Biomechanical characterization of striated muscle cells from R155C VCP knock-in and W2710X filamin C knock-in mice: a novel approach to understand the pathogenesis of myofibrillar myopathies

R155C VCP 敲入和 W2710X filamin C 敲入小鼠横纹肌细胞的生物力学特征：了解肌原纤维肌病发病机制的新方法

• 批准号: 251281920
• 负责人: Professor Dr. Ben Fabry
• 金额: --
• 依托单位: Lehrstuhl für Physikalisch-Medizinische Technik
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2014
• 资助国家: 德国
• 起止时间: 2013-12-31 至 2017-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/251281920?language=en
• 关键词: Biomechanical; characterization; striated; muscle; cells

Myofibrillar myopathies (MFMs) are a group of sporadic and hereditary skeletal and cardiac muscle diseases that lead to severe physical disability and premature death. MFMs are caused by mutations in genes encoding desmin, filamin C, plectin, VCP, FHL1, ZASP, myotilin, alpha-B-crystallin, and BAG3. First results from our biomechanical studies on primary human myoblasts carrying desmin -and plectin mutations showed an increased stiffness and reduced mechanical stress tolerance in the form of higher mechanical vulnerability compared to control cells. We hypothesize that the higher stiffness of mutant cells leads to higher intracellular stress at physiologic stretch and shear deformations, which in turn triggers muscle fiber degeneration. In the present project, we will test this hypothesis using immortalized myoblast cells obtained from two MFM mouse models. Through the DFG research consortium FOR1228, we have access to two knock-in mouse models (R155C VCP, and W2710X filamin C), which harbor the most frequent human pathogenic VCP and filamin C mutations. Using traction force microscopy, magnetic tweezer microrheology, and a cell stretcher together with high resolution (temporal and spatial) confocal microscopy, we will address two key questions: (i) what is the influence of these mutations on the biomechanical function of cultured myoblasts and myotubes derived from skeletal muscle tissue, and (ii) what are the molecular processes that lead to altered mechanical stress tolerance in these cells. This project will provide the first insight into the biomechanical aspects of the pathogenesis of VCP- and filamin C-related myopathies.

肌原纤维性肌病（MFM）是一组散发性和遗传性骨骼肌和心肌疾病，可导致严重的身体残疾和过早死亡。MFM是由编码结蛋白、细丝蛋白C、网蛋白、VCP、FHL 1、ZASP、肌球蛋白、α-B-晶状体蛋白和BAG 3的基因突变引起的。我们对携带结蛋白和网蛋白突变的原代人成肌细胞的生物力学研究的第一个结果显示，与对照细胞相比，刚度增加，机械应力耐受性降低，机械脆弱性更高。我们假设突变细胞的更高刚度导致在生理拉伸和剪切变形时更高的细胞内应力，这反过来又引发肌纤维变性。在本项目中，我们将使用从两个MFM小鼠模型中获得的永生化成肌细胞来验证这一假设。通过DFG研究联盟FOR 1228，我们获得了两种敲入小鼠模型（R155 C VCP和W2710 X细丝蛋白C），它们具有最常见的人类致病性VCP和细丝蛋白C突变。使用牵引力显微镜，磁镊子微观流变学，和细胞担架连同高分辨率（时间和空间）共聚焦显微镜，我们将解决两个关键问题：（i）这些突变对培养的成肌细胞和肌管的生物力学功能的影响来自骨骼肌组织，和（ii）是什么样的分子过程，导致改变这些细胞的机械应力耐受性。该项目将提供第一次深入了解VCP和细丝蛋白C相关肌病发病机制的生物力学方面。