基于ROS-NLRP3炎症小体通路探讨夹脊穴注射糖皮质激素对大鼠带状疱疹神经痛的影响

Effect of Jiaji point injection of glucocorticoid on herpes zoster neuralgia in rats based on ROS-NLRP3 inflammatory body pathway

目的基于活性氧簇(ROS)-核苷酸结合寡聚化结构域样受体蛋白(NLRP)3炎症小体通路探讨夹脊穴注射糖皮质激素(GC)对大鼠带状疱疹神经痛的影响。方法雌性SD大鼠随机分为对照组、模型组、夹脊穴注射GC组(剂量为0.5 mg/kg)、ROS抑制剂N-乙酰-L-半胱氨酸(NAC)(剂量为20 mg/kg)组、夹脊穴注射GC+NAC组(剂量分别为0.5 mg/kg,20 mg/kg),除对照组外,其余各组于大鼠右足蹼底部皮下注射50μl的带状疱疹病毒(VZV)接种液建立带状疱疹神经痛大鼠模型,分别经夹脊穴注射GC、NAC治疗14 d后,以机械性疼痛刺激试验、热敏试验检测各组缩爪阈值、热敏潜伏期;以腺苷检测试剂盒检测各组外周组织液中腺苷水平,以酶联免疫吸附试验(ELISA)检测外周组织液中一磷酸腺苷(AMP)、二磷酸腺苷(ADP)水平;以ELISA检测各组血清肿瘤坏死因子(TNF)-α、白细胞介素(IL)-6水平;以ROS检测试剂盒检测各组脊髓组织ROS水平,Western印迹法检测脊髓组织NLRP3、caspase-1蛋白表达。结果与对照组比较,模型组缩爪阈值、热敏潜伏期、外周组织液中腺苷、AMP及ADP水平明显降低(P<0.05),血清IL-6及TNF-α水平、脊髓组织ROS水平、NLRP3、caspase-1蛋白表达明显升高(P<0.05);与模型组比较,夹脊穴注射GC组、NAC组、夹脊穴注射GC+NAC组缩爪阈值、热敏潜伏期、外周组织液中腺苷、AMP及ADP水平明显升高(P<0.05),血清IL-6及TNF-α水平、脊髓组织ROS水平、NLRP3、caspase-1蛋白表达明显降低(P<0.05);与夹脊穴注射GC组及NAC组分别比较,夹脊穴注射GC+NAC组缩爪阈值、热敏潜伏期、外周组织液中腺苷、AMP及ADP水平明显升高(P<0.05),血清IL-6及TNF-α水平、脊髓组织ROS水平、NLRP3、caspase-1蛋白表达明显降低(P<0.05)。结论夹脊穴注射GC可治疗大鼠带状疱疹神经痛,可能是通过下调ROS-NLRP3炎症信号实现的。

Objective To investigate the effect of Jiaji point injection of glucocorticoid(GC)on herpes zoster neuralgia in rats based on ROS-NLRP3 inflammatory body pathway.Methods SD rats were randomly divided into control,model,Jiaji point injection GC(dose 0.5 mg/kg),NAC(ROS inhibitor,20 mg/kg)and Jiaji point injection GC+NAC groups(0.5 mg/kg,20 mg/kg,respectively),except for the control group,the other groups were subcutaneously injected with 50μl of herpes zoster virus(VZV)vaccine at the bottom of the right webbed foot to establish the rat model of herpes zoster neuralgia.After 14 days of treatment,GC and NAC were injected into Jiaji point,mechanical pain stimulation test and thermal sensitivity test were used to detect the claw shrinkage threshold and thermal latency of rats in each group,adenosine levels in peripheral tissue fluid were detected by adenosine detection kit,and adenosine monophosphate(AMP)and adenosine diphosphate(ADP)levels in peripheral tissue fluid were detected by enzyme-linked immunosorbent assay(ELISA),the levels of serum TNF-αand IL-6 were measured by ELISA,reactive oxygen species(ROS)level in spinal cord tissue of rat in each group was detected by ROS test kit,and the expressions of NLRP3 and caspase-1 proteins in spinal cord tissue were detected by Western blot.Results Compared with control group,the claw shrinkage threshold,thermal latency,adenosine,AMP and ADP levels in peripheral tissue fluid of model group were significantly decreased(P<0.05),while serum IL-6 and TNF-αlevels,ROS level in spinal cord tissue,expressions of NLRP3 and caspase-1 proteins were increased(P<0.05).Compared with model group,the claw shrinkage threshold,thermal latency,adenosine,AMP and ADP levels in peripheral tissue fluid of Jiaji acupoint GC injection group,NAC group and GC+NAC group were significantly increased(P<0.05),while serum IL-6 and TNF-αlevels,ROS level in spinal cord tissue,expressions of NLRP3 and caspase-1 proteins decreased(P<0.05).Compared with the GC group and NAC group,the claw shrinkage threshold,thermal latency,adenosine,AMP and ADP levels in peripheral tissue fluid of the GC+NAC group were significantly increased(P<0.05),while serum IL-6 and TNF-αlevels,ROS level in spinal cord tissue,expressions of NLRP3 and caspase-1 proteins were decreased(P<0.05).Conclusions Injection of GC at Jiaji point ccould treat herpes zoster neuralgia in rats,which is possibly achieved by down-regulating the inflammatory signal of ROS-NLRP3.