S-nitrosylation signaling in asthma

哮喘中的 S-亚硝基化信号传导

基本信息

批准号：
10457996
负责人：
JONATHAN S. STAMLER
金额：
$ 40.25万
依托单位：
INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-08-01 至 2026-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10457996
关键词：
ADRBK1 gene Acute Adrenergic Agents Affect Agonist Air Movements American Arrestins Asthma Biochemistry Bronchodilator Agents Cardiac Cell surface Cells Chronic Clinical Collaborations Complement Complex Coupled Cysteine Data Development Diagnosis Enzymes Epithelial Cells Event Excision Exhibits Functional disorder G protein coupled receptor kinase G-Protein-Coupled Receptors GTP-Binding Proteins Genetic Glutathione Glutathione Reductase Height Heterotrimeric GTP-Binding Proteins Homeostasis Human Inhalation Injury Intervention Kinetics Knock-in Link Lung Maintenance Mediating Mediator of activation protein Modeling Molecular Molecular Weight Mus Mutant Strains Mice Mutate Mutation Nitric Oxide Nitric Oxide Synthase Oxidation-Reduction Oxidoreductase Pathogenesis Pathology Pathway interactions Patients Pharmaceutical Preparations Phosphorylation Phosphotransferases Physiology Point Mutation Positioning Attribute Post-Translational Protein Processing Proteins Publishing Pulmonary Function Test/Forced Expiratory Volume 1 Regulation Resistance Role S-Nitrosoglutathione S-Nitrosothiols SKIL gene Sampling Signal Pathway Signal Transduction Single Nucleotide Polymorphism Site Smooth Muscle Myocytes Sulfhydryl Compounds System Tachyphylaxis Testing Work airway hyperresponsiveness airway inflammation airway remodeling arrestin 1 arrestin 2 asthma model asthmatic asthmatic patient base bench to bedside beta-2 Adrenergic Receptors beta-arrestin biological systems clinically relevant cohort cost desensitization efficacy testing improved inhibitor innovation mortality mouse model patient subsets personalized approach pre-clinical preservation prevent programs protective effect protein function pulmonary function receptor synergism

项目摘要

PROJECT SUMMARY/ABSTRACT Project 1 Asthma afflicts over 7% of Americans and is the result of chronic inflammation of the airways leading to airway remodeling and hyperresponsiveness that impedes air flow. Severe asthma is asthma that remains problematic despite maximal intervention with conventional asthma therapies, and it accounts for the majority of the mortality and cost of asthma. Dysfunction of airway β2-adrenergic receptor (β2AR) signaling contributes to severe asthma pathogenesis, but the precise signaling mechanisms responsible are unclear. While activation of β2AR using inhaled “β-agonist” drugs is a mainstay of acute asthma treatment, overactivation of β2AR is detrimental and can be fatal. Airway β2ARs signal both through heterotrimeric G proteins and through G protein-coupled receptor kinase (GRK)/β-arrestin pathways that also mediate receptor phosphorylation, desensitization, and internalization. Through a long-standing collaboration, the Stamler and Gaston groups have found that airways are regulated by nitric oxide (NO) through S-nitrosylation of thiols to form S-nitrosothiol (SNO), including on cysteine residues in proteins, a post-translational modification that alters protein functions. In addition, SNO forms on low molecular weight thiols, including glutathione to form SNO-glutathione (GSNO). We demonstrated that inhaled GSNO elevates lung protein-SNO and is protective in asthma, identified the enzyme SNO- glutathione reductase (GSNOR) that inactivates GSNO, and demonstrated that mice lacking GSNOR are protected from developing asthma. Since the β2AR can activate NO synthase in the airways to generate NO, there is a need to discover how this promotes endogenous SNO-mediated bronchoprotection. Our recent work has shown that β2AR is S-nitrosylated after activation, and preventing SNO-β2AR with a point mutation augments β2AR signaling. Importantly, mice bearing β2AR with a knock-in of this mutation are protected from developing asthma. We have previously shown that β2AR regulators GRK2 and β-arrestin2 are S-nitrosylated to inhibit their activity to desensitize the β2AR, and mice bearing GRK2-C340S and β-arrestin2-C253S knock-in exhibit heighted β2AR activity and worsened injury in cardiac models. We have begun to test the efficacy of inhaled GSNO to affect bronchorelaxation and improved lung function in patients, and these data and clinical samples uniquely position us to examine the role of S-nitrosylation in regulating the β2AR pathway from bench to bedside. The Central Hypothesis of Project 1 is that the β2AR signaling system is a key target and mediator of SNO-GSNO- GSNOR protective effects in the airways in severe asthma. Our studies will define the role of S-nitrosylation of specific β2AR signaling pathway components in a murine model of asthma, delineate the roles of inhaled GSNO and of GSNO dinitrosylases on β2AR signaling components in murine models of asthma and in human lung primary cells, and demonstrate that inhaled GSNO improves both airway flow and β2-agonist responsiveness in severe asthma. This work complements the clinical aims in Projects 2 and 3 by providing a mechanistic link between NO/GSNO/GSNOR actions and the regulation of β2AR signaling in severe asthma.

项目概要/摘要项目1 超过 7% 的美国人患有哮喘，是导致呼吸道慢性炎症的结果阻碍气流的重塑和高反应性是哮喘的一个难题。尽管对传统哮喘治疗进行了最大程度的干预，但它占了大部分死亡率气道 β2 肾上腺素能受体 (β2AR) 信号传导功能障碍会导致严重哮喘。发病机制，但使用 β2AR 激活的确切信号机制尚不清楚。吸入“β受体激动剂”药物是急性哮喘治疗的主要手段，β2AR过度激活会带来困扰，气道 β2AR 通过异源三聚体 G 蛋白和 G 蛋白偶联受体发出信号。激酶 (GRK)/β-抑制蛋白途径也介导受体磷酸化、脱敏和通过长期合作，斯塔姆勒和加斯顿小组发现了气道。通过硫醇的 S-亚硝基化形成 S-亚硝基硫醇 (SNO)，受到一氧化氮 (NO) 的调节，包括蛋白质中的半胱氨酸残基是一种改变蛋白质功能的翻译后修饰。我们证明了在低分子量硫醇（包括谷胱甘肽）上形成 SNO-谷胱甘肽（GSNO）。吸入 GSNO 会提高肺蛋白 -SNO，对哮喘有保护作用，鉴定出酶 SNO- 谷胱甘肽还原酶（GSNOR）使 GSNO 失活，并证明缺乏 GSNOR 的小鼠由于 β2AR 可以激活气道中的一氧化氮合成酶以产生一氧化氮，我们最近的工作需要了解这如何促进内源性 SNO 介导的支气管保护。已表明β2AR在激活后被S-亚硝基化，并通过点突变阻止SNO-β2AR增强重要的是，带有敲入该突变的 β2AR 的小鼠可以免受发育的影响。我们之前已经证明 β2AR 调节剂 GRK2 和 β-arrestin2 被 S-亚硝基化以抑制其哮喘。使 β2AR 脱敏的活性，并且携带 GRK2-C340S 和 β-arrestin2-C253S 敲入的小鼠表现出更高的活性 β2AR 活性和心脏模型中的急性损伤我们已经开始测试吸入 GSNO 对心脏模型的功效。影响支气管舒张和改善患者肺功能，这些数据和临床样本具有独特的使我们能够研究 S-亚硝基化在调节 β2AR 通路（从实验室到临床）中的作用。项目1的中心假设是β2AR信号系统是SNO-GSNO-的关键靶点和调节者 GSNOR 对严重哮喘气道的保护作用我们的研究将确定 S-亚硝基化的作用。小鼠哮喘模型中特定的 β2AR 信号通路成分，描绘了吸入 GSNO 的作用以及 GSNO 二亚硝基酶对小鼠哮喘模型和人肺中 β2AR 信号成分的影响原代细胞，并证明吸入 GSNO 可以改善气道流量和 β2 激动剂反应性这项工作通过提供机制联系补充了项目 2 和 3 的临床目标。重度哮喘中 NO/GSNO/GSNOR 作用与 β2AR 信号传导的调节之间的关系。