Stem cell-based tissue engineering for myotendinous junction modeling and repair

项目总结/摘要 肌腱连接处是肌肉和肌腱之间的直接界面， 创伤性 肌肉损伤和肌腱断裂。鉴于现有治疗方法的有效性不一致， 肌腱损伤，我们建议开发一种替代方法，使用人类诱导多能干细胞 细胞（iPSC）。这样的细胞有能力创造祖细胞，可以有助于肌肉和肌腱 再生我们这项研究的具体目标是设计iPSC衍生的肌肉肌腱单位， 它们作为研究肌腱连接形成和修复肌肉损伤的体外模型的用途， 肌腱和肌腱连接处。指导这一提议的基本假设是iPSC衍生的 肌肉骨骼祖细胞（骨骼肌祖细胞和肌腱祖细胞）将与 彼此连接并形成具有功能性肌腱连接的肌肉-肌腱单元。这一假设得到了支持 通过我们发表的研究和初步数据，证明了生产人类肌肉骨骼的可行性， 从iPSC中提取的组织。在这个提议中，我们将从iPSC制备人肌腱祖细胞（hTPC）系。 将建立的细胞与iPSC衍生的骨骼肌祖细胞（hSMPC）共培养，使用 用于iPSC的新特征细胞培养系统，二维微图案化培养平台（Aim 1）。 来自微图案的地形和分子指导可以模拟细胞和分子的复杂性 在肌肉骨骼发育和病理学方面的研究。接下来，我们将创建三维肌肉肌腱组织 使用iPSC衍生的肌肉骨骼祖细胞进行培养，并分析它们的解剖和生理特性。 这些特性扩展了iPSC用于建模肌腱连接形成的效用（目的2）。在整个 这些三维培养模型的发展，我们希望确定外源刺激的作用，如 肌-腱分化和肌-腱连接的信号分子和机械负荷 阵最后，我们将测试iPSC衍生的肌肉肌腱组织再生受伤肌肉的能力， 肌腱和肌腱连接处，通过研究在完全肌腱连接处大鼠模型中的植入 破裂（目标3）。这些目标将为使用基于iPSC的体外细胞培养的有效方法提供非常新颖的见解。 建模和治疗。由于iPSC现在可以来源于人类成人体组织，因此这种方法可以 用于开发患者特异性的、基于细胞的体外模型和人类疾病的治疗。的结果 这一项目将加快有效治疗肌肉骨骼疾病患者的进展。 由于缺乏有效的治疗肌腱损伤和随之而来的负担， 社会，这项研究是紧迫和及时的。

PROJECT SUMMARY/ABSTRACT Myotendinous junctions are the direct interface between muscle and tendon and the affected site in traumatic muscle injury and myotendinous rupture. Given the inconsistent effectiveness of existing treatments for myotendinous injuries, we propose developing an alternative approach using human induced pluripotent stem cells (iPSCs). Such cells have the capacity to create progenitor cells that can contribute to muscle and tendon regeneration. Our specific goals of this study are to engineer iPSC-derived muscle-tendon units and evaluate their utility as an in vitro model to study myotendinous junction formation, and to repair damages in muscle, tendon, and myotendinous junctions. The fundamental hypothesis guiding this proposal is that iPSC-derived musculoskeletal progenitor cells (skeletal muscle progenitor cells and tendon progenitor cells) will interact with each other and form a muscle-tendon unit with functional myotendinous junctions. This hypothesis is supported by our published studies and preliminary data demonstrating the feasibility of producing human musculoskeletal tissues from iPSCs. In this proposal, we will prepare lines of human tendon progenitor cells (hTPCs) from iPSCs. The established cells will be co-cultured with iPSC-derived skeletal muscle progenitor cells (hSMPCs) using newly featured cell culture systems for iPSCs, two-dimensional micropatterned culture platforms (Aim 1). Topographical and molecular guidance from the micropatterns can simulate cellular and molecular complexity in musculoskeletal development and pathology. Next, we will create three-dimensional muscle-tendon tissue cultures using iPSC-derived musculoskeletal progenitor cells and analyze their anatomical and physiological properties to extend the utility of iPSCs for modeling myotendinous junction formation (Aim 2). Throughout the development of these 3D culture models, we hope to identify the roles of exogenous stimulations such as signaling molecules and mechanical loads for muscle-tendon differentiation and myotendinous junction formation. Lastly, we will test the capacity of iPSC-derived muscle-tendon tissues to regenerate injured muscle, tendon, and myotendinous junctions by studying implantation in a rat model of complete myotendinous junction rupture (Aim 3). These aims will provide highly novel insights into effective approaches using iPSC-based in vitro modeling and treatments. As iPSCs can now be derived from human adult somatic tissues, this approach can be used to develop patient-specific, cell-based in vitro models and therapy for human disease. The results of this project will accelerate progress towards effective treatments for patients with musculoskeletal disorders. Given the lack of effective treatments for myotendinous injuries and the consequential burden it places on society, this study is both urgent and timely.