Reactive oxygen species and respiratory muscle dysfunction in heart failure

心力衰竭中的活性氧和呼吸肌功能障碍

• 批准号: 9005398
• 负责人: Leonardo Ferreira
• 金额: $ 37.5万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-01 至 2020-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9005398
• 关键词: Actins; Address; Affect; Animal Model; Animals; Antioxidants; Arrhythmia; Biological; Biology; Biopsy; Cardiac Death; Cells; Complex; Congestive Heart Failure; Cysteine; Data; Dithiothreitol; Dyspnea; Electron Transport; Enhancers; Environmental air flow; Event; Exposure to; Fatigue; Fiber; Functional disorder; Goals; Health; Heart failure; Human; Hydrogen Peroxide; In Vitro; Infarction; Injection of therapeutic agent; Intervention; Knock-out; Knockout Mice; Label; Lead; Limb structure; Measurement; Measures; Mediating; Mediator of activation protein; Messenger RNA; Mitochondria; Molecular; Molecular Target; Morbidity - disease rate; Mus; Muscle; Muscle Fibers; Myocardial Infarction; Myofibrils; Myosin ATPase; Myosin Light Chains; NADPH Oxidase; Operative Surgical Procedures; Organ; Oxidants; Oxidation-Reduction; Patients; Physical activity; Pilot Projects; Plasmids; Pleural; Pneumonia; Post-Translational Protein Processing; Prevention; Process; Protein Isoforms; Proteins; Proteomics; Reactive Oxygen Species; Recombinant adeno-associated virus (rAAV); Reducing Agents; Reflex action; Respiratory Diaphragm; Respiratory Muscles; Role; Saponins; Sarcolemma; Sarcoplasmic Reticulum; Skeletal Muscle; Source; Staging; Sulfhydryl Compounds; Testing; Tropomyosin; Up-Regulation; base; catalase; extracellular; heart dimension/size; human data; knock-down; mortality; novel; novel therapeutics; overexpression; oxidation; prevent; promoter; protein biomarkers; protein function; public health relevance; research study; sham surgery; small hairpin RNA; vasoconstriction; vector

 DESCRIPTION (provided by applicant): Diaphragm weakness is a significant health problem in chronic heart failure (CHF) patients because: a) compromises their ability to sustain ventilation and limits physical activity due to dyspnea, b) triggers sympathetic activation that ca lead to cardiac arrhythmias and death or induces vasoconstriction and fatigue in limb muscles, and c) impairs airway clearance, predisposing patients to pneumonia. These observations highlight the importance of understanding the mechanisms underlying diaphragm abnormalities and the need to identify biological targets for prevention of weakness in CHF. CHF diaphragm weakness is predominantly caused by loss of specific force (i.e. contractile dysfunction) due to alterations in myofibrillar proteins, and reactive oxygen species (ROS) have been implicated in this process. The mitochondria electron transport chain has been considered the main source of ROS in muscle. However, our preliminary data from human diaphragm biopsies and animal models suggest that NADPH oxidases (Nox) localized predominantly in diaphragm sarcolemma (Nox2) and mitochondria (Nox4) are involved in the heightened ROS and diaphragm weakness in CHF. Moreover, our pilot studies suggest that myofibrillar protein thiol oxidation is a key molecular event in CHF induced diaphragm weakness. Based on our data, we propose to complete three specific aims: 1) to determine the role of diaphragmatic Nox2 complex on excess ROS and weakness in CHF, 2) to determine whether Nox4 and mitochondrial ROS are mediators of diaphragm weakness in CHF, 3) to define the mechanism of ROS-induced diaphragm contractile dysfunction in CHF. To address these aims, we will use inducible skeletal muscle specific knockout mice and intra-pleural injection of recombinant adeno-associated virus with shRNA or expression plasmids under control of a skeletal muscle-specific promoter to target diaphragm fibers in Sham and CHF mice. We will also isolate diaphragm single fibers from sham and CHF mice and perform experiments to determine if specific force deficits can be rescued in vitro. Finally, we will use global label free proteomics and differential Cysteine labeling to determine the abundance of proteins and redox status of specific Cysteine residues in diaphragm from Sham and CHF animals. Our focus on ROS sources, molecular species and targets, and reversibility is critical to understand the pathophysiology and set the stage for nove therapies to treat diaphragm weakness that contributes to the morbidity and mortality in CHF patients.

 DESCRIPTION (provided by application): Diaphragm weakness is a significant health problem in chronic heart failure (CHF) patients because: a) compromises their ability to sustain ventilation and limits physical activity due to dyspnea, b) triggers sympathetic activation that ca lead to cardiac arrhythmias and death or induces vasoconstriction and fatigue in limb muscles, and c) impairs airway clearance, predisposing patients to 肺炎。这些观察结果突出了理解隔膜异常的机制的重要性，以及需要鉴定预防CHF无力的生物学靶标。 CHF diaphragm弱性主要是由于肌原纤维蛋白改变而导致的特定力损失（即收缩功能障碍）引起的，并且在此过程中已经实现了活性氧（ROS）。线粒体电子传输链被认为是肌肉中ROS的主要来源。然而，我们来自人隔膜活检的初步数据和动物模型表明，NADPH氧化物（NOX）主要在diaphragm骨s骨（NOX2）和线粒体（NOX4）中局部局部，与CHF中的ROS和diaphragm弱点有关。此外，我们的试点研究表明，肌原纤维蛋白氧化物氧化物是CHF诱导的隔膜弱点的关键分子事件。根据我们的数据，我们建议完成三个具体目的：1）确定diaphragmatic nox2复合物对ROS过多和无力的弱点的作用，2）确定NOX4和线粒体ROS是否是diaphragm弱点的介体在CHF中的介体，3）以定义ROS诱导的diaphragragm conscontile conscontile conscontileile conscontileile conscontileile conscontifile conscontifile conscontion conff。为了解决这些目的，我们将使用诱导的骨骼肌特异性敲除小鼠，并在控制骨骼肌特异性启动子的控制下，将重组腺相关病毒与SHRNA或表达质粒对靶向糖尿皮纤维的控制在SHAM和CHF小鼠中。我们还将从假和CHF小鼠中分离隔膜单纤维，并执行实验以确定是否可以在体外检索特定的力。最后，我们将使用全球无标记的无蛋白质组学和差异半胱氨酸标记来确定蛋白质的抽象和隔膜中特定半胱氨酸残基的氧化还原状态。我们对ROS来源，分子物种和靶标的关注以及可逆性对于了解病理生理学和治疗diaphragragm弱点的新疗法的阶段至关重要，这有助于CHF患者的发病率和死亡率。