Department of Orthopaedics, Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi Province, China.
Department of Orthopaedics, First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan Province, China.
Cell Transplant. 2020 Jan-Dec;29:963689720916173. doi: 10.1177/0963689720916173.
MicroRNAs (miRNAs) function as gene expression switches, and participate in diverse pathophysiological processes of spinal cord injury (SCI). Olfactory ensheathing cells (OECs) can alleviate pathological injury and facilitate functional recovery after SCI. However, the mechanisms by which OECs restore function are not well understood. This study aims to determine whether silencing miR-199a-5p would enhance the beneficial effects of the OECs. In this study, we measured miR-199a-5p levels in rat spinal cords with and without injury, with and without OEC transplants. Then, we transfected OECs with the sh-miR-199a-5p lentiviral vector to reduce miR-199a-5p expression and determined the effects of these OECs in SCI rats by Basso-Beattie-Bresnahan (BBB) locomotor scores, diffusion tensor imaging (DTI), and histological methods. We used western blotting to measure protein levels of Slit1, Robo2, and srGAP2. Finally, we used the dual-luciferase reporter assay to assess the relationship between miR-199-5p and Slit1, Robo2, and srGAP2 expression. We found that SCI significantly increased miR-199a-5p levels ( < 0.05), and OEC transplants significantly reduced miR-199a-5p expression ( < 0.05). Knockdown of miR-199a-5p in OECs had a better therapeutic effect on SCI rats, indicated by higher BBB scores and fractional anisotropy values on DTI, as well as histological findings. Reducing miR-199a-5p levels in transplanted OECs markedly increased spinal cord protein levels of Slit1, Robo2, and srGAP2. Our results demonstrated that transplantation of sh-miR-199a-5p-modified OECs promoted functional recovery in SCI rats, suggesting that miR-199a-5p knockdown was more beneficial to the therapeutic effects of OEC transplants. These findings provided new insights into miRNAs-mediated therapeutic mechanisms of OECs, which helps us to develop therapeutic strategies based on miRNAs and optimize cell therapy for SCI.
微小 RNA（miRNA）作为基因表达开关，参与脊髓损伤（SCI）的多种病理生理过程。嗅鞘细胞（OEC）可以减轻 SCI 后的病理损伤并促进功能恢复。然而，OEC 恢复功能的机制尚不清楚。本研究旨在确定沉默 miR-199a-5p 是否会增强 OEC 的有益作用。在这项研究中，我们测量了有和没有损伤的大鼠脊髓中的 miR-199a-5p 水平，以及有和没有 OEC 移植的大鼠。然后，我们用 sh-miR-199a-5p 慢病毒载体转染 OEC，以降低 miR-199a-5p 的表达，并通过 Basso-Beattie-Bresnahan（BBB）运动评分、扩散张量成像（DTI）和组织学方法来确定这些 OEC 在 SCI 大鼠中的作用。我们使用 Western blot 测量 Slit1、Robo2 和 srGAP2 的蛋白水平。最后，我们使用双荧光素酶报告基因检测评估 miR-199-5p 与 Slit1、Robo2 和 srGAP2 表达之间的关系。我们发现 SCI 显著增加了 miR-199a-5p 的水平（<0.05），而 OEC 移植显著降低了 miR-199a-5p 的表达（<0.05）。OEC 中 miR-199a-5p 的敲低对 SCI 大鼠有更好的治疗作用，BBB 评分更高，DTI 的各向异性分数值更高，组织学发现也更好。降低移植 OEC 中的 miR-199a-5p 水平显著增加了脊髓 Slit1、Robo2 和 srGAP2 的蛋白水平。我们的结果表明，sh-miR-199a-5p 修饰的 OEC 移植促进了 SCI 大鼠的功能恢复，这表明 miR-199a-5p 的敲低对 OEC 移植的治疗效果更有益。这些发现为 miRNA 介导的 OEC 治疗机制提供了新的见解，有助于我们基于 miRNA 开发治疗策略并优化 SCI 的细胞治疗。
