Li Jie, Li Liqiang, Shen Yong
Department of Spinal Surgery, The Third Hospital of Hebei Medical University, Shijiazhuang, Hebei 050051, P.R. China.
Exp Ther Med. 2018 Apr;15(4):3563-3569. doi: 10.3892/etm.2018.5829. Epub 2018 Feb 1.
The present study aimed to investigate the role of microRNA (miR)-219-5p in spinal cord injury (SCI) and to examine the underlying molecular mechanism. SCI rat and cell models were conducted in the current study, while reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to detect the level of miR-219-5p in the SCI mice and neurons. Bioinformatics analysis was applied to predict the target genes of miR-219-5p, and dual-luciferase reporter assay was performed to verify the prediction. In addition, MTT assay and flow cytometry were conducted to determine the cell viability and cell apoptosis of the neurons, respectively. Western blot analysis was also performed to detect the expression of associated proteins. The study results demonstrated that miR-219-5p was highly expressed in SCI mice and neurons, and directly targets liver receptor homolog-1 (LRH-1). The neuron viability was significantly reduced by SCI, however, it was recovered upon transfection with an miR-219-5p inhibitor. Neuron apoptosis was notably induced by SCI and inhibited by miR-219-5p inhibition. The LRH-1/Wnt/β-catenin signaling pathway was also inhibited by SCI, while it was significantly enhanced by the miR-219-5p inhibitor. Furthermore, LRH-1 overexpression eliminated the effects of the miR-219-5p inhibitor on SCI. In conclusion, these data indicated that the miR-219-5p inhibitor served a protective role in SCI via regulating the LRH-1/Wnt/β-catenin signaling pathway.
本研究旨在探讨微小RNA(miR)-219-5p在脊髓损伤(SCI)中的作用,并研究其潜在的分子机制。本研究建立了SCI大鼠和细胞模型,同时采用逆转录定量聚合酶链反应(RT-qPCR)检测SCI小鼠和神经元中miR-219-5p的水平。应用生物信息学分析预测miR-219-5p的靶基因,并进行双荧光素酶报告基因检测以验证该预测。此外,进行MTT法和流式细胞术分别测定神经元的细胞活力和细胞凋亡情况。还进行了蛋白质印迹分析以检测相关蛋白的表达。研究结果表明,miR-219-5p在SCI小鼠和神经元中高表达,且直接靶向肝脏受体同源物1(LRH-1)。SCI显著降低了神经元活力,但用miR-219-5p抑制剂转染后可恢复。SCI显著诱导神经元凋亡,而miR-219-5p抑制可抑制凋亡。SCI还抑制了LRH-1/ Wnt/β-连环蛋白信号通路,而miR-219-5p抑制剂可显著增强该通路。此外,LRH-1过表达消除了miR-219-5p抑制剂对SCI的影响。总之,这些数据表明,miR-219-5p抑制剂通过调节LRH-1/ Wnt/β-连环蛋白信号通路在SCI中发挥保护作用。