Department of Otorhinolaryngology and Head and Neck Surgery, Shanghai General Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai, China.
Dis Markers. 2022 Sep 13;2022:7071877. doi: 10.1155/2022/7071877. eCollection 2022.
Peripheral nerve injury (PNI) is often resulting from trauma, which leads to severe and permanently disability. Schwann cells are critical for facilitating the regeneration process after PNI. Adipose-derived mesenchymal stem cells (ADSCs) exosomes have been used as a novel treatment for peripheral nerve injury. However, the underlying mechanism remains unclear. In this study, we isolated ADSCs and extracted exosomes, which were verified by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and western blot (WB). Cocultured with Dorsal Root Ganglion (DRG) and Schwann cells (SCs) to evaluate the effect of exosomes on the growth of DRG axons by immunofluorescence, and the proliferation and migration of SCs by CCK8 and Transwell assays, respectively. Through exosomal miRNA sequencing and bioinformatic analysis, the related miRNAs and target gene were predicted and identified by dual luciferase assay. Related miRNAs were overexpressed and inhibited, respectively, to clarify their effects; the downstream pathway through the target gene was determined by real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) and WB. Results found that ADSC-exosomes could promote the proliferation and migration of SCs and the growth of DRG axons, respectively. Exosomal miRNA-22-3p from ADSCs directly inhibited the expression of Phosphatase and Tensin Homolog deleted on Chromosome 10 (PTEN), activated phosphorylation of the AKT/mTOR axis, and enhanced SCs proliferation and migration. In conclusion, our findings suggest that ADSC-exosomes could promote SCs function through exosomal miRNA-22-3p, which could be used as a therapeutic target for peripheral nerve injury.
周围神经损伤（PNI）通常是由创伤引起的，会导致严重且永久性的残疾。许旺细胞（Schwann cells）对于 PNI 后的再生过程至关重要。脂肪间充质干细胞（adipose-derived mesenchymal stem cells，ADSCs）衍生的外泌体已被用于治疗周围神经损伤。然而，其潜在机制尚不清楚。在本研究中，我们分离了 ADSCs 并提取了外泌体，通过透射电子显微镜（transmission electron microscopy，TEM）、纳米颗粒跟踪分析（nanoparticle tracking analysis，NTA）和 Western blot 进行了验证。我们将外泌体与背根神经节（Dorsal Root Ganglion，DRG）和许旺细胞（Schwann cells，SCs）共培养，通过免疫荧光法评估外泌体对 DRG 轴突生长的影响，通过 CCK8 和 Transwell 实验分别评估对 SCs 增殖和迁移的影响。通过外泌体 miRNA 测序和生物信息学分析，预测和鉴定了相关的 miRNA 和靶基因，并通过双荧光素酶报告基因实验进行验证。过表达和抑制相关 miRNA，分别阐明其作用；通过实时荧光定量聚合酶链反应（real-time fluorescence quantitative polymerase chain reaction，RT-qPCR）和 Western blot 确定通过靶基因的下游途径。结果发现，ADSC 衍生的外泌体可以分别促进 SCs 的增殖和迁移以及 DRG 轴突的生长。ADSCs 来源的外泌体 miRNA-22-3p 可直接抑制第 10 号染色体缺失的磷酸酶和张力蛋白同源物（Phosphatase and Tensin Homolog deleted on Chromosome 10，PTEN）的表达，激活 AKT/mTOR 轴的磷酸化，增强 SCs 的增殖和迁移。综上所述，我们的研究结果表明，ADSC 衍生的外泌体可以通过外泌体 miRNA-22-3p 促进 SCs 的功能，这可能为周围神经损伤的治疗提供新的靶点。
