Sprott Center for Stem Cell Research, Regenerative Medicine Program, Ottawa Hospital Research Institute, 501 Smyth Road, Ottawa, ON, K1H 8L6, Canada.
Department of Cellular and Molecular Medicine, Faculty of Medicine, University of Ottawa, Ottawa, ON, Canada.
Skelet Muscle. 2021 Jan 4;11(1):1. doi: 10.1186/s13395-020-00256-z.
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.
为了理解细胞和特定环境信号传递模式，并了解细胞之间在原位的复杂界面，有必要使干细胞维持在生理相关状态。对人类干细胞功能的理解主要基于组织活检、细胞培养和移植到模式生物中。
在这里，我们描述了一种从死后完整的人类肌肉肌纤维中分离肌肉干细胞并在微环境中培养的方法，以检测细胞动力学、干细胞特性、干细胞层次结构和分化潜能。
我们表明，人类肌纤维培养在培养过程中保持复杂的细胞-细胞接触和细胞外微环境组成。人类卫星细胞至少可以培养 8 天，这代表了激活、分化和新的人类肌纤维形成的时间点。我们证明成年人类肌肉干细胞经历顶端-基底和平面细胞分裂，并表达极化的营养不良蛋白和 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。
该方法为使用人类肌肉评估卫星细胞动力学提供了原理验证，并可应用于针对肌肉修复的治疗方法的临床前评估。
