基于α7nAChRs的针刺对慢性偏头痛大鼠抗炎作用的研究

The Study of Anti-inflammatory Effect of Acupuncture in Rats with Chronic Migraine Based on theα7nAChRs

目的探讨针刺对慢性偏头痛(Chronic Migraine,CM)大鼠α7烟碱型乙酰胆碱受体(α7 nicotinic acetylcholine receptors,α7nAChRs)影响炎症变化的作用。方法将成年雄性SD大鼠根据基础痛阈随机分为对照组(VEH组)、模型组(NTG组)、模型+针刺组(TA组)、模型+针刺+α7nAChRs拮抗剂MLA组(MLA组)、模型+α7nAChRs激动剂PNU-282987组(PNU组)。采用隔日颈背部皮下重复注射硝酸甘油(NTG)建立CM大鼠模型。TA组和MLA组于NTG注射前1 h针刺,20 min/天,连续9天。MLA组于针刺前0.5 h腹腔注射MLA,PNU组于NTG注射前0.5 h腹腔注射PNU-282987,连续9天。采用Von Frey和热辐射测痛仪检测各组大鼠足底机械痛阈值(Paw Withdrawal Mechanical Threshold,PWMT)和甩尾潜伏期(Tail-Flick Latency,TFL)的变化;ELISA法检测各组大鼠血清及TNC中炎性因子IL-1β、TNF-α和TGF-β含量;免疫荧光双标法检测TNC中GFAP和α7nAChRs的平均光密度和共表达情况。结果与VEH组相比,NTG组PWMT、TFL明显降低(P<0.05或P<0.01),血清和TNC中IL-1β(P<0.01)、TNF-α(P<0.01,P<0.05)含量明显增多,TNC中AS明显活化(P<0.01),而α7nAChRs平均光密度和GFAP与α7nAChRs共表达明显减少(P<0.05,P<0.01);与NTG组相比,TA组PWMT、TFL升高(P<0.05或P<0.01),血清和TNC中IL-1β(P<0.01)、TNF-α(P<0.01,P<0.05)含量增多,TNC中GFAP平均光密度明显降低(P<0.01),而α7nAChRs平均光密度和GFAP与α7nAChRs共表达明显增加(P<0.01);与MLA组比较,TA组和PNU组PWMT、TFL明显升高(P<0.05或P<0.01),TA组血清和TNC中IL-1β(P<0.05)、TNF-α(P<0.01)含量增多而TGF-β(P<0.05)含量减少,PNU组血清和TNC中IL-1β含量减少(P<0.01,P<0.05)而TGF-β(P<0.01)含量明显增多。结论针刺能够有效缓解CM炎症反应和痛觉超敏状态,其抗炎镇痛效应可能与上调α7nAChRs的表达有关。

Objective To investigate the effect of acupuncture on the inflammatory changes ofα7 nicotinic acetylcholine receptors(α7 nicotinic acetylcholine receptors,α7nAChRs)in rats with chronic migraine(CM).Methods Adult male SD rats were randomly divided into control group(VEH group),model group(NTG group),acupuncture group(TA group),α7nAChRs antagonist group(MLA group)andα7nAChRs agonist group(TA group)according to basic pain threshold.The rat model of CM was established by repeated subcutaneous injection of nitroglycerin(NTG)at the neck and back every other day.TA group and MLA group received acupuncture treatment one hour before NTG injection,20 minutes every day for 9 consecutive days.The MLA group was intraperitoneally injected withα7nAChRs antagonist MLA half an hours before acupuncture,and the PNU group was intraperitoneally injected withα7nAChRs agonist PNU-282987 half an hours before NTG injection,both groups were injected continuously for 9 days.The changes of paw withdrawal mechanical threshold(Paw Withdrawal Mechanical Threshold,PWMT)and tail-flick latency(Tail-Flick Latency,TFL)were detected by Von Frey and thermal radiation pain meter.The contents of inflammatory factors IL-1β,TNF-αand TGF-βin TNC and serum were determined by ELISA.The relative fluorescence intensity and coexpression of GFAP andα7nAChRs in TNC were detected by immunofluorescence dual labeling method.Results Compared with VEH group,PWMT and TFL in NTG group were significantly decreased(P<0.05 or P<0.01),the contents of IL-1β(P<0.01)and TNF-α(P<0.01,P<0.05)in serum and TNC were significantly increased;and in TNC,the astrocytes were activated significantly(P<0.01),while the relative fluorescence intensity ofα7nAChRs and the coexpression of GFAP andα7nAChRs were significantly decreased(P<0.05,P<0.01).Compared with NTG group,PWMT and TFL in TA group were increased(P<0.05 or P<0.01),the contents of IL-1β(P<0.01)and TNF-α(P<0.01,P<0.05)in serum and TNC were increased;and in TNC,the relative fluorescence intensity of GFAP was significantly decreased(P<0.01),however the relative fluorescence intensity ofα7nAChRs and the co-expression of GFAP andα7nAChRs were significantly increased(P<0.01).Compared with MLA group,PWMT and TFL in TA groups and PNU groups were significantly increased(P<0.05 or P<0.01).The contents of IL-1β(P<0.05)and TNF-α(P<0.01)in serum and TNC of TA group were increased,while the contents of TGF-β(P<0.05)were decreased.The contents of IL-1βin serum and TNC of PNU group were decreased(P<0.01,P<0.05),while the contents of TGF-β(P<0.01)were significantly increased.Conclusions Acupuncture can effectively relieve CM inflammatory response and hypersensitivity to pain,and its anti-inflammatory and analgesic effects may be related to the up-regulation ofα7nAChRs expression.