Functional roles of Nrf2 and NQO1 genetic variants in hyperoxic lung injury-ARDS

描述（由申请人提供）：补充氧气经常用于治疗急性呼吸窘迫综合征（ARDS）患者的肺功能不全，ARDS是一种严重的急性肺损伤（ALI），影响全球数百万人。高氧肺损伤被认为是ALI/ARDS的理想模型。本申请中提出的研究的中心假设是，NF-E2相关因子（Nrf 2）和/或NADPH醌还原酶（NQO 1）基因中的特定单核苷酸多态性（SNP）通过减弱肺和肝功能II期抗氧化酶的表达，导致氧介导的活性氧（ROS）的形成增加，这又导致这些患者对ALI/ARDS的易感性增加以及肺损伤加重。为了实现这些目标，我们提出以下具体目标：1。为了验证以下假设：携带Nrf 2或NQO 1基因上的特定SNP的人比不携带的人更容易发生ALI/ARDS，并且这些个体比携带野生型基因的人对氧化应激的敏感性增加。将分析来自患有ALI/ARDS的个体或对照的气管内抽吸物中SNP、F2-异前列烷/异呋喃、大体积氧化DNA加合物和蛋白质氧化产物的水平的存在。还将使用来自这些个体的气管吸出物的RNA研究基因表达谱。2.为了确定缺乏Nrf 2和NQO 1基因的小鼠对高氧肺损伤更敏感的机制，并检验细胞色素P4501 A（CYP 1A）诱导剂[例如，<$-萘酮（BNF）]或奥美拉唑（OM）将通过需要肝和肺CYP 1A酶的新机制来拯救缺乏功能性Nrf 2或NQO 1基因的小鼠对抗高氧肺损伤。3.确定Nrf 2或NQO 1启动子上的已知SNP在人肺细胞或人源化小鼠体内调节氧毒性的机制。该目标有两个子目标。（i）.验证携带已知Nrf 2或NQO 1基因SNPs的人肺细胞对氧毒性更敏感的假设。(ii)建立表达正常人Nrf 2基因或NQO 1基因的转基因人源化小鼠或这些基因上携带已知SNP的小鼠，并确定SNP在高氧肺损伤中的作用。包括基因组学（微阵列）和蛋白质组学方法在内的组学方法将用于确定Nrf 2或NQO 1变体导致肺损伤的分子机制。目标的成功实现可以导致用于开发新生物标志物以及新方法的创新策略（例如，PPI如OM）用于预防/治疗人的ALI/ARDS的用途。 公共卫生相关性：该项目旨在确定这些NF-E2相关因子（Nrf 2）和/或NADPH醌还原酶（NQO 1）基因的遗传变异导致人类急性肺损伤（ALI）和急性呼吸窘迫综合征（ARDS）的机制。这些目标的成功实现可能会导致开发新的生物标志物的创新策略以及预防/治疗人类ALI/ARDS的新方法。

DESCRIPTION (provided by applicant): Supplemental oxygen is frequently used in the treatment of pulmonary insufficiency in patients with acute respiratory distress syndrome (ARDS), which is a severe form of acute lung injury (ALI), affecting millions worldwide. Hyperoxic lung injury is recognized as an appropriate model for ALI/ARDS. The central hypothesis of the research proposed in this application is that specific single nucleotide polymorphisms (SNPs) in the genes for NF-E2-related factor (Nrf2) and/or NADPH quinone reductase (NQO1) contribute to ALI/ARDS by attenuating the expression of pulmonary and hepatic functional phase II anti-oxidant enzymes, leading to increased formation of oxygen-mediated reactive oxygen species (ROS), which in turn results in increased susceptibility to ALI/ARDS, as well as exacerbated lung damage in these patients. In order to achieve these goals, we propose the following Specific Aims: 1. To test the hypothesis that humans carrying specific SNPs on the Nrf2 or NQO1 genes will be more susceptible to develop ALI/ARDS than those who do not, and these individuals will display increased sensitivity to oxidative stress than those carrying the wild type genes. Endotracheal aspirates from individuals suffering from ALI/ARDS or controls will be analyzed for the presence of SNPs, F2-isoprostanes/isofurans, levels of bulky oxidative DNA adducts, and protein oxidation products. Gene expression profiles will also be studied using RNA from tracheal aspirates of these individuals. 2. To determine the mechanisms by which mice lacking the genes for Nrf2 and NQO1 are more susceptible to hyperoxic lung injury, and test the hypothesis that the cytochrome P4501A (CYP1A) inducer [e.g., ¿-napthoflavone (BNF)] or omeprazole (OM) will rescue the mice lacking functional Nrf2 or NQO1 genes against hyperoxic lung injury, via novel mechanisms entailing hepatic and pulmonary CYP1A enzymes. 3. To determine the mechanisms by which known SNPs on the Nrf2 or NQO1 promoter modulate oxygen toxicity in human lung cells or in humanized mice in vivo. This aim has two sub-aims. (i). To test the hypothesis that human lung cells carrying known SNPs of Nrf2 or NQO1 gene will be more susceptible to oxygen toxicity. (ii) To create transgenic humanized mice expressing the normal human Nrf2 gene or NQO1 gene or those carrying known SNPs on these genes, and determine the role of SNPs in hyperoxic lung injury. Omics' approaches, including genomics (microarrays) and proteomics approaches will be used to determine the molecular mechanisms by which Nrf2 or NQO1 variants contribute to lung injury. Successful accomplishment of the aims could lead to innovative strategies for the development of novel biomarkers as well as new approaches (e.g., use of PPI such as OM) for the prevention/treatment of ALI/ARDS in humans. PUBLIC HEALTH RELEVANCE: This project is aimed at determining the mechanisms by which these genetic variants of NF-E2-related factor (Nrf2) and/or NADPH quinone reductase (NQO1) genes contribute to acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) in humans. Successful accomplishment of the aims could lead to innovative strategies for the development of novel biomarkers as well as novel approaches for the prevention/treatment of ALI/ARDS in humans.