Generation of allogenic and xenogeneic functional muscle stem cells for intramuscular transplantation [scRNA-seq]

Satellite cells, the stem cells of skeletal muscle tissue, hold a remarkable regeneration capacity and therapeutic potential in regenerative medicine. However, low satellite cell yield from autologous or donor-derived muscles hinders the adoption of satellite cell transplantation for the treatment of muscle diseases, including Duchenne muscular dystrophy (DMD). To address this limitation, here we investigated whether satellite cells can be derived in allogeneic or xenogeneic animal hosts. First, injection of CRISPR/Cas9-corrected mouse DMD-induced pluripotent stem cells (iPSCs) into mouse blastocysts carrying an ablation system of host satellite cells gave rise to intraspecies chimeras exclusively carrying iPSC-derived satellite cells. Furthermore, injection of genetically corrected DMD-iPSCs into rat blastocysts resulted in the formation of interspecies rat-mouse chimeras harboring mouse satellite cells. Remarkably, iPSC-derived satellite cells or derivative myoblasts produced in intraspecies or interspecies chimeras restored dystrophin expression in DMD mice following intramuscular transplantation, and contributed to the satellite cell pool. Collectively, this study demonstrates the feasibility of producing therapeutically competent stem cells across divergent animal species, raising the possibility of generating human muscle stem cells in large animals for regenerative medicine purposes.肌卫星细胞（satellite cells）作为骨骼肌组织的成体干细胞，具备优异的再生能力与再生医学领域的治疗应用潜力。然而，从自体或供体来源肌肉中获取的肌卫星细胞产量低下，制约了肌卫星细胞移植疗法在包括杜氏肌营养不良症（Duchenne muscular dystrophy, DMD）在内的肌肉疾病治疗中的应用。为解决这一局限，本研究探究了能否在同种异体或异种动物宿主体内诱导生成肌卫星细胞。首先，将经CRISPR/Cas9基因编辑校正的杜氏肌营养不良症小鼠诱导多能干细胞（iPSCs）注射入携带宿主肌卫星细胞消融系统的小鼠囊胚中，成功获得了仅包含iPSC来源肌卫星细胞的种内嵌合体。此外，将经基因校正的杜氏肌营养不良症小鼠iPSCs注射入大鼠囊胚，成功构建了携带有小鼠肌卫星细胞的跨物种大鼠-小鼠嵌合体。值得注意的是，从种内或跨物种嵌合体中获取的iPSC来源肌卫星细胞及其分化而成的成肌细胞，在肌肉注射移植至杜氏肌营养不良症小鼠体内后，可恢复其抗肌萎缩蛋白的表达，并可整合至宿主的肌卫星细胞库中。综上，本研究证实了跨不同动物物种制备具备治疗效能的干细胞的可行性，为在大型动物体内培育人类肌肉干细胞以用于再生医学研究开辟了可能性。