Department of Spine Surgery, The First People's Hospital of Lianyungang, Lianyungang, Jiangsu 222000, P.R. China.
Mol Med Rep. 2020 Jul;22(1):328-336. doi: 10.3892/mmr.2020.11071. Epub 2020 Apr 16.
MicroRNAs (miRs) play an important role in the development and progression of spinal cord injury (SCI). The role of miR‑138‑5p in SCI was investigated in the present study. The anti‑inflammatory effects of miR‑138‑5p and underlying mechanisms were investigated in an SCI rat model and in vitro model. Reverse transcription‑quantitative PCR (RT‑qPCR) was used to examine the expression of miR‑138‑5p in the SCI in vivo and in vitro models, as well as patients with SCI; it was found that miR‑138‑5p was significantly upregulated in SCI. Bioinformatics and dual‑luciferase reporter assays were performed to predict and confirm the binding sites between miR‑138‑5p and the 3'untranslated region of sirtuin 1 (SIRT1). Then, the expression of SIRT1 was detected via RT‑qPCR and western blotting, indicating downregulation of SIRT1 in SCI. PC12 cells were transfected with miR‑138‑5p inhibitor, inhibitor control or miR‑138‑5p inhibitor + SIRT1 small interfering RNA for 48 h, and then subjected to lipopolysaccharide (100 ng/ml) treatment for 4 h. Then, MTT assay, flow cytometry and ELISA experiments were performed to analyze cell viability, apoptosis, and the levels of tumor necrosis factor‑α, interleukin (IL)‑1β and IL‑6. Findings suggested that downregulation of miR‑138‑5p increased PC12 cell viability, inhibited cell apoptosis and attenuated proinflammatory responses, which may result in amelioration of SCI. However, all these effects were reversed by SIRT1 knockdown. Finally, it was observed that miR‑138‑5p altered the related protein expression of the PTEN/AKT pathway. These results indicated that miR‑138‑5p could regulate inflammatory responses and cell apoptosis in SCI models by modulating the PTEN/AKT signaling pathway via SIRT1, thus playing an important role in the development of SCI. Collectively, the present study demonstrated that miR‑138‑5p may be a novel therapeutic target for the treatment of SCI.
微小 RNA(miRs)在脊髓损伤(SCI)的发展和进展中发挥重要作用。本研究旨在探讨 miR-138-5p 在 SCI 中的作用。在 SCI 大鼠模型和体外模型中研究了 miR-138-5p 的抗炎作用及其潜在机制。逆转录定量聚合酶链反应(RT-qPCR)用于检测体内和体外模型以及 SCI 患者中 miR-138-5p 的表达,结果发现 miR-138-5p 在 SCI 中显著上调。通过生物信息学和双荧光素酶报告基因检测来预测和确认 miR-138-5p 与沉默信息调节因子 1(SIRT1)3'非翻译区之间的结合位点。然后,通过 RT-qPCR 和蛋白质印迹法检测 SIRT1 的表达,结果表明 SIRT1 在 SCI 中下调。PC12 细胞转染 miR-138-5p 抑制剂、抑制剂对照或 miR-138-5p 抑制剂+SIRT1 小干扰 RNA 48 h 后,用脂多糖(100 ng/ml)处理 4 h。然后,通过 MTT 测定、流式细胞术和 ELISA 实验分析细胞活力、细胞凋亡以及肿瘤坏死因子-α(TNF-α)、白细胞介素(IL)-1β和 IL-6 的水平。结果表明,下调 miR-138-5p 可增加 PC12 细胞活力,抑制细胞凋亡并减轻炎症反应,从而改善 SCI。然而,SIRT1 敲低逆转了所有这些作用。最后,观察到 miR-138-5p 改变了 PTEN/AKT 通路的相关蛋白表达。这些结果表明,miR-138-5p 可能通过 SIRT1 调节 PTEN/AKT 信号通路来调节 SCI 模型中的炎症反应和细胞凋亡,从而在 SCI 的发展中发挥重要作用。综上所述,本研究表明 miR-138-5p 可能成为治疗 SCI 的新的治疗靶点。