Bone marrow mesenchymal stem cell (BMSC)-derived small extracellular vesicles (sEV) but not fibroblast sEV provide retinal ganglion cell (RGC) neuroprotection both in vitro and in vivo, with miRNAs playing an essential role. More than 40 miRNAs were more abundant in BMSC-sEV than in fibroblast-sEV. The purpose of this study was to test the in vitro and in vivo neuroprotective and axogenic properties of six candidate miRNAs (miR-26a, miR-17, miR-30c-2, miR-92a, miR-292, and miR-182) that were more abundant in BMSC-sEV than in fibroblast-sEV. Adeno-associated virus 2 (AAV2) expressing a combination of three of the above candidate miRNAs were added to heterogenous adult rat retinal cultures or intravitreally injected into rat eyes one week before optic nerve crush (ONC) injury. Survival and neuritogenesis of βIII-tubulin⁺ RGCs was assessed in vitro, as well as the survival of RBPMS⁺ RGCs and regeneration of their axons in vivo. Retinal nerve fiber layer thickness (RNFL) was measured to assess axonal density whereas positive scotopic threshold response electroretinography amplitudes provided a readout of RGC function. Qualitative retinal expression of PTEN, a target of several of the above miRNAs, was used to confirm successful miRNA activity. AAV2 reliably transduced RGCs in vitro and in vivo. Viral delivery of miRNAs in vitro showed a trend towards neuroprotection but remained insignificant. Delivery of selected combinations of miRNAs (miR-17-5p, miR-30c-2 and miR-92a; miR-92a, miR-292 and miR-182) before ONC provided significant therapeutic benefits according to the above measurable endpoints. However, no single miRNA appeared to be responsible for the effects observed, whilst positive effects observed appeared to coincide with successful qualitative reduction in PTEN immunofluorescence in the retina. Viral delivery of miRNAs provides a possible neuroprotective strategy for injured RGCs that is conducive to therapeutic manipulation.

骨髓间充质干细胞 (BMSC) 衍生的小细胞外囊泡 (sEV) 但不是成纤维细胞 sEV，可在体外和体内提供视网膜神经节细胞 (RGC) 神经保护，其中 miRNA 发挥重要作用。与成纤维细胞-sEV 相比，BMSC-sEV 中的 40 多种 miRNA 更丰富。本研究的目的是在体外和体内测试六种候选 miRNA（miR-26a、miR-17、miR-30c-2、miR-92a、miR-292 和 miR-182）的神经保护和轴生特性在 BMSC-sEV 中比在成纤维细胞-sEV 中更丰富。在视神经挤压 (ONC) 损伤前一周，将表达上述三种候选 miRNA 组合的腺相关病毒 2 (AAV2) 添加到异质成年大鼠视网膜培养物中或玻璃体内注射到大鼠眼中。存活和βIII微管蛋白的轴突⁺视网膜神经节细胞中评估在体外，以及RBPMS的生存⁺视网膜神经节细胞和再生轴突的体内. 测量视网膜神经纤维层厚度 (RNFL) 以评估轴突密度，而正暗视阈值响应视网膜电图振幅提供了 RGC 功能的读数。PTEN 的定性视网膜表达是上述几种 miRNA 的靶标，用于确认成功的 miRNA 活性。AAV2在体外和体内可靠地转导 RGC 。体外miRNA 的病毒传递显示出神经保护的趋势，但仍然不显着。根据上述可测量终点，在 ONC 之前递送选定的 miRNA 组合（miR-17-5p、miR-30c-2 和 miR-92a；miR-92a、miR-292 和 miR-182）可提供显着的治疗益处。然而，似乎没有单个 miRNA 对观察到的影响负责，而观察到的积极影响似乎与视网膜中 PTEN 免疫荧光的成功定性降低相吻合。miRNA 的病毒传递为受伤的 RGC 提供了一种可能的神经保护策略，有利于治疗操作。