摘要
目的 从三叉神经血管系统中过氧化物酶体增殖物激活受体γ(PPARγ)/核转录因子κB(NF-κB)通路探讨电针风池、阳陵泉、外关治疗急性偏头痛发作期的可能机制.方法 40只SD大鼠随机分为空白组、模型组、电针组、电针+抑制剂组、激动剂组,每组8只.除空白组外,其余各组大鼠采用炎症汤颅内导管内注射建立急性偏头痛大鼠模型.造模后30 min,电针组予0.9％氯化钠溶液腹腔注射后电针干预"风池""阳陵泉""外关"30 min;电针+抑制剂组予PPARγ抑制剂T0070907(1.5 mg/kg)腹腔注射后再予电针干预上述穴位30 min;激动剂组予PPARγ激动剂盐酸吡格列酮(10mg/kg)腹腔注射;空白组与模型组大鼠予0.9％氯化钠溶液腹腔注射后捆绑束缚30 min.在造模前、造模30 min后(干预前)及干预后进行行为学评分;最后一次行为学检测后,采用实时荧光定量PCR法检测大鼠硬脑膜中PPARγ、NF-κB p65 mRNA含量;蛋白印迹法检测大鼠三叉神经脊束核中PPARγ、NF-κB p65、白细胞介素6(IL-6)、肿瘤坏死因子α(TNF-α)的蛋白表达;免疫荧光双标法检测大鼠三叉神经脊束核中PPARγ、NF-κB p65荧光强度.结果 与空白组同时间比较,其余各组大鼠造模后及干预后行为学评分均升高(P＜0.01).与模型组同时间比较,干预后电针组、电针+抑制剂组、激动剂组大鼠行为学评分均降低(P＜0.01).与电针组同时间比较,干预后激动剂组行为学评分降低,电针+抑制剂组行为学评分升高(P＜0.01).与空白组比较,模型组大鼠脑膜中PPARγ、NF-κB p65 mRNA表达升高,三叉神经脊束核中PPARγ、NF-κB p65、IL-6、TNF-α蛋白表达增强,PPARγ、NF-κB p65荧光强度增强(P＜0.01).与模型组比较,各干预组硬脑膜PPARγ mRNA表达升高、NF-κB p65 mRNA表达降低,三叉神经脊束核中PPARγ蛋白表达增强,NF-κB p65、IL-6、TNF-α蛋白表达减弱;电针组、激动剂组PPARγ荧光强度增强,各干预组NF-κB p65荧光强度均减弱(P＜0.05或P＜0.01).与电针组比较,电针+抑制剂组PPARγmRNA、蛋白表达及荧光强度降低,NF-κB p65 mRNA、蛋白表达及荧光强度升高,IL-6、TNF-α蛋白表达增强;激动剂组PPARγ mRNA、蛋白表达升高,NF-κB p65 mRNA、蛋白表达降低,IL-6、TNF-α蛋白表达降低(P＜0.05或P＜0.01).结论 电针"风池""阳陵泉""外关"可减轻急性偏头痛模型大鼠发作症状,其机制可能通过上调PPARγ表达、降低NF-κB表达,从而抑制神经源性炎症反应有关.
Abstract
Objective To investigate the possible mechanism of electroacupuncture at Fengchi(GB 20),Yanglingquan(GB 34)and Waiguan(TE 5)for acute migraine attacks by peroxisome proliferator-activated receptor y/nuclear transcription factor-κB pathway in the trigeminal neurovascular system.Methods Forty SD rats were ran-domly divided into blank group,model group,electroacupuncture group,electroacupuncture plus inhibitor group,and agonist group,8 rats in each group.Except for the blank group,rats were injected with inflammation decoction in-tracranial catheter to establish rat models of acute migraine.Thirty minutes after modelling,the electroacupuncture group was given 0.9％NaCl solution by intraperitoneal injection and then electroacupuncture at"Fengchi"(GB 20),"Yanglingquan"(GB 34),and"Waiguan"(TE 5)for 30 mins;the electroacupuncture plus inhibitor group was given PPARy inhibitor T0070907(1.5 mg/kg)and eclectroacupuncture as the above for 30 mins;the agonist group was given PPARγ inhibitor pioglitazone hydrochloride(10 mg/kg)for 30 mins.Rats in the blank group and the model group were injected intraperitoneally with 0.9％NaCl solution and then bound and restrained for 30 mins.Behavioural scores were evaluated before modelling,30 mins after modelling(pre-intervention)and post-intervention;after the last behavioural test,PPARγ,NF-κB p65 mRNA content in rat dura mater was detected by real-time fluorescence quantitative PCR;protein expression of PPARy,NF-κB p65,interleukin 6(IL-6),tumour necrosis factor α(TNF-α)in spinal trigeminal nucleus of rats was detected by protein blotting;immunofluorescence double-labelling method was used to detect the fluorescence intensity of PPARy,NF-κB p65 in spinal trigeminal nucleus of rats.Results Compared with the blank group at the same time,the behavioural scores of rats in the remaining groups increased after modelling and after intervention(P＜0.01).Compared with the model group at the same time,the beha-vioural scores of rats in the electroacupuncture group,electroacupuncture plus inhibitor group,and agonist group decreased after the intervention(P＜O.01).Compared with the electroacupuncture group at the same time,beha-vioural scores reduced in the agonist group and elevated in the electroacupuncture plus inhibitor group after interven-tion(P＜0.01).Compared with the blank group,expression of PPARγ and NF-κB p65 mRNA elevated in the dura mater of rats in the model group,and expression of PPARγ,NF-κB p65,IL-6,and TNF-α protein enhanced in spi-nal trigeminal nucleus,and the fluorescence intensity of PPARγ and NF-κB p65 enhanced(P＜0.01).Compared with the model group,PPARγ mRNA expression in dura mater elevated and NF-κB p65 mRNA expression reduced in each intervention group,PPARγ protein expression in spinal trigeminal nucleus enhanced,and NF-κB p65,IL-6,and TNF-α protein expression weakened;in the electroacupuncture group and the agonist group,PPARγ fluores-cence intensity enhanced,and the fluorescence intensity of NF-κB p65 weakened in each intervention group(P＜0.05 or P＜0.01).Compared with the electroacupuncture group,in the electroacupuncture plus inhibitor group,PPARy mRNA,protein expression and fluorescence intensity reduced,NF-κB p65 mRNA,protein expression and fluorescence intensity elevated,and IL-6 and TNF-α protein expression enhanced;in the agonist group,PPARγmRNA and protein expression elevated,NF-κB p65 mRNA and protein expression reduced,and IL-6 and TNF-α pro-tein expression decreased(P＜0.05 or P＜0.01).Conclusion Electroacupuncture at"Fengchi"(GB 20),"Yanglingquan"(GB 34)and"Waiguan"(TE 5)can can reduce the symptoms of acute migraine attacks in rats,and its mechanism may be related to the up-regulation of PPARγ expression and the reduction of NF-κB expression,thus inhibiting the neurogenic inflammatory response.
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