Inhibition of miR-27a suppresses the inflammatory response via the p38/MAPK pathway in intervertebral disc cells
Inhibition of miR-27a suppresses the inflammatory response via the p38/MAPK pathway in intervertebral disc cells
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DOI：
10.3892/etm.2017.5053
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发表时间：
2017-11-01
影响因子：
2.7
通讯作者：
Chen, Liang
中科院分区：
文献类型：
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作者：
Cao, Zhenguo;Chen, Liang
本研究旨在探讨miR-27 a在椎间盘退变（IDD）中的作用并研究其潜在机制。采用定量聚合酶链反应（qPCR）检测IDD患者髓核（NP）组织中miR-27 a的表达水平，结果显示IDD患者髓核组织中miR-27 a的表达水平高于对照组。为了进一步研究miR-27 a在IDD中的作用，通过用慢病毒antigomiR-27 a抑制剂转染细胞来产生具有低miR-27 a表达的稳定的人NP细胞系。此外，通过脂多糖（LPS; 10 μ M）刺激建立人NP细胞炎症模型。通过qPCR测定NP细胞中的miR-27 a表达，而通过蛋白质印迹分析测量其靶蛋白; p-p38和核因子（NF-κ B）的表达。此外，还分别通过qPCR和ELISA评价了促炎因子（包括白细胞介素（IL）-1 β、IL-6和肿瘤坏死因子-α（TNF-α））的mRNA和蛋白表达水平。目前的结果证实，miR-27 a在IDD中显著上调。在体外，通过转染miR-27 a抑制剂下调LPS刺激的NP细胞中的miR-27 a导致p-p38和NF-κ B表达水平的抑制。此外，在具有下调的miR-27 a的LPS刺激的NP细胞中，促炎因子IL-1 β、IL-6和TNF-α的产生显著减少。结论：miR-27 a可能在IDD的发生发展中起促进作用，而抑制miR-27 a可能通过调节p38/丝裂原活化蛋白激酶（MAPK）信号通路抑制椎间盘细胞释放的促炎因子。
The current study aimed to investigate the role of miR-27a in intervertebral disc degeneration (IDD) and to examine the underlying mechanisms. Quantitative polymerase chain reaction (qPCR) was performed to detect the expression level of miR-27a in the nucleus pulposus (NP) tissues of patients with IDD, and the results revealed an increasing expression of miR-27a in IDD compared with the control. To further investigate the role of miR-27a in IDD, a stable human NP cell line with low miR-27a expression was generated by transfecting cells with a lentiviral antigomiR-27a inhibitor. In addition, a human NP cell inflammation model was established by lipopolysaccharide (LPS; 10 mu M) stimulation. The miR-27a expression in NP cells was determined by qPCR, while the expression of its target proteins; p-p38 and nuclear factor (NF-kappa B) was measured by western blot analysis. Furthermore, the mRNA and protein expression levels of proinflammatory factors, including interleukin (IL)-1 beta, IL-6 and tumor necrosis factor-alpha (TNF-alpha), were also evaluated by qPCR and ELISA, respectively. The current results confirmed that miR-27a was significantly upregulated in IDD. In vitro, downregulation of miR-27a in LPS-stimulated NP cells by transfection with the miR-27a inhibitor resulted in suppression of p-p38 and NF-kappa B expression levels. Furthermore, the production of the proinflammatory factors IL-1 beta, IL-6 and TNF-alpha was significantly reduced in LPS-stimulated NP cells with down-regulated miR-27a. In conclusion, miR-27a may function as a promoter in IDD development, while inhibition of miR-27a may suppress proinflammatory factors released by intervertebral disc cells by regulating the p38/mitogen-activated protein kinase (MAPK) signaling pathway.