Mondragon-Gonzalez Ricardo, Selvaraj Sridhar, Perlingeiro Rita C R
Lillehei Heart Institute, Department of Medicine, University of Minnesota, Minneapolis, MN, USA.
Stem Cell Institute, University of Minnesota, Minneapolis, MN, USA.
Methods Mol Biol. 2023;2640:129-142. doi: 10.1007/978-1-0716-3036-5_10.
Pluripotent stem cells have a multitude of potential applications in the areas of disease modeling, drug screening, and cell-based therapies for genetic diseases, including muscular dystrophies. The advent of induced pluripotent stem cell technology allows for the facile derivation of disease-specific pluripotent stem cells for any given patient. Targeted in vitro differentiation of pluripotent stem cells into the muscle lineage is a key step to enable all these applications. Transgene-based differentiation using conditional expression of the transcription factor PAX7 leads to the efficient derivation of an expandable and homogeneous population of myogenic progenitors suitable for both in vitro and in vivo applications. Here, we describe an optimized protocol for the derivation and expansion of myogenic progenitors from pluripotent stem cells using conditional expression of PAX7. Importantly, we further describe an optimized procedure for the terminal differentiation of myogenic progenitors into more mature myotubes, which are better suited for in vitro disease modeling and drug screening studies.
多能干细胞在疾病建模、药物筛选以及针对包括肌肉萎缩症在内的遗传性疾病的细胞疗法等领域有众多潜在应用。诱导多能干细胞技术的出现使得为任何特定患者轻松获取疾病特异性多能干细胞成为可能。将多能干细胞定向体外分化为肌肉谱系是实现所有这些应用的关键步骤。利用转录因子PAX7的条件性表达进行基于转基因的分化，可高效获得适合体外和体内应用的可扩增且同质的成肌祖细胞群体。在此，我们描述了一种使用PAX7条件性表达从多能干细胞中获取和成肌祖细胞并进行扩增的优化方案。重要的是，我们进一步描述了将成肌祖细胞终末分化为更成熟肌管的优化程序，这些肌管更适合体外疾病建模和药物筛选研究。
