依托咪酯对慢性神经病理性疼痛大鼠脊髓小胶质细胞活化的影响及其作用机制

伍兰; 田彬; 向红; 张贵升; 董航

doi:10.16190/j.cnki.45-1211/r.2023.03.015

依托咪酯对慢性神经病理性疼痛大鼠脊髓小胶质细胞活化的影响及其作用机制

Effect of etomidate on spinal microglia activation in rats with chronic neuropathic pain and its mechanism

摘要

摘要: 目的：探究依托咪酯（Eto）调节CXC趋化因子配体12（CXCL12）/CXC趋化因子受体4（CXCR4）信号通路对慢性神经病理性疼痛（NP）大鼠脊髓小胶质细胞活化的影响。方法：取SD 大鼠36 只，随机分为6 组：Sham 组、Model 组、阳性药对照组（150 mg/kg 阿魏酸钠）、Eto 低剂量组（0.3 mg/kg）、Eto 中剂量组（0.9 mg/kg）、Eto 高剂量组（2.7 mg/kg），每组6 只。除Sham 组外，其余各组采用坐骨神经慢性压迫损伤法（CCI）构建大鼠NP模型。造模成功后，尾静脉注射Eto或灌胃给予阿魏酸钠，每天1次，连续4周。分别于术前2 h和术后3 d、5 d、7 d、10 d测定各组大鼠机械性缩足反射阈值（MWT）和热刺激缩足反射潜伏期（TWL）；对脊髓L4～L6行免疫组化染色，观察小胶质细胞离子钙接头蛋白分子1（Iba-1）蛋白表达，实时荧光定量PCR（qPCR）检测Iba-1 mRNA 表达，酶联免疫吸附（ELISA）法检测脊髓中肿瘤坏死因子（TNF）-α、白细胞介素（IL）-6、IL-1β 含量，western blotting 法检测脊髓CXCL12、CXCR4、Iba-1 蛋白表达。结果：与Sham 组相比，Model 组大鼠的MWT、TWL 显著降低，Iba-1 表达的IOD 值，Iba-1 mRNA 表达，TNF-α、IL-6、IL-1β 含量及CXCL12、CXCR4、Iba-1 蛋白表达升高（P＜0.05）；与Model 组相比，Eto低、中、高剂量组大鼠的MWT、TWL显著升高，Iba-1表达的IOD值，Iba-1 mRNA表达，TNF-α、IL-6、IL-1β含量及CXCL12、CXCR4、Iba-1 蛋白表达降低（P＜0.05），且Eto 高剂量组与阳性药对照组大鼠的上述指标均无显著差异（P＞ 0.05）。结论：Eto可能通过抑制CXCL12/CXCR4信号通路，抑制炎症反应与脊髓小胶质细胞活化，从而起到缓解NP的作用。

Abstract: Objective: To explore the effect of etomidate (Eto) on the activation of spinal microglia in chronic neuropathic pain (NP) rats by regulating CXC chemokine ligand 12 (CXCL12)/CXC chemokine receptor 4 (CXCR4) signal pathway.Methods: A total of 36 SD rats were randomly divided into 6 groups (6 rats/group): Sham group, Model group, positive control group (150 mg/kg sodium ferulate), Eto low dose group (0.3 mg/kg), Eto medium dose group(0.9 mg/kg), and Eto high dose group(2.7 mg/kg).Except for the Sham group, the NP model of rats was established by chronic compression injury of sciatic nerve (CCI).After successful modeling, Eto was injected through tail vein or sodium ferulate was administered by gavage once a day for 4 weeks.Mechanical foot contraction reflex threshold (MWT) and thermal stimulation foot contraction reflex latency (TWL) were measured 2 hours before operation and 3, 5, 7 and 10 days after operation respectively; immunohistochemical staining was performed on L4-L6 spinal cords to observe the expression of ionic calcium junction protein 1 (Iba-1) protein in microglia; the expression of Iba-1 mRNA was detected by real-time fluorescent quantitative polymerase chain reaction (RT-qPCR); the contents of tumor necrosis factor(TNF)-α, interleukin(IL)-6 and IL-1β in the spinal cord were detected by enzyme-linked immunosorbent assay(ELISA); western blotting was used to detect the expressions of CXCL12, CXCR4 and Iba-1 proteins in the spinal cord.Results: Compared with Sham group, the MWT and TWL in rats of Model group were obviously reduced, while the IOD value of Iba-1 expression and Iba-1 mRNA expression, contents of TNFα, IL-6, IL-1β, expressions of CXCL12, CXCR4, Iba-1 protein increased (P＜0.05); compared with the Model group, the MWT and TWL in rats of Eto low, medium and high dose groups obviously increased, the IOD value of Iba-1 expression, Iba-1 mRNA expression, contents of TNF-α, IL-6 and IL-1β and expressions of CXCL12, CXCR4, Iba-1 protein decreased (P＜0.05), and there was no obvious difference between Eto high dose group and positive control group in the above indicators(P＞ 0.05).Conclusion: Eto may play a role in relieving NP by inhibiting CXCL12/CXCR4 signal pathway, inhibiting inflammatory reaction and activation of spinal microglia.

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