“三法三穴”推拿手法对坐骨神经损伤大鼠痛觉功能和脊髓背角CX3CL1/CX3CR1表达的调节

目的 通过观察脊髓背角趋化因子(C-X3-C基元)配体1 (CX3CL1)/趋化因子(C-X3-C基元)受体1 (CX3CR1)蛋白和mRNA表达变化,探讨“三法三穴”推拿手法对坐骨神经损伤大鼠痛觉功能改善的作用机制。方法 74只雄性Sprague-Dawley大鼠,随机分为正常组(n = 12)、假手术组(n = 24)、模型组(n = 25)和三法三穴组(n = 13)。模型组和三法三穴组制作坐骨神经损伤模型。假手术组仅暴露坐骨神经。三法三穴组造模后第7天起用按摩推拿手法模拟仪按摩殷门、承山、阳陵泉三穴。分别于造模后7 d、干预20 d进行光热耐痛阈测定;造模后7 d、干预10 d、干预20 d进行累积疼痛评分测定;并于造模后7 d和干预20 d取材,对脊髓背角CX3CL1/CX3CR1表达进行Western blotting和RT-PCR检测,干预20 d对脊髓背角小胶质细胞进行免疫荧光观察。结果 造模后7 d,与正常组比较,模型组和假手术组光热耐痛阈升高(P < 0.05);与假手术组比较,模型组和三法三穴组累积疼痛评分升高( P < 0.05);干预10 d后,三法三穴组累积疼痛评分低于模型组( P < 0.05);干预20 d后,三法三穴组光热耐痛阈和累积疼痛评分均低于模型组( P < 0.05)。造模后7 d和干预20 d后,各组CX3CL1/CX3CR1蛋白和mRNA的表达变化均无显著性差异( P > 0.05)。干预20 d后模型组小胶质细胞呈部分活化或完全活化状态,三法三穴组呈未活化或部分活化状态。 结论 “三法三穴”推拿手法对坐骨神经损伤大鼠痛觉功能有改善,可能是通过CX3CL1/CX3CR1以外的途径调节小胶质细胞实现的。

Effect of Three Handing-Three Points on Pain Function and Expression of CX3CL1/CX3CR1 in Spinal Cord Dorsal Horn of Rats with Sciatic Nerve Injury

Objective To investigate the mechanism of Three Handing-Three Points on pain function in sciatic nerve injury rats by observing the changes of chemokine(C-X3-C motif)ligand 1,CX3 CL1)/chemokine(C-X3-C motif)receptor 1(CX3 CR1)protein and mRNA expression in spinal dorsal horn.Methods A total of 74 male Sprague-Dawley rats were randomly divided into normal group(n=12),sham group(n=24),model group(n=25),and Three Handing-Three Points group(Tuina group,n=13).The model group and Tuina group prepared the sciatic nerve injury model.The sham group exposed sciatic nerve only.Tuina group received Tuina on Yinmen(BL37),Chengshan(BL57)and Yanglingquan(GB34)with Tuina manipulation emulator.The photothermal pain threshold was measured seven days after modeling and after 20 days of intervention;cumulative pain score was measured seven days after modeling,and after ten days and 20 days of intervention.The spinal dorsal horn tissues were extracted to detect the protein and mRNA expression of CX3 CL1/CX3 CR1 with Western blotting and RT-PCR seven days after modeling and after 20 days of intervention.The microglia morphology in spinal dorsal horn was observed with immunofluorescence after 20 days of intervention.Results Seven days after modeling,compared with the normal group,the photothermal pain tolerance threshold increased in the model group and the sham group(P<0.05);compared with the sham group,the cumulative pain score increased in the model group and Tuina group(P<0.05).After ten days of intervention,the cumulative pain score was lower in Tuina group than in the model group(P<0.05).After 20 days of intervention,both the photothermal pain tolerance threshold and cumulative pain score were lower in Tuina group than in the model group(P<0.05).There was no significant difference in the expression of CX3 CL1/CX3 CR1 protein and mRNA on the seven days after modeling and after 20 days of intervention(P>0.05).The microglia in the model group were partially activated or completely activated,while those in Tuina group were unactivated or partially activated after 20 days of intervention.Conclusion Three Handing-Three Points can improve the pain function of sciatic nerve injured rats,which may associate with regulating microglia through the pathway other than CX3 CL1/CX3 CR1.