miR-381-3p 的过表达促进脊髓损伤的恢复。
Overexpression of miR-381-3p promotes the recovery of spinal cord injury.
机构信息
Department of Orthopedics, The Second Affiliated Hospital of Nanchang University, Nanchang, China.
出版信息
Eur Rev Med Pharmacol Sci. 2018 Sep;22(17):5429-5437. doi: 10.26355/eurrev_201809_15802.
OBJECTIVE
To study the effects of miR-381-3p on spinal cord injury and its underlying mechanism.
MATERIALS AND METHODS
After the spinal cord injury rat model of was established, Sprague Dawley (SD) rats were randomly divided into the control group and the acute spinal cord injury (ASCI) group. Microglial BV2 cells were used as experimental cells, and the cells were divided into the control group and the lipopolysaccharide (LPS) group. The mRNA and protein expression level of miR-381-3p, IKKβ, inflammatory factors, and p-p65 were detected by quantitative Reverse Transcriptase-Polymerase Chain Reaction (qRT-PCR) and Western blot, respectively. Dual-luciferase reporter gene assay and Western blot were used to detect the regulatory effect of IKKβ on miR-381-3p. Changes in grip ability and rotary performance of rats in the ASCI group were evaluated after miR-381-3p overexpression in vivo.
RESULTS
The expression of miR-381-3p was downregulated in rats of the ASCI group, while the expression of IKKβ and p-p65 were upregulated. In vitro experiments demonstrated that LPS could inhibit the expression of miR-381-3p and promote the upregulation of IKKβ and p-p65. Overexpression of miR-381-3p could inhibit the mRNA and protein expression of IKKβ. The upregulated expression of IKKβ, p-p65, tumor necrosis factor-alpha (TNF-α), and interleukins-1β (IL-1β) induced by LPS in BV2 cells were reversed by miR-381-3p mimic transfection. Besides, upregulated TNF-α and IL-1β induced by miR-381-3p inhibitor in BV2 cells were reversed by IKKβ inhibitor (BMS-345541). Results of animal experiments indicated that miR-381-3p was overexpressed in rats of the ASCI group. The protein levels of IKKβ and p-p65, and the mRNA expression levels of inflammatory cytokines TNF-α and IL-1β were remarkably decreased in the ASCI group than those of the control group. The grip ability, coordination, and anti-fatigue performance of rats in the ASCI group recovered quicker than those of the control group.
CONCLUSIONS
MiR-381-3p was downregulated in ASCI rats. The overexpression of miR-381-3p could recover the motor ability of rats in the ASCI group earlier and might inhibit injury aggravation by inhibiting inflammatory responses via the IKKβ-NF-κB pathway.
目的
研究 miR-381-3p 对脊髓损伤的影响及其作用机制。
材料与方法
建立大鼠急性脊髓损伤模型后，将 Sprague Dawley（SD）大鼠随机分为对照组和急性脊髓损伤（ASCI）组。将小胶质细胞 BV2 细胞作为实验细胞，分为对照组和脂多糖（LPS）组。通过定量逆转录-聚合酶链反应（qRT-PCR）和 Western blot 分别检测 miR-381-3p、IKKβ、炎症因子和 p-p65 的 mRNA 和蛋白表达水平。双荧光素酶报告基因检测和 Western blot 用于检测 IKKβ 对 miR-381-3p 的调控作用。体内过表达 miR-381-3p 后，评估 ASCI 组大鼠的抓握能力和旋转性能变化。
结果
ASCI 组大鼠 miR-381-3p 表达下调，而 IKKβ 和 p-p65 表达上调。体外实验表明，LPS 可抑制 miR-381-3p 的表达，促进 IKKβ 和 p-p65 的上调。过表达 miR-381-3p 可抑制 IKKβ 的 mRNA 和蛋白表达。LPS 诱导的 BV2 细胞中 IKKβ、p-p65、肿瘤坏死因子-α（TNF-α）和白细胞介素-1β（IL-1β）的上调表达可被 miR-381-3p 模拟物转染逆转。此外，BV2 细胞中 miR-381-3p 抑制剂上调的 TNF-α和 IL-1β可被 IKKβ 抑制剂（BMS-345541）逆转。动物实验结果表明，ASCI 组大鼠 miR-381-3p 过表达。与对照组相比，ASCI 组大鼠 IKKβ 和 p-p65 蛋白水平以及 TNF-α和 IL-1β 炎症因子的 mRNA 表达水平均显著降低。ASCI 组大鼠的抓握能力、协调性和抗疲劳性能恢复速度快于对照组。
结论
ASCI 大鼠中 miR-381-3p 表达下调。过表达 miR-381-3p 可通过抑制 IKKβ-NF-κB 通路抑制炎症反应，更早恢复 ASCI 大鼠的运动能力，可能抑制损伤加重。
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