Muscle-derived stem/progenitor cells (MDSPCs) have been demonstrated as a promising source of cellular therapy for the regeneration of peripheral nerves and how to improve the therapeutic potential of these cells is the focus of future research. This study aims to investigate the role of hypoxia on functional effects in MDSPCs. In this study, MDSPCs were isolated from skeletal muscles of limbs; meanwhile, the flow cytometry analysis, immunocytochemistry and multilineage differentiation were performed to characterize MDSPCs. We set cultured MDSPCs in a hypoxic condition with 3% oxygen. The quantitative RT–PCR results showed that hypoxia inhibits myogenic differentiation genes expression, while it is beneficial for the stemness and pro-angiogenic genes. The CCK-8, flow cytometry, Hoechst 33342 and trypan blue dye assays revealed that hypoxia promotes survival and reduces apoptotic cells from oxidative stress damage. The western blot analysis indicated increased expression of HIF-1α and its downstream proteins by hypoxia. Moreover, the angiogenesis assay showed that condition media extracted from hypoxia-treated MDSPCs support endothelial cells’ survival, migration and tube formation in vitro. The cultured cells under hypoxia appear as a salutary strategy to improve the survival and stemness of MDSPCs and enhance angiogenesis capacity in tissue injury repair and reconstruction applications.

Abbreviations: MDSPCs, Muscle-derived stem/progenitor cells; ESCs, embryonic stem cells; PPSCs, induced pluripotent stem cells; ACSs, adult stem cells; PNI, Peripheral nerve injuries; BMSCs, bone marrow mesenchymal stem cells; NSCs, neural stem cells; ADSCs, adipose-derived stem cells; HIF-1α; Hypoxia-inducible factor-1α ; VEGF, vascular endothelial growth factor; SDF-1, stromal-derived factor 1; PM, proliferation media; DMEM, Dulbecco’s modified eagle medium; FBS, fetal bovine serum; MYHC, Myosin heavy chain; CCK-8, cell counting kit-8; H₂O₂, hydrogen peroxide; CM, conditioned media; CON, control group; qPCR, Quantitative real-time polymerase chain reaction; MRL/MpJ, Murphy Roths Large

肌源性干/祖细胞 (MDPCs) 已被证明是用于周围神经再生的细胞疗法的有希望的来源，如何提高这些细胞的治疗潜力是未来研究的重点。本研究旨在探讨缺氧对 MDPCs 功能影响的作用。本研究从四肢骨骼肌中分离出MDPCs；同时，进行流式细胞术分析、免疫细胞化学和多向分化以表征 MDPCs。我们将培养的 MDPCs 置于含 3% 氧气的缺氧条件下。定量RT-PCR结果显示缺氧抑制生肌分化基因的表达，而有利于干性和促血管生成基因。CCK-8，流式细胞术，Hoechst 33342 和台盼蓝染料测定显示缺氧促进存活并减少氧化应激损伤的凋亡细胞。蛋白质印迹分析表明缺氧导致 HIF-1α 及其下游蛋白的表达增加。此外，血管生成分析表明，从低氧处理的 MDPCs 中提取的条件培养基支持内皮细胞的存活、迁移和管形成体外。在缺氧条件下培养的细胞似乎是一种有益的策略，可提高 MDPCs 的存活率和干性，并在组织损伤修复和重建应用中增强血管生成能力。

缩写：MDPCs，肌肉来源的干/祖细胞；胚胎干细胞，胚胎干细胞；PPSCs，诱导多能干细胞；ACSs，成体干细胞；PNI，周围神经损伤；BMSCs，骨髓间充质干细胞；NSCs，神经干细胞；ADSCs，脂肪来源的干细胞；HIF-1α; 缺氧诱导因子-1α ; VEGF，血管内皮生长因子；SDF-1，基质衍生因子1；PM，扩散媒体；DMEM，Dulbecco 改良的鹰培养基；FBS，胎牛血清；MYHC，肌球蛋白重链；CCK-8，细胞计数试剂盒-8；H ₂ O ₂，过氧化氢；CM，条件培养基；CON，对照组；qPCR，定量实时聚合酶链反应；MRL/MpJ，墨菲罗斯大