Department of Clinical Application, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan.
Methods Mol Biol. 2023;2640:143-157. doi: 10.1007/978-1-0716-3036-5_11.
Most muscular dystrophies are the result of genetic disorders. There is currently no effective treatment for these progressive diseases except palliative therapy. Muscle stem cells with potent self-renewal and regenerative potential are considered a target for treating muscular dystrophy. Human induced pluripotent stem cells have been expected as a source of MuSCs because of their infinite proliferation potential and less immunogenicity. However, the generation of engraftable MuSCs from hiPSCs is relatively difficult and encounters low efficiency and reproducibility. Here, we introduce a transgene-free protocol of hiPSCs differentiating into fetal MuSCs by identifying them as MYF5-positive cells. Flow cytometry analysis detected around 10% of MYF5-positive cells after 12 weeks of differentiation. Approximately 50 ~ 60% of MYF5-positive cells were positively identified using Pax7 immunostaining. This differentiation protocol is expected to be useful for not only the establishment of cell therapy but also the future drug discovery using patient-derived hiPSCs.
大多数肌肉萎缩症是遗传疾病的结果。除姑息疗法外,目前尚无有效治疗这些进行性疾病的方法。具有强大自我更新和再生能力的肌肉干细胞被认为是治疗肌肉萎缩症的一个目标。由于具有无限增殖潜力和较低的免疫原性,人类诱导多能干细胞被认为是 MuSCs 的来源。然而,从 hiPSCs 中产生可植入的 MuSCs 相对困难,效率和重现性较低。在这里,我们通过鉴定其为 MYF5 阳性细胞,介绍了一种无转基因的 hiPSCs 分化为胎儿 MuSCs 的方案。流式细胞术分析显示,分化 12 周后约有 10%的 MYF5 阳性细胞。使用 Pax7 免疫染色,约 50%~60%的 MYF5 阳性细胞被阳性鉴定。该分化方案不仅有望用于细胞治疗的建立,也有望用于未来使用患者来源的 hiPSCs 进行药物发现。