基于TLR2靶点探讨脐灸对寒凝血瘀型痛经大鼠穴区启动-神经-内分泌-靶器官的调控作用

Discussion of regulation effect of umbilical moxibustion on the acupoint initiationneuron-endocrine-target organs in dysmenorrhea rats with cold congelation and blood stasis based on TLR2 as a target

目的:观察脐灸法对寒凝血瘀型痛经大鼠扭体反应及神阙穴区、下丘脑、脊髓、子宫组织中Toll样受体2(TLR2)表达的影响,初步探索脐灸法的穴区启动-神经-内分泌-靶器官的调控作用,完善脐灸镇痛效应信号通路。方法:将50只成年雌性Wistar大鼠随机分为5组,对照组、模型组、隔药灸脐组、药物贴脐组、艾条灸脐组。除对照组外均造寒凝血瘀型痛经模型,造模后给予相应治疗;采用免疫荧光法、蛋白免疫印迹法(Western Blot)、实时荧光定量PCR法(RT-PCR)检测神阙穴区、下丘脑、脊髓、子宫组织中TLR2蛋白及mRNA表达。结果:①脐灸组与模型组相比痛经模型大鼠寒凝血瘀状态改善,扭体潜伏期显著延长、扭体次数和评分降低(P<0.01),脐灸法具有显著的镇痛疗效,且隔药灸脐组优于艾条灸脐组及药物贴脐组(P<0.05,P<0.01)。②脐灸组与模型组相比可上调神阙穴区TLR2蛋白及mRNA表达(P<0.05,P<0.01),其中艾条灸脐组疗效优于隔药灸脐组及药物贴脐组。③脐灸组与模型组相比可下调下丘脑、脊髓及子宫TLR2蛋白及mRNA表达(P<0.01),其中下丘脑组织中的艾条灸脐组疗效优于隔药灸脐组及药物贴脐组(P<0.05,P<0.01),子宫及脊髓组织中的隔药灸脐组疗效最优(P<0.05,P<0.01)。结论:脐灸法通过上调神阙穴区TLR2蛋白和mRNA表达以激活穴区信息启动及下调下丘脑-脊髓-子宫中TLR2蛋白及mRNA表达以抑制中枢神经系统及靶器官中肥大细胞功能进而发挥镇痛疗效。

Objective:To observe the effects of umbilical moxibustion on the writhing reaction and the expression of Toll-like receptor 2(TLR2)in Shenque(CV 8)acupoint area,hypothalamus,spinal cord and uterus of dysmenorrhea rats with cold congelation and blood stasis.And to explore the acupoint initiation-neuronal-endocrine-target organ regulation,so as to improve the analgesic effect signal pathway of umbilical moxibustion.Methods:Fifty adult female Wistar rats were randomly divided into 5 groups:control group,model group,herb-partition moxibustion group(HM),drug-coving navel therapy group(DC)and moxibustion navel therapy group(MB).Except for the control group,all rats were made cold coagulation and blood stasis model,and the corresponding treatment was given after modeling.The expression of TLR2 protein and mRNA in the Shenque(CV 8)acupoint area,hypothalamus,spinal cord and uterus tissue were detected by immunofluorescence,Western Blot and Realtime PCR methods.Results:①Compared with the model group,the cold coagulation and blood stasis state of dysmenorrhea model rats in umbilical moxibustion therapy group were improved.At the same time,the incubation period of writhing was significantly prolonged,and the times and scores of writhing were decreased(P<0.01).Frequency of writhing response and has obvious analgesic effect on dysmenorrhea rats.The umbilicus moxibustion had significant analgesic effect,and the HM group was better than DC group and MB group(P<0.01,P<0.05).②Compared with the model group,the umbilical moxibustion therapy group could up-regulate the expression of TLR2 protein and mRNA in Shenque(CV 8)acupoint area(P<0.05,P<0.01),and the therapeutic effect of MB group was better than that of HM group and DC group.③Compared with the model group,the umbilical moxibustion therapy group could down regulate the expression of TLR2 protein and mRNA in the hypothalamus,spinal cord and uterus tissue(P<0.01).The effect of the MB group in the hypothalamic tissue was better than that of the HM group and the DC group(P<0.05,P<0.01).The effect of the HM group in the uterus and spinal cord tissue was the best(P<0.05,P<0.01).Conclusion:Umbilical therapy activates the acupoint information by up-regulating the expression of TLR2 protein and mRNA in Shenque(CV 8)acupoint area and inhibits mast cell function in the central nervous system and target organs by down-regulating TLR2 protein and mRNA expression in the hypothalamus-spinal cord-uterus tissues.