Lillehei Heart Institute, Department of Medicine, University of Minnesota, 4-128 CCRB, 2231 6th Street SE, Minneapolis, MN 55455, USA; Department of Genetic, Cell Biology, and Development, University of Minnesota, Minneapolis, MN, USA.
Lillehei Heart Institute, Department of Medicine, University of Minnesota, 4-128 CCRB, 2231 6th Street SE, Minneapolis, MN 55455, USA.
Stem Cell Reports. 2021 Jan 12;16(1):10-19. doi: 10.1016/j.stemcr.2020.11.001. Epub 2020 Dec 3.
Inducible expression of PAX7 in differentiating pluripotent stem cells (PSCs) allows massively scalable generation of human myogenic progenitors, which upon transplantation into dystrophic muscles give rise to donor-derived myofibers and satellite cells. Therefore, PSC-derived PAX7 myogenic progenitors represent an attractive therapeutic approach to promote muscle regeneration. Work to date has used lentiviral vectors (LVs) that randomly integrate inducible PAX7 transgenes. Here, we investigated whether equivalent induction of the myogenic program could be achieved by targeting the PAX7 transgene into genomic safe harbor (GSH) sites. Across multiple PSC lines, we find that this approach consistently generates expandable myogenic progenitors in vitro, although scalability of expansion is moderately reduced compared with the LV approach. Importantly, transplantation of GSH-targeted myogenic progenitors produces robust engraftment, comparable with LV counterparts. These findings provide proof of concept for the use of GSH targeting as a potential alternative approach to generate therapeutic PSC-derived myogenic progenitors for clinical applications.
诱导多能干细胞 (PSCs) 中 PAX7 的表达可大规模生成人类肌源性祖细胞，这些祖细胞在移植到萎缩肌肉后，可产生供体来源的肌纤维和卫星细胞。因此，源自 PSC 的 PAX7 肌源性祖细胞代表了一种有吸引力的治疗方法，可促进肌肉再生。迄今为止的工作使用了随机整合诱导型 PAX7 转基因的慢病毒载体 (LVs)。在这里，我们研究了将 PAX7 转基因靶向基因组安全港 (GSH) 位点是否可以实现等效的肌生成程序诱导。在多个 PSC 系中，我们发现尽管与 LV 方法相比，扩展的可扩展性适度降低，但这种方法始终可在体外生成可扩展的肌源性祖细胞。重要的是，GSH 靶向肌源性祖细胞的移植可产生强大的植入，与 LV 相当。这些发现为使用 GSH 靶向作为生成治疗性 PSC 衍生肌源性祖细胞的潜在替代方法用于临床应用提供了概念验证。
