电针对骨癌痛-吗啡耐受大鼠蓝斑核μ阿片受体表达的干预
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摘要
目的:观察电针对骨癌痛-吗啡耐受(cancer pain and morphine tolerance)大鼠痛行为的影响及部分作用机制。方法:将42只健康雌性SD大鼠完全随机分为5组:假手术组(7只)、骨癌痛组(8只)、吗啡耐受组(9只)、电针组(9只)和假电针组(9只)。假手术组于左胫骨髓腔内注射无菌磷酸盐缓冲液,其余4组大鼠均于左胫骨髓腔内注射MRMT-1乳腺癌细胞以制备骨癌痛模型。假手术组及骨癌痛组术后均不予干预。吗啡耐受组、电针组及假电针组于术后第11d对骨癌痛制备成功大鼠行盐酸吗啡注射液腹腔注射,每12h注射1次,连续注射11d诱导骨癌痛-吗啡耐受模型。此模型建立后第1d起,吗啡耐受组每12h(早上9:00,晚上9:00)行吗啡注射,连续7d;电针组、假电针组均于早上9:00行吗啡注射,30min后分别给予电针(2Hz/100Hz)和假电针(仅刺入皮下,破皮即可)治疗,穴取双侧"足三里"和"昆仑",每次30min,每日1次,连续治疗7d。分别于癌细胞接种前1d,术后第6d、8d、10d,吗啡注射第1d、5d、9d、11d的30min后及电针治疗第1d、3d、5d、7d的30min后检测各组大鼠患侧机械缩足阈(paw withdrawal threshold,PWT)的变化。于吗啡注射第11d,采用HE染色检测假手术组、骨癌痛组、吗啡耐受组大鼠(每组随机选2只)胫骨组织形态学变化;电针治疗第7d采用荧光免疫组织化学法观察各组大鼠(每组随机选4只)蓝斑核μ阿片受体(μ-opioid receptor,MOR)阳性细胞表达情况。结果:癌细胞接种后第10d,28只骨癌痛造模成功大鼠(骨癌痛组8只、吗啡耐受组8只、电针组6只、假电针组6只)PWT较7只假手术组大鼠明显下降(P<0.01)。吗啡注射第1d,吗啡耐受组、电针组及假电针组大鼠PWT均明显高于骨癌痛组(均P<0.01);吗啡连续注射第11d,吗啡耐受组、电针组、假电针组大鼠PWT与骨癌痛组比较差异均无统计学意义(均P>0.05)。吗啡注射第11d,骨癌痛组、吗啡耐受组大鼠胫骨上1/3处均可见癌细胞致肿块,且胫骨髓腔内布满MRMT-1癌细胞;假手术组大鼠胫骨未见异常变化。电针治疗第1d、3d、5d、7d,骨癌痛组、吗啡耐受组和假电针组大鼠患侧PWT均明显低于电针组(均P<0.01)。电针治疗第7d,骨癌痛组、吗啡耐受组、电针组、假电针组蓝斑核MOR阳性表达均低于假手术组(P<0.01,P<0.05),且骨癌痛组、吗啡耐受组和假电针组均低于电针组(均P<0.01)。结论:电针可提高骨癌痛-吗啡耐受大鼠的机械痛阈,改善模型大鼠痛觉异常;该效应可能与电针提高大鼠蓝斑核MOR阳性细胞表达有关。
Objective To observe the effects of electroacupuncture(EA)on pain behavior in rats with bone cancer pain and morphine tolerance,and to explore partial action mechanism.Methods Forty-two SD healthy female rats were randomly divided into a sham operation group(7rats),a bone cancer pain group(8rats),a morphine tolerance group(9rats),an EA group(9rats)and a sham EA group(9rats).The rats in the sham operation group were treated with injection of phosphate buffer saline at medullary cavity of left-side tibia,and the rats in the remaining groups were injected with MRMT-1breast cancer cells.After operation,no treatment was given to rats in the sham operation group and bone cancer pain group.11 days after operation,rats in the morphine tolerance group,EA group and sham EA group were treated with intraperitoneal injection of morphine hydrochloride,once every 12 hours,for 11 days to establish the model of bone cancer pain and morphine tolerance.One day after the establishment of this bone cancer pain model,the rats in the morphine tolerance group were injected with morphine,once every 12hours(9:00a.m.and 9:00p.m.)for 7days;the rats in the EA group and sham EA group were injected with morphine at 9:00a.m.,and treated with EA(2Hz/100Hz)and sham EA(only injected into the subcutaneous tissue)at bilateral"Zusanli"(ST 36)and"Kunlun"(BL 60),30 min per treatment,once a day for 7days.One day before cancer cell injection,6days,8days,10 days after operation,after 30 min on 1days,5days,9days,11 days of morphine injection,and after 30 min on 1days,3days,5days,7days of EA treatment,the paw withdrawal threshold(PWT)was measured in each group.On 11 day of morphine injection,HE staining was applied to observe the morphology and structure change of tibia in the sham operation group,bone cancer pain group and morphine tolerance group,random 2rats in each group.On 7days of EA treatment,fluorescent immunohistochemical method was applied to observe the expression ofμ-opioid receptor positive cells in nucleus ceruleus in each group,random 4rats in each one.Results After 10 days of the cancer cells injection,the PWT of 28 rats of bone cancer pain model(8rats in the bone cancer pain group,8rats in the morphine tolerance group,6rats in the EA group and 6rats in the sham EA group)was significantly lower than that of 7rats in the sham operation group(P<0.01).After one day of morphine injection,the PWT of the morphine tolerance group,EA group and sham EA group was higher than that of the bone cancer pain group(all P<0.01);on 11 dof morphine injection,the PWT of the morphine tolerance group,EA group and sham EA group was not significantly different from that of the bone cancer pain group(all P>0.05).On 11 dof morphine injection,the tumor induced by cancer cells was observed in upper 1/3tibia in the bone cancer pain group and morphine tolerance group,and the marrow cavity was filled with MRMT-1cancer cells;no abnormal change was observed in the sham operation group.On1 d,3d,5dand 7dof EA treatment,the PWT of the cancer pain group,morphine tolerance group and sham EA group was lower than that of the EA group(all P<0.01).On 7dof EA treatment,the positive expression of MOR in nucleus ceruleus in the cancer pain group,morphine tolerance group,EA group and sham EA group was lower than that in the sham operation group(P<0.01,P<0.05),and that in the cancer pain group,morphine tolerance group and sham EA group was lower than that in the EA group(all P<0.01).Conclusion EA can improve mechanical pain threshold in rats with bone cancer pain-morphine tolerance,and improve the abnormal pain,which is likely to be involved with improvement of the MOR positive cells expression in nucleus ceruleus by EA.
Objective To observe the effects of electroacupuncture(EA)on pain behavior in rats with bone cancer pain and morphine tolerance,and to explore partial action mechanism.Methods Forty-two SD healthy female rats were randomly divided into a sham operation group(7rats),a bone cancer pain group(8rats),a morphine tolerance group(9rats),an EA group(9rats)and a sham EA group(9rats).The rats in the sham operation group were treated with injection of phosphate buffer saline at medullary cavity of left-side tibia,and the rats in the remaining groups were injected with MRMT-1breast cancer cells.After operation,no treatment was given to rats in the sham operation group and bone cancer pain group.11 days after operation,rats in the morphine tolerance group,EA group and sham EA group were treated with intraperitoneal injection of morphine hydrochloride,once every 12 hours,for 11 days to establish the model of bone cancer pain and morphine tolerance.One day after the establishment of this bone cancer pain model,the rats in the morphine tolerance group were injected with morphine,once every 12hours(9:00a.m.and 9:00p.m.)for 7days;the rats in the EA group and sham EA group were injected with morphine at 9:00a.m.,and treated with EA(2Hz/100Hz)and sham EA(only injected into the subcutaneous tissue)at bilateral"Zusanli"(ST 36)and"Kunlun"(BL 60),30 min per treatment,once a day for 7days.One day before cancer cell injection,6days,8days,10 days after operation,after 30 min on 1days,5days,9days,11 days of morphine injection,and after 30 min on 1days,3days,5days,7days of EA treatment,the paw withdrawal threshold(PWT)was measured in each group.On 11 day of morphine injection,HE staining was applied to observe the morphology and structure change of tibia in the sham operation group,bone cancer pain group and morphine tolerance group,random 2rats in each group.On 7days of EA treatment,fluorescent immunohistochemical method was applied to observe the expression ofμ-opioid receptor positive cells in nucleus ceruleus in each group,random 4rats in each one.Results After 10 days of the cancer cells injection,the PWT of 28 rats of bone cancer pain model(8rats in the bone cancer pain group,8rats in the morphine tolerance group,6rats in the EA group and 6rats in the sham EA group)was significantly lower than that of 7rats in the sham operation group(P<0.01).After one day of morphine injection,the PWT of the morphine tolerance group,EA group and sham EA group was higher than that of the bone cancer pain group(all P<0.01);on 11 dof morphine injection,the PWT of the morphine tolerance group,EA group and sham EA group was not significantly different from that of the bone cancer pain group(all P>0.05).On 11 dof morphine injection,the tumor induced by cancer cells was observed in upper 1/3tibia in the bone cancer pain group and morphine tolerance group,and the marrow cavity was filled with MRMT-1cancer cells;no abnormal change was observed in the sham operation group.On1 d,3d,5dand 7dof EA treatment,the PWT of the cancer pain group,morphine tolerance group and sham EA group was lower than that of the EA group(all P<0.01).On 7dof EA treatment,the positive expression of MOR in nucleus ceruleus in the cancer pain group,morphine tolerance group,EA group and sham EA group was lower than that in the sham operation group(P<0.01,P<0.05),and that in the cancer pain group,morphine tolerance group and sham EA group was lower than that in the EA group(all P<0.01).Conclusion EA can improve mechanical pain threshold in rats with bone cancer pain-morphine tolerance,and improve the abnormal pain,which is likely to be involved with improvement of the MOR positive cells expression in nucleus ceruleus by EA.
引文
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[2]Connor M,Osborne PB,Christie MJ.Mu-opioid receptor desensitization:is morphine different?[J].Br J Pharmacol,2004,143(6):685-696.
[3]吕庆琴,陈霆隽,洪炎国.μ阿片受体的内吞抑制吗啡耐受的形成[J].中国临床药理学与治疗学,2012,17(10):1185-1190.
[4]Paley CA,Johnson MI,Tashani OA,et al.Acupuncture for cancer pain in adults[J].Cochrane Database Syst Rev,2011,19(1):CD007753.
[5]Lu W,Rosenthal DS.Acupuncture for cancer pain and related symptoms[J].Curr Pain Headache Rep,2013,17(3):321-328.
[6]ZHANG Y,LI A,LAO L,et al.Rostral ventromedial medulla mu,but not kappa,opioid receptors are involved in electroacupuncture anti-hyperalgesia in an inflammatory pain rat model[J].Brain Res,2011,1395:38-45.
[7]司马蕾,刘波涛,厉建春,等.电针对骨癌痛-吗啡耐受大鼠痛行为和降钙素基因相关肽的影响[J].中华行为医学与脑科学杂志,2013,22(5):388-390.
[8]Ducourneau VR,Dolique T,Hachem-Delaunay S,et al.Cancer pain is not necessarily correlated with spinal overexpression of reactive glia markers[J].Pain,2014,155:275-291.
[9]Medhurst SJ,Walker K,Bowes M,et al.A rat model of bone cancer pain[J].Pain,2002,96:129-140.
[10]华兴邦,周浩良.大鼠穴位图谱的研制[J].实验动物与动物实验,1991(1):1-5.
[11]Paxinos G,Watson C.The rat brain[M].Elsevier Inc:George Paxinos and Charles Watson,2007.
[12]Anand KJ,Willson DF,Berger J,et al.Tolerance and withdrawal from prolonged opioid use in critically ill children[J].Pediatrics,2010,125(5):1208-1225.
[13]唐淑洁,张佳琦,刘洋,等.洛芬待因与吗啡交替服用减少吗啡在癌痛患者中的耐药性[J].中国医学创新,2016,13(6):29-31.
[14]黄秋明,李俊.硫酸吗啡控释片直肠给药治疗晚期癌痛的Meta分析[J].药物流行病学杂志,2011,20(2):72-75.
[15]张亚军,田玉科,杨承祥,等.大鼠骨癌痛-吗啡耐受模型建立[J].中华麻醉学杂志,2011,31(1):63-66.
[16]梁宜,杜俊英,房军帆,等.常用骨癌痛动物模型的共性特点和特征分析[J].中华肿瘤杂志,2014,36(12):949-951.
[17]Hassanzadeh K,Khodadadi B,Moloudi MR,et al.A new pharmacological role for thalidomide:attenuation of morphine-induced tolerance in rats[J].Acta Anaesthesiol Taiwan,2016,54(2):65-69.
[18]司马蕾,厉建春,蔡淑呈,等.P物质和降钙素基因相关肽在骨癌痛-吗啡耐受模型中的表达[J].中国癌症杂志,2012,22(8):561-565.
[19]Mercadante S.Intravenous morphine for management of cancer pain[J].Lancet Oncol,2010,11(5):484-489.
[20]Paley CA,Bennett MI,Johnson MI.Acupuncture for cancer-induced bone pain?[J].Evid Based Complement Alternat Med,2011,2011:1043-1051.
[21]李娜,李为民,陈颖波,等.电针对完全弗氏佐剂性小鼠外周慢性炎症痛的缓解作用[J].中国针灸,2008,28(2):122-126.
[22]王珂,刘惠芬,周文华.“足三里”和“肾俞”穴位埋线对大鼠吗啡镇痛和运动行为敏化的影响[J].中国针灸,2008,28(7):509-513.
[23]杜俊英,房军帆,陈宜恬,等.电针抗大鼠骨癌痛的参数优选及其对阿片受体和前体mRNA表达的干预[J].中国针灸,2015,35(2):161-168.
[24]Hamza MA,White PF,Ahmed HE,et al.Effect of the frequency of transcutaneous electrical nerve stimulation on the postoperative opioid analgesic requirement and recovery profile[J].Anesthesiology,1999,91(5):1232-1238.
[25]郑宇欣,于泳浩,王国林.电针刺激对炎性痛大鼠吗啡耐受形成影响的研究[J].中国现代医学杂志,2009,19(9):1315-1318.
[26]Yamamoto J,Kawamata T,Niiyama Y,et al.Downregulation of mu opioid receptor within distinct subpopulations of dorsal root ganglion neurons ina murine model of bone cancer pain[J].Neuroscience,2008,151(3):843-853.
[27]BAO Y,GAO Y,HOU W,et al.Engagement of signaling pathways of protease-activated receptor 2andμ-opioid receptor in bone cancer pain and morphine tolerance[J].Int J Cancer,2015,137(6):1475-1483.
[28]Sim-Selley LJ,Selley DE,Vogt LJ,et al.Chronic heroin self-administration desensitizes mu opioid receptor-activated G-proteins in specific regionsofrat brain[J].Neurosci,2000,20(12):4555-4562.
[29]Campana G,Sarti D,Spampinato S,et al.Longterm intrathecal morphine and bupivacaine upregulate MOR gene in lymphocytes[J].Int Immunopharmacol,2010,10(9):1149-1152.
[30]JIANG Z,WU S,WU X,et al.Blocking mammalian target of rapamycin alleviates bone cancer pain and morphine tolerance viaμ-opioid receptor[J].Int J Cancer,2016,138(8):2013-2020.
[31]Zhang M,Wang K,Ma M,et al.Low-dose cannabinoid type 2receptor agonist attenuates tolerance to repeated morphine administration via regulatingμ-Opioid receptor expression in Walker 256tumor-bearing rats[J].Anesth Analg,2016,122(4):1031-1037.
[32]Dang VC,Christie MJ.Mechanisms of rapid opioid receptor desensitization resensitization and tolerance in brain neurons[J].Br J Pharmacol,2012,165(6):1704-1716.
[33]Lowe JD,Sanderson HS,Cooke AE,et al.Role of Gprotein-coupled receptor kinases 2and 3inμ-opioid receptor desensitization and internalization[J].Mol Pharmacol,2015,88(2):347-356.
[2]Connor M,Osborne PB,Christie MJ.Mu-opioid receptor desensitization:is morphine different?[J].Br J Pharmacol,2004,143(6):685-696.
[3]吕庆琴,陈霆隽,洪炎国.μ阿片受体的内吞抑制吗啡耐受的形成[J].中国临床药理学与治疗学,2012,17(10):1185-1190.
[4]Paley CA,Johnson MI,Tashani OA,et al.Acupuncture for cancer pain in adults[J].Cochrane Database Syst Rev,2011,19(1):CD007753.
[5]Lu W,Rosenthal DS.Acupuncture for cancer pain and related symptoms[J].Curr Pain Headache Rep,2013,17(3):321-328.
[6]ZHANG Y,LI A,LAO L,et al.Rostral ventromedial medulla mu,but not kappa,opioid receptors are involved in electroacupuncture anti-hyperalgesia in an inflammatory pain rat model[J].Brain Res,2011,1395:38-45.
[7]司马蕾,刘波涛,厉建春,等.电针对骨癌痛-吗啡耐受大鼠痛行为和降钙素基因相关肽的影响[J].中华行为医学与脑科学杂志,2013,22(5):388-390.
[8]Ducourneau VR,Dolique T,Hachem-Delaunay S,et al.Cancer pain is not necessarily correlated with spinal overexpression of reactive glia markers[J].Pain,2014,155:275-291.
[9]Medhurst SJ,Walker K,Bowes M,et al.A rat model of bone cancer pain[J].Pain,2002,96:129-140.
[10]华兴邦,周浩良.大鼠穴位图谱的研制[J].实验动物与动物实验,1991(1):1-5.
[11]Paxinos G,Watson C.The rat brain[M].Elsevier Inc:George Paxinos and Charles Watson,2007.
[12]Anand KJ,Willson DF,Berger J,et al.Tolerance and withdrawal from prolonged opioid use in critically ill children[J].Pediatrics,2010,125(5):1208-1225.
[13]唐淑洁,张佳琦,刘洋,等.洛芬待因与吗啡交替服用减少吗啡在癌痛患者中的耐药性[J].中国医学创新,2016,13(6):29-31.
[14]黄秋明,李俊.硫酸吗啡控释片直肠给药治疗晚期癌痛的Meta分析[J].药物流行病学杂志,2011,20(2):72-75.
[15]张亚军,田玉科,杨承祥,等.大鼠骨癌痛-吗啡耐受模型建立[J].中华麻醉学杂志,2011,31(1):63-66.
[16]梁宜,杜俊英,房军帆,等.常用骨癌痛动物模型的共性特点和特征分析[J].中华肿瘤杂志,2014,36(12):949-951.
[17]Hassanzadeh K,Khodadadi B,Moloudi MR,et al.A new pharmacological role for thalidomide:attenuation of morphine-induced tolerance in rats[J].Acta Anaesthesiol Taiwan,2016,54(2):65-69.
[18]司马蕾,厉建春,蔡淑呈,等.P物质和降钙素基因相关肽在骨癌痛-吗啡耐受模型中的表达[J].中国癌症杂志,2012,22(8):561-565.
[19]Mercadante S.Intravenous morphine for management of cancer pain[J].Lancet Oncol,2010,11(5):484-489.
[20]Paley CA,Bennett MI,Johnson MI.Acupuncture for cancer-induced bone pain?[J].Evid Based Complement Alternat Med,2011,2011:1043-1051.
[21]李娜,李为民,陈颖波,等.电针对完全弗氏佐剂性小鼠外周慢性炎症痛的缓解作用[J].中国针灸,2008,28(2):122-126.
[22]王珂,刘惠芬,周文华.“足三里”和“肾俞”穴位埋线对大鼠吗啡镇痛和运动行为敏化的影响[J].中国针灸,2008,28(7):509-513.
[23]杜俊英,房军帆,陈宜恬,等.电针抗大鼠骨癌痛的参数优选及其对阿片受体和前体mRNA表达的干预[J].中国针灸,2015,35(2):161-168.
[24]Hamza MA,White PF,Ahmed HE,et al.Effect of the frequency of transcutaneous electrical nerve stimulation on the postoperative opioid analgesic requirement and recovery profile[J].Anesthesiology,1999,91(5):1232-1238.
[25]郑宇欣,于泳浩,王国林.电针刺激对炎性痛大鼠吗啡耐受形成影响的研究[J].中国现代医学杂志,2009,19(9):1315-1318.
[26]Yamamoto J,Kawamata T,Niiyama Y,et al.Downregulation of mu opioid receptor within distinct subpopulations of dorsal root ganglion neurons ina murine model of bone cancer pain[J].Neuroscience,2008,151(3):843-853.
[27]BAO Y,GAO Y,HOU W,et al.Engagement of signaling pathways of protease-activated receptor 2andμ-opioid receptor in bone cancer pain and morphine tolerance[J].Int J Cancer,2015,137(6):1475-1483.
[28]Sim-Selley LJ,Selley DE,Vogt LJ,et al.Chronic heroin self-administration desensitizes mu opioid receptor-activated G-proteins in specific regionsofrat brain[J].Neurosci,2000,20(12):4555-4562.
[29]Campana G,Sarti D,Spampinato S,et al.Longterm intrathecal morphine and bupivacaine upregulate MOR gene in lymphocytes[J].Int Immunopharmacol,2010,10(9):1149-1152.
[30]JIANG Z,WU S,WU X,et al.Blocking mammalian target of rapamycin alleviates bone cancer pain and morphine tolerance viaμ-opioid receptor[J].Int J Cancer,2016,138(8):2013-2020.
[31]Zhang M,Wang K,Ma M,et al.Low-dose cannabinoid type 2receptor agonist attenuates tolerance to repeated morphine administration via regulatingμ-Opioid receptor expression in Walker 256tumor-bearing rats[J].Anesth Analg,2016,122(4):1031-1037.
[32]Dang VC,Christie MJ.Mechanisms of rapid opioid receptor desensitization resensitization and tolerance in brain neurons[J].Br J Pharmacol,2012,165(6):1704-1716.
[33]Lowe JD,Sanderson HS,Cooke AE,et al.Role of Gprotein-coupled receptor kinases 2and 3inμ-opioid receptor desensitization and internalization[J].Mol Pharmacol,2015,88(2):347-356.