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: 9199443
• 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/9199443
• 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; Impairment; Intensive Care Units; Knockout Mice; Lead; Link; Location; Lung; Lung diseases; Mechanical Stress; Mechanical ventilation; Mechanics; Mediating; Microfilaments; Modeling; Molecular; Morphology; Mus; Muscle; Mutation; Oxygen; Patients; Periodicity; 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; clinically relevant; connectin; innovation; mechanical load; mechanotransduction; mortality; mouse model; muscle form; novel; novel therapeutics; preconditioning; protein degradation; public health relevance; response; sensor; tool; uptake

DESCRIPTION (provided by applicant): 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. Th search for the molecular triggers for the development of diaphragm weakness during mechanical unloading is ongoing. The potential role of mechano-sensor proteins, that link diaphragm unloading to protein turnover, is unexplored but is an exciting and novel concept that needs to be studied. A candidate mechano-sensor 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 musle 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 mechano-sensing 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.

描述（由申请人提供）：该提案的长期目标是详细了解膈肌（呼吸的主要肌肉）如何响应机械卸载而迅速减弱，以及巨型弹性蛋白肌联蛋白影响这一机制的机制回复。 隔膜是一种独特的肌肉，因为它不断承受机械负荷。最近的研究表明，膈肌强度对机械卸载非常敏感，就像 ICU 机械通气期间发生的那样。人们对卸载如何影响隔膜强度知之甚少。加深这种理解至关重要：机械通气期间膈肌卸载在数小时内会导致 ICU 患者膈肌无力，从而导致患者难以脱离通气支持并导致死亡。机械卸载过程中膈肌无力发展的分子触发因素的研究仍在进行中。将隔膜卸载与蛋白质周转联系起来的机械传感器蛋白质的潜在作用尚未被探索，但这是一个令人兴奋的新颖概念，需要研究。一种候选的机械传感器是肌联蛋白，这是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。阐明肌动蛋白在机械通气期间膈肌营养性和膈肌无力中的作用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），这种情况在临床上很重要，并且是这项工作的一个很好的工具，因为它会导致肌肉快速肥大。 由于周期性被动拉伸而失去神经的半膈肌。我将在两种新型肌联蛋白 KO 小鼠模型中研究 UDD：一种是通过删除 Ig 结构域 (Ig KO) 来增加肌联蛋白硬度，另一种是通过删除肌联蛋白剪接因子 rbm20 (Rbm20 KO) 来降低肌联蛋白硬度。我预计 UDD 后的肥大反应在 Ig KO 小鼠中被夸大，而在 Rmb20 KO 小鼠中减弱，并且这种反应是由改变的 titin 信号介导的。目标 2 将研究低肌动蛋白刚度是​​否可以保护膈肌在机械通气引起的卸载过程中免受削弱，并将使用低肌动蛋白刚度的大鼠模型。如果基于肌动蛋白的机械传感介导隔膜对机械卸载的响应，那么我预计，通过预处理隔膜以减少机械传感，低肌动蛋白刚度会减弱这种响应。目标 3 将首次使用从 ICU 机械通气患者活检中分离出的膈肌纤维来研究机械通气 ICU 患者膈肌无力的机制基础。目的是调查动物研究的结果是否可以推断到患者身上。 该提案的创新之处在于具有创新指导假设的新研究焦点、新颖的小鼠模型、来自机械通气ICU患者的独特隔膜活检以及新颖的实验工具。该提案的综合方法预计将在我们对膈肌功能和肌联蛋白在其中的作用的理解方面向前迈出重要一步。