乌头对寒、热痹证大鼠痛觉敏感干预作用的药效学及作用机制研究

Study on pharmacodynamics of the intervention effect of aconite on pain sensitivity of rats with syndromes of cold and heat bi and its mechanism of action

目的通过温热药乌头对寒、热痹证的干预作用,以确定其对寒痹、热痹作用的差异。方法本实验采用完全弗式佐剂致大鼠足跖关节炎症,在此基础上叠加致病因素风寒湿、风热湿复制寒痹、热痹证模型。造模前随机分为9组,分别是对照组、布洛芬组、模型组、热痹证组、寒痹证组、乌头热痹生药0.8 g/kg组、乌头热痹生药1.6 g/kg组、乌头寒痹痹生药0.8 g/kg组、乌头寒痹生药/kg组,造模后每天灌胃给药1次,共14天。并分别在第0天、3天、7天、10天、14天测量每组大鼠右后肢的痛反应时间,第15天取材后,用蛋白质印迹法(Western blot)法测定大鼠脊髓中不规则趋化因子(fractalkine,FKN)、趋化因子受体(chemokine CX3C receptor1, CX3CR1)和N-甲基-D-天门冬氨酸受体(N-methyl-D-aspartate receptor,NMDAR)的表达,用免疫组织化学染色法检测大鼠背根神经节(DRG)中FKN、CX3CR1、CD11b/c和NMDAR的表达。结果模型组大鼠患足的痛反应时间以明显低于对照组(与对照组比较,P<0.01),寒凉因素可抑制大鼠患足对温度刺激敏感性的升高,湿热因素引入后作用不明显,最终热痹证的痛反应时间均低于模型组。温热药乌头可延长寒痹证组的痛反应时间,对热痹证组的作用明显相反。脊髓和被根神经节的实验结果与痛反应时间结果基本保持一致。结论温热药乌头对寒、热痹证大鼠痛觉敏感的干预作用,与其抑制脊髓和背根神经节中的FKN、CX3CR1、CD11b/c和NMDAR的表达存在关联性。

Objective To determine the effect differences of aconite between cold and heat bi,by exploring the intervention effect of aconite with warm-hot nature on syndromes of cold and heat bi.Methods In this experiment,complete Freund’ s adjuvant was used to induce the inflammation of foot metatarsal joint in rats,and on this basis,the pathogenic factors of wind-cold-dampness and wind-heatdampness were superimposed to replicate the model rats with syndromes of cold and heat bi.Before modeling,rats were randomly divided into 9 groups,including control group,ibuprofen group,model group,heat bi group,cold bi group,aconite-heat bi-crude drug group(0.8 g/kg),aconite-heat bi-crude drug group(1.6 g/kg),aconite-cold bi-crude drug group(0.8 g/kg),aconite-cold bi-crude drug group(1.6 g/kg),and aconite-cold bi-crude drug group(1.6 g/kg).After the model was established,the drug was administered by gavage once a day,for a total of 14 days.The pain reaction time of the right hind limb of rats in each group was measured at day 0,day 3,day 7,day 10 and day 14,respectively.The expression of fractalkine(FKN),chemokine CX3 C receptor(CX3 CR1) and N-methyl-D-aspartate receptor(NMDAR) in rats’ spinal cord was determined by Western blot on the 15 th day after sampling.The expressions of FKN,CX3 CR1,CDllb/c and NMDAR in dorsal root ganglion(DRG) of rats were detected by immunohistochemical staining.Results The pain reaction time of rats in the model group was significantly lower than lower than that in the control group(compared with the control group,P<0.01).The cold and cool factors could inhibit the growth of rats’ disease foot being sensitive totemperature stimulation,after the introduction of heat and humid factors,the effect was not obvious,and finally the pain reaction time of heat bi syndrome was lower than that in the model group.Aconite with warm-hot nature can prolong the pain reaction time of rats in the cold bi syndrome group,but it has opposite effect on the heat bi syndrome group.The experiment results of spinal cord and dorsal root ganglia were consistent with the results of pain reaction time.Conclusion Aconite with warm-hot nature have the intervention effect on rats with syndromes of cod and heat bi being sensitive to pain.It was associated with inhibiting the expressions of FKN,CX3 CR1,CDllb/c and NMDAR in spinal cord and dorsal root ganglia.