Institute of Biotechnology, University of Lausanne, Lausanne, Switzerland.
Department of Pathology and Immunology, Faculty of Medicine, University of Geneva, 1211 Geneva, Switzerland.
Mol Ther. 2018 Apr 4;26(4):1093-1108. doi: 10.1016/j.ymthe.2018.01.021. Epub 2018 Feb 2.
Duchenne muscular dystrophy (DMD) is a lethal muscle-wasting disease currently without cure. We investigated the use of the PiggyBac transposon for full-length dystrophin expression in murine mesoangioblast (MABs) progenitor cells. DMD murine MABs were transfected with transposable expression vectors for full-length dystrophin and transplanted intramuscularly or intra-arterially into mdx/SCID mice. Intra-arterial delivery indicated that the MABs could migrate to regenerating muscles to mediate dystrophin expression. Intramuscular transplantation yielded dystrophin expression in 11%-44% of myofibers in murine muscles, which remained stable for the assessed period of 5 months. The satellite cells isolated from transplanted muscles comprised a fraction of MAB-derived cells, indicating that the transfected MABs may colonize the satellite stem cell niche. Transposon integration site mapping by whole-genome sequencing indicated that 70% of the integrations were intergenic, while none was observed in an exon. Muscle resistance assessment by atomic force microscopy indicated that 80% of fibers showed elasticity properties restored to those of wild-type muscles. As measured in vivo, transplanted muscles became more resistant to fatigue. This study thus provides a proof-of-principle that PiggyBac transposon vectors may mediate full-length dystrophin expression as well as functional amelioration of the dystrophic muscles within a potential autologous cell-based therapeutic approach of DMD.
杜氏肌营养不良症（DMD）是一种致命的肌肉消耗性疾病，目前尚无治愈方法。我们研究了利用 PiggyBac 转座子在鼠中胚层血管前体细胞（MAB）祖细胞中全长表达抗肌萎缩蛋白。用全长抗肌萎缩蛋白的转座表达载体转染 DMD 鼠 MAB，并通过肌内或动脉内移植到 mdx/SCID 小鼠中。动脉内递送表明 MAB 可以迁移到再生肌肉中，介导抗肌萎缩蛋白的表达。肌内移植使 11%-44%的肌纤维在鼠肌肉中表达抗肌萎缩蛋白，在评估的 5 个月期间保持稳定。从移植肌肉中分离的卫星细胞构成了 MAB 衍生细胞的一部分，表明转染的 MAB 可能定殖于卫星干细胞龛。通过全基因组测序进行转座子整合位点映射表明，70%的整合发生在基因间，而在基因内则没有观察到。原子力显微镜进行的肌肉阻力评估表明，80%的纤维表现出弹性特性恢复到野生型肌肉的水平。体内测量表明，移植的肌肉对疲劳的抵抗力增强。因此，这项研究提供了一个原理证明，即 PiggyBac 转座子载体可以介导全长抗肌萎缩蛋白的表达，并改善 DMD 潜在的自体细胞治疗方法中肌肉的功能。
