S1脑区CXCL1/CXCR2参与针刺改善佐剂性关节炎大鼠炎性痛的作用机制  

Acupuncture alleviates inflammatory pain by activating CXCL1/CXCR2 signaling in the primary somatosensory cortex in adjuvant induced arthritis rats

作　　者：李艳伟 窦报敏 王慎军 吕中茜[1] 李威 徐枝芳 刘阳阳[1] 徐媛[1] 房钰鑫[1] 郭永明[1] 郭义[1] LI Yan-wei;DOU Bao-min;WANG Shen-jun;LÜZhong-xi;LI Wei;XU Zhi-fang;LIU Yang-yang;XU Yuan;FANG Yu-xin;GUO Yong-ming;GUO Yi(Research Center of Experimental Acupuncture-moxibustion,Tianjin University of Traditional Chinese Medicine,Tianjin 301617,China)

机构地区：[1]天津中医药大学实验针灸学研究中心,天津301617

出　　处：《针刺研究》2024年第4期331-340,共10页Acupuncture Research

基　　金：国家自然科学基金重点项目(No.82030125);国家自然科学基金面上项目(No.81873369、81973939)。

摘　　要：目的:探究大脑初级体感皮层(S1)脑区趋化因子CXC配体1(CXCL1)及趋化因子CXC受体2(CXCR2)参与针刺镇痛的作用机制,进一步揭示针刺镇痛的神经-免疫学机制。方法:将热痛阈值相对稳定的BALB/c小鼠足底注射0.05 mL完全弗氏佐剂(CFA)建立佐剂性关节炎(AIA)模型后,随机分为单纯荧光组和荧光标记CXCL1组,每组2只。荧光标记CXCL1组小鼠尾静脉注射AF750荧光标记CXCL1重组蛋白,并通过小动物活体成像技术,观察荧光及荧光标记CXCL1在AIA小鼠体内的主要富集脏器及脑的荧光富集情况。另取热痛阈值相对稳定的Wistar大鼠,右足底注射0.1 mL CFA建立AIA模型,随机分为模型组、针刺组,每组12只,取12只Wistar大鼠右足底注射等体积的0.9%氯化钠溶液作为盐水组。针刺组大鼠针刺“足三里”1次/d,针刺持续7 d,治疗结束后采用热痛仪测量大鼠右侧足底热缩足潜伏期即热痛阈值;利用ELISA、免疫荧光及实时荧光定量PCR技术,检测文献报道的可能表达CXCR2及阿片肽受体的多个疼痛相关脑区CXCL1蛋白含量及S1脑区CXCL1阳性细胞数与mRNA表达水平,以期探明前期研究结果中针刺后升高的血清CXCL1的作用脑区;采用ELISA法检测各组大鼠S1脑区中CXCR2蛋白含量,采用实时荧光定量PCR法检测S1脑区中CXCR2、μ-阿片肽受体(MOR)的mRNA表达,采用免疫荧光双标法将CXCR2分别与星形胶质细胞标记物胶质纤维酸性蛋白(GFAP)、神经元标记物NeuN及MOR共染,检测CXCR2在S1脑区中的表达细胞及是否与MOR共表达。结果:与单纯荧光组相比,荧光标记CXCL1组小鼠脑组织中有较强的荧光信号。与盐水组相比,模型组大鼠右侧足底热痛阈值均明显降低(P<0.01),S1脑区中CXCR2 mRNA表达升高(P<0.05);与模型组相比,针刺组第2~7天热痛阈值、S1脑区中CXCL1蛋白含量、CXCR2蛋白含量及mRNA表达、MOR mRNA表达均升高(P<0.05,P<0.01);针刺组大鼠S1脑区中CXCR2分别与NeuN及MOR有共�Objective To observe whether acupuncture up-regulates chemokine CXC ligand 1(CXCL1)in the brain to play an analgesic role through CXCL1/chemokine CXC receptor 2(CXCR2)signaling in adjuvant induced arthritis(AIA)rats,so as to reveal its neuro-immunological mechanism underlying improvement of AIA.Methods BALB/c mice with relatively stable thermal pain reaction were subjected to planta injection of complete Freund adjuvant(CFA)for establishing AIA model,followed by dividing the AIA mice into simple AF750(fluorochrome)and AF750+CXCL1 groups(n=2 in each group).AF750 labeled CXCL1 recombinant protein was then injected into the mouse’s tail vein to induce elevation of CXCL1 level in blood for simulating the effect of acupuncture stimulation which has been demonstrated by our past study.In vivo small animal imaging technology was used to observe the AF750 and AF750+CXCL1-labelled target regions.After thermal pain screening,the Wistar rats with stable pain reaction were subjected to AIA modeling by injecting CFA into the rat’s right planta,then were randomized into model and manual acupuncture groups(n=12 in each group).Other 12 rats that received planta injection of saline were used as the control group.Manual acupuncture(uniform reinforcing and reducing manipulations)was applied to bilateral“Zusanli”(ST36)for 4×2 min,with an interval of 5 min between every 2 min,once daily for 7 days.The thermal pain threshold was assessed by detecting the paw withdrawal latency(PWL)using a thermal pain detector.The contents of CXCL1 in the primary somatosensory cortex(S1),medial prefrontal cortex,nucleus accumbens,amygdala,periaqueductal gray and rostroventromedial medulla regions were assayed by using ELISA,and the expression levels of CXCL1,CXCR2 and mu-opioid receptor(MOR)mRNA in the S1 region were detected using real time-quantitative polymerase chain reaction.The immune-fluorescence positive cellular rate of CXCL1 and CXCR2 in S1 region was observed after immunofluorescence stain.The immunofluorescence double-stain of CXC

关 键 词：针刺镇痛 佐剂性关节炎 炎性痛 CXC配体1/CXC受体2 大脑初级体感皮层区(S1) 阿片肽受体 

分 类 号：R245[医药卫生—针灸推拿学]