, 2) to determine the role of transfected iNOS on the development of pulmonary hypertension, and 3) to compare intravascular and intratracheal delivery of the iNOS gene in terms of gene expression, vascular reactivity and toxicity. These goals are addressed in the following specific aims: Specific Aim number 1: Assess the effectiveness of adenovirus-mediated iNOS gene transfection in attenuating acute pulmonary vasoconstrictor responses. Specific Aim number 2: Assess iNOS gene transfection-mediated effects on the development of chronic hypoxia-induced pulmonary hypertension. Specific Aim number 3: Assess the efficacy and toxicity of intravascularly and intratracheally administered adenoviral iNOS constructs. The methods will involve using adenovirus constructs containing the gene for iNOS or beta-gal that will be administered intravascularly; the lungs will then be studied to determine vascular reactivity, NO production, and localization of transfected iNOS. Some rats will be transfected and exposed to chronic hypoxia. Finally, intravascular and intratracheal delivery will be compared in terms of gene localization and toxicity.