DAPC structural adaption in regulating nanotopography-responsive myotube orientation

DAPC结构适应调节纳米形貌响应肌管方向

• 批准号: 10592954
• 负责人: JOACHIM D MUELLER
• 金额: $ 35.42万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-20 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10592954
• 关键词: Affect; Age; Biochemical; Biocompatible Materials; Brain; Cells; Cessation of life; Cues; Cytoskeleton; Data; Dimensions; Disease; Duchenne muscular dystrophy; Dystrophin; Dystrophin-Associated Protein Complex; Engineering; Exhibits; Extracellular Matrix; Foundations; Future; Genetic Diseases; Heart failure; Human; Individual; Integral Membrane Protein; Knowledge; Laminin; Lateral; Length; Link; Location; Mechanics; Mediating; Microscopy; Muscle; Muscle Cells; Muscle Fibers; Muscle function; Muscular Dystrophies; Mutation; Myofibrils; Myopathy; Nanotopography; Neuromuscular Diseases; Organ; Outcome; Patients; Pattern; Phenotype; Positioning Attribute; Proteins; Respiratory Failure; Role; Signal Transduction; Technology; Testing; Tissues; Width; cell behavior; drug development; imaging modality; induced pluripotent stem cell; insight; matrigel; mechanotransduction; nanoarchitecture; nanocluster; nanoscale; new technology; novel; phenotypic biomarker; protein complex; renal epithelium; response; single molecule; submicron; superresolution imaging; therapy development; transmission process; ultra high resolution

Project Summary The Dystrophin-Associated-Protein-Complex (DAPC) is an important transmembrane protein complex that structurally, mechanically, and functionally links cytoskeleton and the extracellular matrix and serves as both mechanical and signaling hubs in regulating muscle and non-muscle cells. Mutations affecting DAPC components are associated with a wide range of diseases such as muscular dystrophies. Although there is a wealth of knowledge on the DAPC composition, the mechanisms underlying how the DAPC senses and adapts to biochemical, mechanical, and topographic cues in cell microenvironments and consequently regulates cell phenotypes and functions remain unknown. We recently discovered that substrates patterned with submicron ridges/grooves and functionalized with Matrigel or laminin present an engineered cell microenvironment to allow myotubes derived from non-diseased human induced pluripotent stem cells (hiPSCs) to align nearly perpendicular to the ridges, while myotubes derived from less-affected and severely-affected Duchenne Muscular Dystrophy (a genetic disorder resulting from mutations in dystrophin and often leading to death at an age of 20-30s due to cardiac and respiratory failure) cells exhibit prominent differences in alignment and orientation, providing a sensitive phenotypic biomarker to distinguish these cells, which may serve as a phenotypic readout for high throughput therapy development. Our preliminary data suggest that this nanotopography-responsive myotube orientation is regulated by the DAPC; however, details of how nanotopography regulates myotube orientation through the DAPC are unclear. We hypothesize that the perpendicular myotube orientation is caused by structural adaption of the anisotropic DAPC on the nanotopography to remain its stability. We expect that super-resolution Single Molecule Localization Microscopy (SMLM) will enable us to examine the DAPC nanoarchitecture and its structural adaption in response to nanotopography and verify our hypothesis. Successful accomplishment of this project will reveal unprecedented nanoscale details of the DAPC, DAPC structural adaption on nanotopography, and the mechanism underlying the nanotopography-responsive myotube orientation, which are all unknown currently. This project will lay the foundation for future studies that combine SMLM and biomaterials engineering to elucidate more mechanistic details underlying DAPC-mediated force transmission, mechanosensing, and mechanochemical transduction in normal muscle function and in various muscular and neuromuscular disorders. Verification of the role of the DAPC in regulating nanotopography-responsive myotube orientation may extend it as a phenotypic biomarker for many other muscular dystrophies associated with defective DAPC components. This study will also reveal novel engineering approaches to regulate cell behavior and cell fate through topographic control of the DAPC.

项目摘要 肌营养不良蛋白相关的蛋白质复合物（DAPC）是一种重要的跨膜蛋白复合物 在结构，机械上和功能上连接细胞骨架和细胞外基质，并用作两者兼而有之 调节肌肉和非肌肉细胞的机械和信号枢纽。影响DAPC的突变 成分与多种疾病有关，例如肌肉营养不良。虽然有一个 关于DAPC组成的丰富知识，DAPC如何适应的机制 细胞微环境中的生化，机械和地形线索，因此调节细胞 表型和功能仍然未知。我们最近发现，用亚微米构图的底物 脊/凹槽，用矩阵或层粘连蛋白功能化，将工程的细胞微环境提交给 允许源自未切除的人类诱导多能干细胞（HIPSC）的肌管几乎对齐 垂直于山脊，而肌管则来自影响较小且受到严重影响的Duchenne 肌肉营养不良（由肌营养不良蛋白突变引起的遗传疾病，通常导致死亡 由于心脏和呼吸衰竭而导致的20-30岁）细胞在对齐和 方向，提供敏感的表型生物标志物来区分这些细胞，这些细胞可以用作 高通量疗法开发的表型读数。我们的初步数据表明这一点 DAPC调节了纳米形态响应的肌管方向；但是，详细信息如何 通过DAPC调节肌管方向的纳米形造影尚不清楚。我们假设 垂直肌管的方向是由各向异性DAPC在该上的结构适应引起的 纳米造影保持其稳定性。我们期望超分辨率的单分子定位 显微镜（SMLM）将使我们能够检查DAPC纳米结构及其结构适应 对纳米形态的反应并验证我们的假设。这个项目的成功完成将揭示 DAPC的前所未有的纳米级细节，DAPC结构适应纳米形态以及 纳米形态响应性肌管方向的基础机制，目前都是未知的。 该项目将为将来的研究奠定基础，将SMLM和生物材料工程结合到 阐明DAPC介导的力传输，机械感应和 正常肌肉功能以及各种肌肉和神经肌肉的机械化学转导 疾病。验证DAPC在调节纳米形态响应性肌管方向方面的作用 可以将其扩展为与DAPC有缺陷的许多其他肌肉营养不良的表型生物标志物 成分。这项研究还将揭示调节细胞行为和细胞命运的新型工程方法 通过DAPC的地形控制。