Maintaining stem cells in physiologically relevant states is necessary to understand cell and context-specific signalling paradigms and to understand complex interfaces between cells in situ. Understanding human stem cell function is largely based on tissue biopsies, cell culture, and transplantation into model organisms. Here, we describe a method to isolate post-mortem intact human muscle myofibers and culture muscle stem cells within the niche microenvironment to assay cellular dynamics, stem cell identity, stem cell hierarchy, and differentiation potential. We show human myofiber culture maintains complex cell-cell contacts and extracellular niche composition during culture. Human satellite cells can be cultured at least 8 days, which represents a timepoint of activation, differentiation, and de novo human myofiber formation. We demonstrate that adult human muscle stem cells undergo apicobasal and planar cell divisions and express polarized dystrophin and EGFR. Furthermore, we validate that stimulation of the EGFR pathway stimulates the generation of myogenic progenitors and myogenic differentiation. This method provides proof of principle evidence for the use of human muscle to evaluate satellite cell dynamics and has applications in pre-clinical evaluation of therapeutics targeting muscle repair.

中文翻译：

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培养成人骨骼肌肌纤维的人类卫星细胞动力学分析


将干细胞维持在生理相关状态对于了解细胞和特定环境的信号传导范式以及了解原位细胞之间的复杂界面是必要的。了解人类干细胞功能主要基于组织活检、细胞培养和移植到模型生物体中。在这里，我们描述了一种分离死后完整的人类肌肉纤维并在利基微环境中培养肌肉干细胞的方法，以测定细胞动力学、干细胞身份、干细胞层次结构和分化潜力。我们证明人类肌纤维培养在培养过程中保持复杂的细胞间接触和细胞外生态位组成。人类卫星细胞可以培养至少8天，这代表了激活、分化和人类肌纤维从头形成的时间点。我们证明成人肌肉干细胞经历顶端和平面细胞分裂并表达极化肌营养不良蛋白和 EGFR。此外，我们验证了 EGFR 通路的刺激会刺激肌源性祖细胞的产生和肌源性分化。该方法为使用人体肌肉评估卫星细胞动力学提供了原理证据，并可应用于针对肌肉修复的治疗的临床前评估。