Mechanism of Metabolism of S-Nitrosothiols

S-亚硝基硫醇的代谢机制

• 批准号: 7259906
• 负责人: NEIL HOGG
• 金额: $ 26.19万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-04-01 至 2011-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7259906
• 关键词: Acids; Address; Affect; Amino Acid Transport System L; Apoptosis; Area; Asthma; Automobile Driving; Back; Behavior; Binding; Biological; Blood Vessels; Bronchoalveolar Lavage; Bronchodilator Agents; Cell physiology; Cells; Coculture Techniques; Cultured Cells; Cyclic GMP; Cysteine; Cystic Fibrosis; Data; Detection; Diffusion; Enzymes; Erythrocytes; Event; Functional disorder; Funding; Gel; Glutathione; Guanylate Cyclase; Heme; Hemeproteins; Hemoglobin; Homeostasis; Inflammation; Inflammatory; Light; Liquid substance; Location; Lung; Lung diseases; Mediating; Metabolism; Methodology; Methods; Modeling; Modification; Molecular Weight; NADH; NADP; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Signaling Pathway; Nitrite Reductase; Nitrites; Nitrosation; Numbers; Outcome; Oxidative Stress; Oxygen; Pathway interactions; Post-Translational Protein Processing; Prosthesis; Proteins; Proteome; Proteomics; Publishing; Pulmonary Hypertension; Range; Reaction; Research Personnel; Role; Scheme; Signal Transduction; Soluble Guanylate Cyclase; Sulfhydryl Compounds; System; Techniques; Testing; Thinking; Tissues; Transducers; Work; caspase-3; concept; enzyme activity; functional group; improved; inhaled nitric oxide; programs; pulmonary function; response; uptake

DESCRIPTION (provided by applicant): This proposal seeks funds to examine the fundamental cellular behavior of nitric oxide (NO) and S- nitrosothiols (RSNO). Nitric oxide (NO) is an essential component of vascular function in pulmonary and all other tissues. In the lung, NO has particular importance as inhaled NO is used as a therapy for pulmonary hypertension, and alterations in NO homeostasis have been appreciated in many pulmonary diseases including asthma and cystic fibrosis. In addition S-nitrosoglutathione (GSNO) has been detected in micro molar concentration in lung lavage and is decreased in asthma. In general RSNO have been invoked as 'carriers' of NO bioactivity and RSNO have been used in cell culture studies as 'NO-donors'. Our studies in the previous funding period have highlighted the fact that NO and RSNO affect cellular functions by clearly different mechanisms. In particular we characterized the mechanisms by which RSNO can be transported into cells and have shown that these compounds can affect cellular functions via both NO-dependent and NO-independent pathways. This has led to the overall hypothesis of this proposal, that lmwtRSNO formation represents a bifurcation in NO signaling. We will address this hypothesis by the following three Specific Aims. 1) We will further characterize how lmwtRSNO are synthesized and transported into cells and how the glutathione/NADH/NADPH can control the activity and stability of RSNO. 2) Examine the differential sensitivity of selected pathways to modulation by RSNO and NO. 3) Define modifications of intracellular proteins using a range of lmwtRSNO and NO concentrations to uncover the differential sensitivity of proteins and pathways to S-nitrosation and S-thiolation. We will accomplish these aims using lung-cell specific cell culture models, specialized chemiluminescence detection methodologies and proteomic techniques. Successful completion of these studies will disentangle the cellular effects of NO from the direct effects of RSNO to more fully understand how these species affect cellular function. Importantly, these studies will point to a role of RSNO formation not simply as 'carriers' of NO bioactivity but as a distinct bifurcation in NO signaling pathways.

描述（由申请人提供）：本提案寻求资金来研究一氧化氮（NO）和S-亚硝基硫醇（RSNO）的基本细胞行为。一氧化氮（NO）是肺和所有其他组织血管功能的重要组成部分。在肺中，NO具有特别的重要性，因为吸入NO可用于治疗肺动脉高压，并且在许多肺部疾病（包括哮喘和囊性纤维化）中已发现NO稳态的改变。此外，肺灌洗液中检测到s -亚硝基谷胱甘肽（GSNO）的微摩尔浓度，哮喘患者的GSNO浓度降低。一般来说，RSNO被称为NO生物活性的“载体”，RSNO被用作细胞培养研究中的“NO供体”。我们在之前资助期的研究强调了NO和RSNO通过明显不同的机制影响细胞功能的事实。我们特别描述了RSNO转运进入细胞的机制，并表明这些化合物可以通过no依赖性和no非依赖性途径影响细胞功能。这导致了本提案的总体假设，即lmwtRSNO的形成代表了NO信号的分支。我们将通过以下三个具体目标来解决这一假设：1)我们将进一步表征lmwtRSNO如何合成和转运到细胞中，以及谷胱甘肽/NADH/NADPH如何控制RSNO的活性和稳定性。2)检查所选途径对RSNO和NO调制的差异敏感性。3)使用一系列lmwtRSNO和NO浓度定义细胞内蛋白质的修饰，以揭示蛋白质对s -亚硝化和s -硫基化的差异敏感性和途径。我们将使用肺细胞特异性细胞培养模型，专门的化学发光检测方法和蛋白质组学技术来实现这些目标。这些研究的成功完成将解开NO对细胞的影响与RSNO的直接影响，从而更全面地了解这些物种如何影响细胞功能。重要的是，这些研究将指出RSNO的形成不仅是NO生物活性的“载体”，而且是NO信号通路中的一个明显的分支。