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

肌联蛋白在机械通气期间膈肌无力病理生理学中的作用

基本信息

批准号：
10659578
负责人：
Coen Ottenheijm
金额：
$ 55.61万
依托单位：
UNIVERSITY OF ARIZONA
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-01-01 至 2027-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10659578
关键词：
Actins Adopted Affect Animal Model Atrophic Attention Attenuated Binding Biochemical Biological Assay Biopsy Contractile Proteins Data Depressed mood Disease Elasticity Failure Filament Functional disorder Genetic Engineering Goals Hand Health Heart Diseases Hour Individual Laparotomy Length Light Lung Mass Spectrum Analysis Measures Mechanical ventilation Mechanics Mental Depression Microfilaments Modeling Motor Mus Muscle Muscle Proteins Muscular Atrophy Myosin ATPase Oxygen Patients Pharmaceutical Preparations Phosphorylation Play Positioning Attribute Post-Translational Protein Processing Production Protein Dephosphorylation Proteins Proteomics Rattus Relaxation Research Respiration Respiratory Diaphragm Respiratory Muscles Role Running Sarcomeres Skeletal Muscle Testing Therapeutic Ventilator Weaning Vertebral column Weaning Work X ray diffraction analysis clinically relevant connectin experimental study in vivo innovation insight interdisciplinary approach mechanical load mouse model nanometer resolution novel preservation programs response tool uptake ventilation

项目摘要

Summary Our long-term goal is to understand diaphragm contractility in health and disease, and the role of titin therein. The diaphragm muscle drives respiration and is constantly subjected to mechanical loading. Changes in mechanical loading rapidly affect diaphragm contractility, e.g., within hours of diaphragm unloading during mechanical ventilation (MV) in ICU patients, severe diaphragm weakness ensues, contributing to difficult ventilator weaning. The pathophysiology of diaphragm weakness is incompletely understood. In the proposal at hand, we will study the role of depressed interactions between myosin and actin, the two key contractile proteins in myofibers, and the role of titin in force depression. Our pilot data suggest that unloading of the diaphragm causes the myosin motors to adopt the so-called ‘super-relaxed state’ (SRX). Once in the SRX state, less myosin motors are available for binding to actin, and less force can be generated. We will also study the role of the giant protein titin in releasing myosin from the SRX state and investigate whether during MV-induced unloading of the diaphragm, this mechanism is perturbed. Aim 1 will determine the SRX state of myosin in the diaphragm of ventilated ICU patients. We will use our diaphragm biopsies of ICU patients to study the contractile force of diaphragm myofibers and determine the number of myosin motors that attach to actin during activation. We will combine mechanics with X-ray diffraction and biochemical assays to resolve with nanometer resolution the position of myosin motors relative to actin during activation and the proportion of myosin in the SRX state. In Aim 2 we will determine the role of posttranslational modifications in the SRX of myosin. Phosphorylation of regulatory light chains (RLC) regulates the SRX state of myosin and we will apply mass-spectrometry on the diaphragm biopsies to determine the phospho-proteome, and we will establish whether there is a cause-and-effect relation between RLC phosphorylation and SRX by performing RLC exchange experiments into patients’ myofibers. To critically test whether changes in RLC are caused by diaphragm inactivity, we will ventilated healthy rats, and study RLC phosphorylation. Aim 3 will determine whether, in addition to RLC phosphorylation, direct mechanical effects of titin on myosin induce SRX. Titin-based passive tension strains the myosin filament which regulates the SRX state. We will study whether the reduction in titin-based passive tension, due to diaphragm unloading during MV, reduces the strain in the myosin filament and increases SRX. We will use mouse models with genetically engineered increased or decreased titin-based passive tension and study the effect of MV on the SRX state of myosin. The innovation of this proposal lies in the novel research foci and guiding hypotheses, unique diaphragm biopsies from patients, and its novel tools and mouse models, The proposal’s integrative approach is expected to lead to a major step forward in our understanding of diaphragm function and titin’s role therein.

概括我们的长期目标是了解健康和疾病中的膈肌收缩力，以及肌联蛋白在其中的作用。膈肌驱动呼吸并不断承受机械负荷的变化。机械负荷会迅速影响膈肌收缩力，例如，在隔膜卸载后的数小时内 ICU 患者进行机械通气 (MV) 时，会出现严重的膈肌无力，导致呼吸困难呼吸机脱机的提案尚未完全了解膈肌无力的病理生理学。另一方面，我们将研究肌球蛋白和肌动蛋白（两种关键的收缩蛋白）之间抑制相互作用的作用肌纤维中的肌纤维，以及肌动蛋白在力抑制中的作用，我们的试验数据表明隔膜的卸载。导致肌球蛋白马达进入所谓的“超级放松状态”(SRX)，一旦进入 SRX 状态，肌球蛋白就会减少。电机可用于与肌动蛋白结合，并且可以产生较小的力我们还将研究巨人的作用。蛋白肌联从 SRX 状态释放肌球蛋白，并研究在 MV 诱导的肌球蛋白卸载过程中是否隔膜，该机制受到干扰。目标 1 将确定 ICU 通气患者膈肌中肌球蛋白的 SRX 状态。对 ICU 患者进行膈肌活检，研究膈肌纤维的收缩力并确定激活期间附着于肌动蛋白的肌球蛋白马达的数量我们将把力学与 X 射线衍射结合起来。和生化测定以纳米分辨率解析肌球蛋白马达相对于肌动蛋白的位置在激活过程中以及处于 SRX 状态的肌球蛋白的比例中，我们将确定的作用。调节轻链 (RLC) 的 SRX 的翻译后修饰。调节肌球蛋白的 SRX 状态，我们将对隔膜活检应用质谱分析以确定磷酸化蛋白质组，我们将确定 RLC 之间是否存在因果关系通过对患者的肌纤维进行 RLC 交换实验来严格测试磷酸化和 SRX。 RLC的变化是否是由膈肌不活动引起的，我们将对健康大鼠进行通气，并研究RLC 目标 3 将确定除了 RLC 磷酸化之外，是否还包括直接机械磷酸化。肌球蛋白对肌球蛋白的影响诱导基于肌球蛋白的被动张力使调节肌球蛋白丝紧张。我们将研究基于肌动蛋白的被动张力是否会因隔膜卸载而减少。在 MV 期间，减少肌球蛋白丝的应变并增加 SRX 我们将使用小鼠模型。基因工程增加或减少基于 titin 的被动张力，并研究 MV 对肌球蛋白的 SRX 状态。该提案的创新之处在于新颖的研究焦点和指导假设、独特的膈肌活检来自患者及其新颖的工具和小鼠模型，该提案的综合方法预计将引领我们对膈肌功能和肌动蛋白在其中的作用的理解向前迈出了一大步。

项目成果

期刊论文数量（18）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Unaffected contractility of diaphragm muscle fibers in humans on mechanical ventilation.

机械通气时人体膈肌纤维的收缩力不受影响。

DOI：
发表时间：
2014-09-15
期刊：
影响因子：
0
作者：
Hooijman, Pleuni E;Paul, Marinus A;Stienen, Ger J M;Beishuizen, Albertus;Van Hees, Hieronymus W H;Singhal, Sunil;Bashir, Muhammad;Budak, Murat T;Morgen, Jacqueline;Barsotti, Robert J;Levine, Sanford;Ottenheijm, Coen A C
通讯作者：
Ottenheijm, Coen A C

Diaphragm contractile weakness due to reduced mechanical loading: role of titin.

由于机械负荷减少而导致膈肌收缩无力：肌动蛋白的作用。

DOI：
发表时间：
2019
期刊：
影响因子：
0
作者：
van der Pijl, Robbert J;Granzier, Henk L;Ottenheijm, Coen A C
通讯作者：
Ottenheijm, Coen A C

Diaphragm muscle fiber weakness and ubiquitin-proteasome activation in critically ill patients.

危重患者膈肌纤维无力和泛素蛋白酶体激活。

DOI：
发表时间：
2015-05-15
期刊：
影响因子：
24.7
作者：
Hooijman, Pleuni E;Beishuizen, Albertus;Witt, Christian C;de Waard, Monique C;Girbes, Armand R J;Spoelstra;Niessen, Hans W M;Manders, Emmy;van Hees, Hieronymus W H;van den Brom, Charissa E;Silderhuis, Vera;Lawlor, Michael
通讯作者：
Lawlor, Michael

Diaphragm Pathology in Critically Ill Patients With COVID-19 and Postmortem Findings From 3 Medical Centers.

COVID-19 危重患者的膈肌病理学和 3 个医疗中心的尸检结果。

DOI：
发表时间：
2021
期刊：
影响因子：
39
作者：
Shi, Zhonghua;de Vries, Heder J;Vlaar, Alexander P J;van der Hoeven, Johannes;Boon, Reinier A;Heunks, Leo M A;Ottenheijm, Coen A C;Dutch COVID
通讯作者：
Dutch COVID

Preserved single muscle fiber specific force in facioscapulohumeral muscular dystrophy.

面肩肱型肌营养不良症中保留的单肌纤维比力。

DOI：
发表时间：
2020
期刊：
影响因子：
9.9
作者：
Lassche, Saskia;Voermans, Nicol C;van der Pijl, Robbert;van den Berg, Marloes;Heerschap, Arend;van Hees, Hieronymus;Kusters, Benno;van der Maarel, Silvère M;Ottenheijm, Coen A C;van Engelen, Baziel G M
通讯作者：
van Engelen, Baziel G M