Human muscle-derived CLEC14A-positive cells regenerate muscle independent of PAX7

Skeletal muscle stem cells, called satellite cells, are responsible for postnatal muscle growth, homeostasis and regeneration. Attempts to utilize the regenerative potential of muscle stem cells for therapeutic purposes so far failed. The transcription factor Pax7 defines satellite cells across species 1-4 . We previously established human PAX7-positive cell colonies with high regenerative potential 5 . We now identified PAX7-negative human muscle-derived cell colonies (PAX7neg) also positive for the myogenic markers desmin and MYF5. These included cells from a unique myopathic patient with rigid spine and respiratory insufficiency due to a homozygous PAX7 c.86-1G>A mutation (PAX7null). Single cell and bulk transcriptome analysis showed high intra- and inter-donor heterogeneity and revealed the endothelial cell marker CLEC14A to be highly expressed in PAX7null cells. All PAX7neg cell populations, including PAX7null, formed myofibers after transplantation into mice, and regenerated muscle after reinjury. Transplanted PAX7neg cells repopulated the satellite cell niche where they re-expressed PAX7. Strikingly, PAX7null cells expressing CLEC14A were also identified below the basal lamina. In summary, transplanted human cells do not depend on PAX7 for muscle regeneration. Thus, Pax7 is not a suitable marker for selection of optimal cells for muscle regenerative therapies.

骨骼肌干细胞（satellite cells，卫星细胞）负责出生后肌肉的生长、稳态维持与再生修复。迄今为止，利用肌肉干细胞再生潜能开展治疗性研究的尝试均以失败告终。转录因子Pax7是跨物种卫星细胞的特异性标记物（参考文献1-4）。本团队此前已成功构建具备高再生潜能的人源PAX7阳性细胞集落（参考文献5）。本研究此次鉴定出一类人源肌肉源性PAX7阴性细胞集落（PAX7neg），该类细胞同时表达肌源性标志物结蛋白（desmin）与MYF5。该类细胞的样本取自1名特殊肌病患者：该患者因PAX7基因纯合突变c.86-1G>A（PAX7null）引发刚性脊柱综合征与呼吸功能不全。单细胞与批量转录组（bulk transcriptome）分析显示，样本供体间及供体内均存在高度异质性，同时发现内皮细胞标志物CLEC14A在PAX7null细胞中呈高表达。所有PAX7neg细胞群（含PAX7null细胞）在移植至小鼠体内后均可形成肌纤维，并在再次损伤后完成肌肉再生修复。移植后的PAX7neg细胞可定植于卫星细胞龛（satellite cell niche），并重新表达Pax7。值得注意的是，表达CLEC14A的PAX7null细胞同样可被检测于基膜（basal lamina）下方。综上，移植的人源细胞进行肌肉再生并不依赖Pax7。因此，Pax7并不适合作为筛选肌肉再生治疗最优细胞的标记物。