Role of titin in the pathophysiology of diaphragm weakness during mechanical vent

肌联蛋白在机械通气期间膈肌无力病理生理学中的作用

• 批准号: 8614046
• 负责人: Coen Ottenheijm
• 金额: $ 37.12万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-01-01 至 2018-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8614046
• 关键词: Affect; Animal Model; Animals; Antibodies; Applications Grants; Atrophic; Binding Proteins; Biophysics; Biopsy; Breathing; Clinical; Data; Denervation; Development; Environmental air flow; Failure; Fiber; Functional disorder; Gene Targeting; Genetic Engineering; Goals; Growth; Hour; Human; Hypertrophy; Intensive Care Units; Lead; Link; Location; Lung; Lung diseases; Mechanical Stress; Mechanical ventilation; Mechanics; Mediating; Microfilaments; Modeling; Molecular; Morphology; Mus; Muscle; Mutation; Oxygen; Patients; Pilot Projects; Play; Property; Proteins; RNA Recognition Motif; RNA Splicing; Rattus; Research; Respiration; Respiratory Diaphragm; Roentgen Rays; Role; Signal Pathway; Signal Transduction; Stretching; Structure; Testing; Time; Vent; Weaning; Work; X ray diffraction analysis; X-Ray Diffraction; base; connectin; innovation; link protein; mortality; mouse model; muscle form; novel; novel therapeutics; preconditioning; protein degradation; public health relevance; response; sensor; tool; uptake

7. Summary. The long-term goal of this proposal is to gain detailed understanding of how the diaphragm - the main muscle of respiration - rapidly weakens in response to mechanical unloading and of the mechanisms whereby the giant elastic protein titin influences this response. The diaphragm is a unique muscle in that it is constantly subjected to mechanical loading. Recent work suggests that diaphragm strength is remarkably sensitive to mechanical unloading, as occurs during mechanical ventilation in the ICU. How unloading affects diaphragm strength is poorly understood. Increasing this understanding is critically important: within hours, diaphragm unloading during mechanical ventilation causes diaphragm weakness in ICU patients, which leads to difficulties in weaning patients from ventilatory support and contributes to mortality. The search for the molecular triggers for the development of diaphragm weakness during mechanical unloading is ongoing. The potential role of mechanosensor proteins, that link diaphragm unloading to protein turnover, is unexplored but is an exciting and novel concept that needs to be studied. A candidate mechanosensor is titin, a giant elastic protein that has been suggested to sense mechanical stress and link this to trophic signalling pathways. The elucidation of titin's role in diaphragm trophicity and in diaphragm weakness during mechanical ventilation is central to this grant proposal. Aim 1 will critically test whether titin affects muscle trophicity. I will use unilateral diaphragm denervation (UDD), a condition that is clinically important and that presents itself as a great tool for this work as it induces rapid hypertrophy of the denervated hemidiaphragm due to cyclic passive stretch. I will study UDD in two novel titin KO mouse models: one in which titin stiffness is increased through deletion of Ig domains (Ig KO) and another in which titin stiffness is decreased through deletion of the titin splice factor rbm20 (Rbm20 KO). I anticipate that the hypertrophic response following UDD is exaggerated in Ig KO mice and blunted in Rmb20 KO mice, and that this response is mediated by altered titin signaling. Aim 2 will study whether low titin stiffness protects the diaphragm from weakening during mechanical ventilation-induced unloading and will use a rat model with low titin stiffness. If titin-based mechanosensing mediates the response of the diaphragm to mechanical unloading, then I anticipate that low titin stiffness, by preconditioning the diaphragm to reduced mechanosensing, blunts this response. Aim 3 will study the mechanistic basis for diaphragm weakness in mechanically ventilated ICU patients using, for the first time, diaphragm fibers isolated from biopsies of mechanically ventilated ICU patients. The goal is to investigate whether the findings of animal studies extrapolate to patients. The innovation of this proposal lies in the novel research foci with innovative guiding hypotheses, its novel mouse models, unique diaphragm biopsies from mechanically ventilated ICU patients, and its novel experimental tools. The proposal's integrative approach is expected to lead to a significant step forward in our understanding of diaphragm function and the role of titin therein.

7. 总结。 该提案的长期目标是详细了解隔膜（主要肌肉）如何 呼吸作用 - 响应机械卸载和呼吸机制而迅速减弱 巨大的弹性蛋白肌联蛋白影响这种反应。 隔膜是一种独特的肌肉，因为它不断承受机械负荷。最近的工作 表明隔膜强度对机械卸载非常敏感，如在 ICU 内的机械通气。人们对卸载如何影响隔膜强度知之甚少。增加 这种理解至关重要：在机械通气过程中，数小时内隔膜就会卸载 导致 ICU 患者膈肌无力，导致患者脱机困难 支持并导致死亡率。寻找膈肌发育的分子触发因素 机械卸载期间的弱点仍在持续。机械传感器蛋白的潜在作用，将 隔膜卸载到蛋白质周转尚未被探索，但这是一个令人兴奋且新颖的概念，需要加以研究 研究过。一种候选的机械传感器是肌联蛋白，一种巨大的弹性蛋白，被认为可以感知 机械应力并将其与营养信号通路联系起来。肌联蛋白在膈肌中的作用的阐明 机械通气期间的营养性和膈肌无力是本拨款提案的核心。 目标 1 将严格测试肌联蛋白是否影响肌肉营养性。我将使用单侧膈肌去神经术 （UDD），一种临床上很重要的疾病，它本身就是这项工作的一个很好的工具，因为它会引起 由于周期性被动拉伸，去神经支配的膈肌迅速肥大。我会在两本小说中学习UDD 肌动蛋白 KO 小鼠模型：一种通过删除 Ig 结构域 (Ig KO) 来增加肌动蛋白硬度的小鼠模型 另一种方法是通过删除肌动蛋白剪接因子 rbm20 (Rbm20 KO) 来降低肌动蛋白硬度。我 预计 UDD 后的肥大反应在 Ig KO 小鼠中被夸大，并在 20 元人民币后减弱 KO 小鼠，并且这种反应是由改变的肌动蛋白信号传导介导的。目标 2 将研究是否存在低肌蛋白 刚度可保护隔膜在机械通气引起的卸载过程中免受削弱，并将使用 具有低肌动蛋白硬度的大鼠模型。如果基于肌动蛋白的机械传感介导隔膜对 机械卸载，然后我预计通过预处理隔膜来降低钛刚度 机械传感会削弱这种反应。目标 3 将研究膈肌无力的机制基础 机械通气 ICU 患者首次使用从活检组织中分离出的隔膜纤维 ICU 机械通气患者。目的是调查动物研究的结果是否 推断到患者身上。 本方案的创新点在于研究重点新颖、指导假设新颖、 小鼠模型、来自机械通气 ICU 患者的独特隔膜活检及其新颖 实验工具。该提案的综合方法预计将在以下方面向前迈出重要一步： 我们对膈肌功能和肌联蛋白在其中的作用的理解。