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: 8982039
• 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/8982039
• 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; Health; Hour; Human; Hypertrophy; 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; 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; 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)：这项提案的长期目标是详细了解横隔膜--呼吸的主要肌肉--如何在机械卸载时迅速减弱，以及巨大的弹性蛋白质titin如何影响这种反应的机制。 横隔膜是一种独特的肌肉，因为它不断地受到机械载荷的作用。最近的研究表明，隔膜强度对机械卸载非常敏感，就像在ICU机械通风期间发生的那样。卸载是如何影响横隔膜强度的，目前还知之甚少。提高这一认识是至关重要的：在几个小时内，机械通气过程中的隔膜卸载会导致ICU患者的隔膜薄弱，从而导致患者难以脱离呼吸机支持，并可能导致死亡。在机械卸载过程中，寻找导致横隔膜薄弱的分子诱因正在进行中。机械感受器蛋白将隔膜卸载与蛋白质周转联系起来，其潜在作用尚未被探索，但这是一个令人兴奋的新概念，需要研究。一种候选的机械传感器是肌动蛋白，它是一种巨大的弹性蛋白质，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风期间横隔膜营养丰富和横隔膜薄弱中的作用是这项赠款提案的核心。目标1将严格测试肌动蛋白是否影响肌肉的营养活性。我将使用单侧隔膜去神经(UDD)，这是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，因为它会导致快速肥大的 周期性被动牵拉所致的失神经横隔膜。我将在两个新的titin KO小鼠模型中研究UDD：一个通过缺失Ig结构域(Ig KO)来增加ittin硬度，另一个通过缺失titin剪接因子RBM20(RBM20 KO)来降低ittin硬度。我预计UDD后的肥大反应在Ig KO小鼠中被夸大，在RMB20 KO小鼠中被钝化，并且这种反应是由改变的肌动蛋白信号介导的。目的2研究低Titin硬度是否能在机械通气诱导的卸荷过程中保护横隔膜免受削弱，并将使用低Titin硬度的大鼠模型。如果基于Titin的机械传感调节了横隔膜对机械卸载的反应，那么我预计低的Titin硬度，通过减少机械传感来预先调节横隔膜，使这种反应变钝。目的3首次利用从ICU患者的活检组织中分离的横隔膜纤维，研究机械通气ICU患者横隔膜无力的机制基础。目的是调查动物研究的结果是否适用于患者。 这一方案的创新之处在于具有创新的指导假设的新的研究重点，其新颖的小鼠模型，来自机械通气的ICU患者的独特的横隔膜活检，以及其新颖的实验工具。该提案的综合方法有望使我们对横隔膜功能和Titin在其中的作用的理解向前迈进一大步。