In Vivo Molecular Probes for the Membrane Repair Pathway in Muscle

肌肉膜修复途径的体内分子探针

基本信息

批准号：
9060873
负责人：
Daniel E Michele
金额：
$ 17.11万
依托单位：
UNIVERSITY OF MICHIGAN AT ANN ARBOR
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-08-01 至 2018-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9060873
关键词：
Actins Animals Binding Biological Assay Calcium Cell membrane Cells Cessation of life Communities Complex Confocal Microscopy Cytoskeleton DYSF gene Data Deterioration Disease Outcome Dyes Dystrophin Early Ambulation Engineering Event Face Family Fiber Future Genes Glycoproteins Goals Gold Health Hereditary Disease Human Inherited Injury Knockout Mice Label Lasers Lesion Life Limb-Girdle Muscular Dystrophies Lipid Binding Measures Mechanics Membrane Membrane Protein Traffic Methodology Methods Mitotic Modeling Molecular Molecular Probes Monitor Mus Muscle Muscle Cells Muscle Fibers Muscle Weakness Muscular Dystrophies Mutation Myopathy Natural regeneration Organelles Outcome Pathway interactions Patients Peptides Phospholipids Physiological Process Proteins Recruitment Activity Recycling Reporter Research Respiratory physiology Role Sarcolemma Series Site Striated Muscles Testing Therapeutic Time Total Internal Reflection Fluorescent Transgenic Organisms Vesicle Work base cell injury gamma Actin improved in vivo injury and repair link protein loss of function loss of function mutation member mouse model muscle degeneration muscle form novel prevent repaired research study response sensor synaptotagmin targeted treatment tool trafficking two-photon uptake wound

项目摘要

DESCRIPTION (provided by applicant): Muscular dystrophies, particularly those associated with disruption of the function of the dystrophin glycoprotein complex, are characterized by muscles that are sensitive to mechanical damage. A key feature in the mechanism of muscle damage is the loss of muscle fiber plasma membrane integrity which ultimately results in severe muscle fiber degeneration, loss of muscle fibers, and consequently loss of muscle mass and progressive weakness. Post-mitotic muscle fibers, like many cells, have a remarkable capacity to repair membrane lesions, but little is known about the mechanisms of membrane repair in muscle fibers. Mutations in the protein dysferlin, are associated with LGMD 2B and Myoshi Myopathy in humans. Due to the similarity of dysferlin to other ferlins and synaptotagmin, dysferlin is believed to be involved in membrane trafficking. Loss of dysferlin in mice appears to disrupt the normal membrane repair pathway. One of the major limitations in studying membrane repair is that most of the approaches to study membrane repair are indirect, only showing how well membranes can effectively exclude membrane impermeant probes but doesn't necessarily differentiate differences in the magnitude of membrane wounding versus differences in efficiency of repair. The overall goal of this proposal is to develop a novel set of live cell molecular probes to specifically label the membrane repair pathway in live muscle fibers and muscles, and study the mechanisms of membrane trafficking directly in muscle fibers in response to experimental and physiological injury. Our preliminary data using one of these novel reporters challenges the current model that pre-existing vesicle or organelle compartments containing dysferlin are responsible for repairing plasma membrane lesions. Therefore, the proposed work will focus on two major aims: 1) Develop a set of live cell molecular probes in living mice to study the membrane repair pathway under experimental and physiological muscle injury. 2) Dissect how the submembrane actin cytoskeleton contributes to membrane repair by recruiting dysferlin to the site of membrane injury. The long term goal is to identify the mechanisms of membrane repair that hopefully can be exploited or enhanced in order to repair muscle damage in several forms of inherited muscular dystrophy.

描述（由申请人提供）：肌营养不良症，特别是与肌营养不良蛋白糖蛋白复合物功能破坏相关的肌营养不良症，其特征是肌肉对机械损伤敏感，肌肉损伤机制的一个关键特征是肌纤维的损失。质膜完整性最终导致严重的肌纤维变性、肌纤维损失以及由此导致的肌肉质量损失和进行性无力。像许多细胞一样，有丝分裂后肌纤维具有修复膜损伤的显着能力，但人们知之甚少。关于Dysferlin 蛋白的突变与人类 LGMD 2B 和 Myoshi 肌病有关，因为 Dysferlin 与其他 ferlin 和突触结合蛋白相似，因此人们认为 Dysferlin 参与了膜运输的丢失。小鼠体内的 Dysferlin 似乎会破坏正常的膜修复途径，研究膜修复的主要局限性之一是大多数研究膜修复的方法都是间接的，只能显示膜如何有效排除。膜不透性探针，但不一定区分膜损伤程度的差异与修复效率的差异。该提案的总体目标是开发一套新颖的探针。活细胞分子探针专门标记活肌肉纤维和肌肉中的膜修复途径，并研究直接在肌肉纤维中响应实验和生理损伤的膜运输机制，我们使用这些新型生产者之一的初步数据挑战了当前的模型。预先存在的含有dysferlin的囊泡或细胞器区室负责修复质膜损伤，因此，拟议的工作将集中于两个主要目标：1）在活体小鼠中开发一套活细胞分子探针，以研究膜修复途径。实验和生理学2) 剖析膜下肌动蛋白细胞骨架如何通过将 Dysferlin 募集到膜损伤部位来促进膜修复。长期目标是确定有望利用或增强的膜修复机制来修复肌肉损伤。多种形式的遗传性肌营养不良症。