早期骨边刺结合电针抗骨癌痛大鼠吗啡耐受效应及其对背根神经节HDAC与MOR表达的影响

Early intervention of bone-nearby acupuncture combined with electroacupuncture on morphine tolerance in bone cancer pain rats and its effect on the expression of HDAC and MOR in dorsal root ganglia

目的:观察早期介入骨边刺结合电针对骨癌痛-吗啡耐受模型大鼠背根神经节(DRG)中组蛋白去乙酰化酶1(HDAC1)、组蛋白去乙酰化酶2(HDAC2)和μ阿片受体(MOR)蛋白表达的影响,探讨骨边刺结合电针抗骨癌痛大鼠吗啡耐受效应的可能作用机制。方法:将35只SD大鼠随机分为假手术组(6只)、骨癌痛组(7只)、吗啡耐受组(7只)、骨边刺结合电针组(8只)和假骨边刺组(7只)。除假手术组外,其余4组大鼠于左胫骨行癌细胞接种术制备骨癌痛模型;吗啡耐受组、骨边刺结合电针组和假骨边刺组大鼠予腹腔注射盐酸吗啡注射液诱导骨癌痛-吗啡耐受模型。骨边刺结合电针组于癌细胞接种术后15 d予骨边刺结合电针治疗,穴取双侧"足三里""昆仑",疏密波,频率2 Hz/100 Hz,电流强度0.5~1.5 mA;假骨边刺组介入时间及取穴同骨边刺结合电针组,仅刺破皮肤,不连接电针,两组均留针30 min,每日1次,连续治疗7 d。分别于癌细胞接种术前,术后10、11、15、22 d测量各组大鼠左侧机械缩足阈值(PWT)。采用Westernblot检测各组大鼠左侧DRG中HDAC1、HDAC2和MOR蛋白表达情况。结果:行癌细胞接种术后10d,与假手术组比较,骨癌痛组、吗啡耐受组、骨边刺结合电针组、假骨边刺组大鼠PWT均降低(P<0.01)。术后11 d,与骨癌痛组比较,吗啡耐受组、骨边刺结合电针组、假骨边刺组大鼠PWT明显升高(P<0.01)。术后22d,吗啡耐受组与骨癌痛组比较,差异无统计学意义(均P>0.05);与吗啡耐受组及假骨边刺组比较,骨边刺结合电针组大鼠PWT明显升高(P<0.01)。与假手术组比较,骨癌痛组大鼠DRG中HDAC1、HDAC2蛋白表达均升高(P<0.05),MOR蛋白表达降低(P<0.01)。与骨癌痛组比较,吗啡耐受组大鼠DRG中HDAC1蛋白表达升高(P<0.05)。与吗啡耐受组及假骨边刺组比较,骨边刺结合电针组大鼠DRG中HDAC1蛋白相对表达降低(P<0.01,P<0.05),MOR蛋白相对表达升高(均P<0.01)。结论:早期骨边刺结合电针干预可减轻骨癌痛模型大鼠的吗啡耐受现象,该效应可能与下调DRG中HDAC1蛋白表达、上调MOR蛋白表达有关。

Objective To observe the effect of early intervention of bone-nearby acupuncture(BNA)combined with electroacupuncture(EA)on the expression of histone deacetylasel(HDAC1),histone deacetylase 2(HDAC2)andμ-opioid recepter(MOR)in dorsal root ganglia(DRG)of bone cancer pain-morphine tolerance(BCP-MT)rats,and to explore its possible mechanism.Methods A total of 35 SD rats were randomized into a sham BCP group(n=6),a BCP group(n=7),a MT group(n=7),a BNA+EA group(n=8)and a shame BNA group(n=7).Except of the sham BCP group,cancer cell inoculation operation at left tibia was given in the other 4 groups to establish the bone cancer pain model.In the MT group,the BNA+EA group and the shame BNA group,intraperitoneal injection of morphine hydrochloride was given to establish the morphine tolerance model.After the operation,bone-nearby acupuncture combined with electroacupuncture was applied at"Zusanli"(ST 36)and"Kunlun"(BL 60)in the BNA+EA group,with dilatational wave,2 Hz/100 Hz in frequency,0.5 to 1.5 mA in intensity.Intervention in the shame BNA group was applied at the same time and acupoints as those in the BNA+EA group,the needles were pierced the skin without any electrical stimulation.The needles were retained for 30 min,once a day for continuous 7 days in both BNA+EA and shame BNA groups.Before and 10,11,15,22 days after the operation,the left paw withdrawal threshold(PWT)was measured in the 5 groups.The levels of HDAC1,HDAC2 and MOR in DRG were detected by Western blot.Results Ten days after the cancer cell inoculation operation,the PWT of the BCP,MT,BNA+EA and sham BNA groups was decreased compared with the sham BCP group(P<0.01).Eleven days after the operation,the PWT of the MT,BNA+EA and sham BNA groups was increased compared with the BCP group(P<0.01).Twenty-two days after the operation,the difference was no significant between the BCP group and MT group(P>0.05);the PWT of the BNA+EA group was increased compared with the MT and sham BNA group(P<0.01).In the BCP group,the DRG levels of HDAC1 and HDCA2 were increased,while the level of MOR was decreased compared with the sham BCP group(P<0.05,P<0.01).In the MT group,the DRG level of HDAC1 was increased compared with the BCP group(P<0.05).In the BNA+EA group,the DRG level of HDAC1 was decreased compared with the MT group and the sham BNA group(P<0.01,P<0.05),while the level of MOR was increased(P<0.01).Conclusion Early intervention of bone-nearby acupuncture combined with electroacupuncture can relieve the morphine tolerance in bone cancer pain rats,it may relate to down-regulating the expression of HD AC 1 and up-regulating the expression of MOR in the dorsal root ganglia.