电针联合天麻素对阿尔茨海默病大鼠海马CA 1区沉默信息调节因子2同源蛋白1和过氧化物酶体增殖物激活受体γ辅激活子1 ɑ表达的影响
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摘要
目的:探讨电针联合天麻素治疗阿尔茨海默病(AD)的作用机制。方法:SD大鼠随机分为正常组、假手术组、模型组、电针组、天麻素组和针药联合组,每组10只。腹腔注射D-半乳糖联合双侧海马注射β淀粉样蛋白1-40制备AD大鼠模型。电针组和针药联合组给予"百会""大椎"、双侧"足三里"穴电针刺激,每次30min,1次/d,连续4周;天麻素组和针药联合组腹腔注射天麻素注射液,每日1次,连续4周。Morris水迷宫检测各组大鼠学习记忆能力;尼氏染色观察海马CA 1区神经元形态;免疫组织化学法检测各组大鼠海马CA 1区沉默信息调节因子2同源蛋白1(SIRT 1)和过氧化物酶体增殖物激活受体γ辅激活子(PGC-1ɑ)的表达。结果:Morris水迷宫结果显示,与正常组及假手术组比较,模型组大鼠逃避潜伏期延长(P<0.05),平台象限停留时间百分比、穿台次数降低(P<0.05);与模型组比较,电针与天麻素及联合使用均可降低AD大鼠的逃避潜伏期(P<0.05),提高平台象限停留时间百分比(P<0.05),增加穿台次数(P<0.05);针药联合组的效果优于电针组及天麻素组(P<0.05)。尼氏染色结果显示,与正常组及假手术组比较,模型组大鼠海马CA 1区神经元数量减少,排列紊乱;各治疗组CA 1区神经元数量较模型组明显增多,排列规则。与正常组及假手术组比较,模型组大鼠海马CA 1区SIRT 1和PGC-1ɑ阳性表达水平降低(P<0.05);与模型组比较,电针组和天麻素组CA 1区SIRT 1和PGC-1ɑ阳性表达水平升高(P<0.05);针药联合组的表达水平高于电针组和天麻素组(P<0.05)。结论:电针与天麻素均能够改善AD大鼠的学习记忆能力,且电针联合天麻素的效果更为显著,提示其可能通过上调海马SIRT 1和PGC-1ɑ蛋白的表达,发挥对AD大鼠神经元的保护作用。
Objective To observe the effect of electroacupuncture(EA)combined with Gastrodin on learning-memory ability and expression of silent information regulator 2 homologous protein 1(SIRT 1)and peroxisome proliferator activated receptorγcoactivator(PGC-1α)of hippocampal CA 1 region in Alzheimer's disease(AD)rats,so as to explore its mechanism underlying improvement of AD.Methods Sixty male SD rats were randomly divided into normal control(normal),sham operation(sham),model,EA,Gastrodin and EA+ Gastrodin groups(n=10 in each).The AD model was established by intraperitoneal injection of D-Galactose(120 mg·kg~(-1)·d~(-1))combined with bilateral hippocampal injection ofβamyloid 1-40(Aβ1-40).EA was applied at"Baihui"(GV 20),"Dazhui"(GV 14)and"Zusanli"(ST 36)for 30 min,once daily for 4 weeks.For rats of the Gastrodin group and EA+ Gastrodin group,intraperitoneal injection of gastrodin(10 mg/kg)was conducted once daily for 4 weeks.Morris water maze tests were used to assess the rat's learning-memory ability.Nissl staining was used to assess the morphological changes of neurons in the hippocampal CA 1 area.The expression of SIRT 1 and PGC-1αof hippocampal CA 1 region was measured by immunohistochemical staining.Results 1)Morris water maze tests showed that,compared with the normal and sham group,the escape latency was significantly prolonged(P<0.05),and the percentage of platform quadrant residence duration and the platform crossing times were considerably decreased in the model group(P<0.05).After the intervention,the escape latency was obviously shortened(P<0.05),and the percentage of platform quadrant residence duration and the platform crossing times were markedly increased in the EA,Gastrodin and EA+Gastrodin groups relevant to the model group(P<0.05).2)Nissl staining showed that,in comparison with the normal group or sham group,the number of cells in the hippocampal CA 1 area was decreased and the arrangement was disorganized in the model group.The number of cells in CA 1 area was relatively higher in the3 treatment groups than in the model group.3)The expression levels of SIRT 1 and PGC-1αproteins in the hippocampal CA 1 area were significantly down-regulated in the model group than in the normal and sham groups(P<0.05).After the intervention,the expression levels of SIRT 1 and PGC-1αin the EA,Gastrodin and EA+Gastrodin groups were significantly up-regulated compared with the model group(P<0.05).The effects of EA+Gastrodin were significantly superior to those of simple EA and simple Gastrodin in shortening the escape latency,up-regulating the expression levels of SIRT 1 and PGC-1αas well as in increasing the percentage of platform quadrant residence time and platform crossing times(P<0.05).Conclusion Both EA and Gastrodin can improve the learning-memory ability of AD rats,which may be related to their effects in up-regulating the expression of SIRT 1 and PGC-1αand reducing neuronal injury in the CA 1 region of hippocampus,suggesting aprotective role of EA on hippocampal neurons.The effect of EA combined with Gastrodin is markedly better than that of EA and Gastrodin alone.
Objective To observe the effect of electroacupuncture(EA)combined with Gastrodin on learning-memory ability and expression of silent information regulator 2 homologous protein 1(SIRT 1)and peroxisome proliferator activated receptorγcoactivator(PGC-1α)of hippocampal CA 1 region in Alzheimer's disease(AD)rats,so as to explore its mechanism underlying improvement of AD.Methods Sixty male SD rats were randomly divided into normal control(normal),sham operation(sham),model,EA,Gastrodin and EA+ Gastrodin groups(n=10 in each).The AD model was established by intraperitoneal injection of D-Galactose(120 mg·kg~(-1)·d~(-1))combined with bilateral hippocampal injection ofβamyloid 1-40(Aβ1-40).EA was applied at"Baihui"(GV 20),"Dazhui"(GV 14)and"Zusanli"(ST 36)for 30 min,once daily for 4 weeks.For rats of the Gastrodin group and EA+ Gastrodin group,intraperitoneal injection of gastrodin(10 mg/kg)was conducted once daily for 4 weeks.Morris water maze tests were used to assess the rat's learning-memory ability.Nissl staining was used to assess the morphological changes of neurons in the hippocampal CA 1 area.The expression of SIRT 1 and PGC-1αof hippocampal CA 1 region was measured by immunohistochemical staining.Results 1)Morris water maze tests showed that,compared with the normal and sham group,the escape latency was significantly prolonged(P<0.05),and the percentage of platform quadrant residence duration and the platform crossing times were considerably decreased in the model group(P<0.05).After the intervention,the escape latency was obviously shortened(P<0.05),and the percentage of platform quadrant residence duration and the platform crossing times were markedly increased in the EA,Gastrodin and EA+Gastrodin groups relevant to the model group(P<0.05).2)Nissl staining showed that,in comparison with the normal group or sham group,the number of cells in the hippocampal CA 1 area was decreased and the arrangement was disorganized in the model group.The number of cells in CA 1 area was relatively higher in the3 treatment groups than in the model group.3)The expression levels of SIRT 1 and PGC-1αproteins in the hippocampal CA 1 area were significantly down-regulated in the model group than in the normal and sham groups(P<0.05).After the intervention,the expression levels of SIRT 1 and PGC-1αin the EA,Gastrodin and EA+Gastrodin groups were significantly up-regulated compared with the model group(P<0.05).The effects of EA+Gastrodin were significantly superior to those of simple EA and simple Gastrodin in shortening the escape latency,up-regulating the expression levels of SIRT 1 and PGC-1αas well as in increasing the percentage of platform quadrant residence time and platform crossing times(P<0.05).Conclusion Both EA and Gastrodin can improve the learning-memory ability of AD rats,which may be related to their effects in up-regulating the expression of SIRT 1 and PGC-1αand reducing neuronal injury in the CA 1 region of hippocampus,suggesting aprotective role of EA on hippocampal neurons.The effect of EA combined with Gastrodin is markedly better than that of EA and Gastrodin alone.
引文
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[10]丁见,吴锋,缪化春,等.电针对脑缺血大鼠丘脑腹后外侧核神经生长因子和生长休止特定蛋白7表达的影响[J].针刺研究,2017,42(2):125-130.
[11]CHEN J H,KE K F,LU J H,et al.Protection of TGF-β1against neuroinflammation and neurodegeneration in Aβ1-42-induced Alzheimers disease model rats[J].PLoS One,2015,10(2):e 0116549.
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[13]黄金兰,秦嫦云,周楠,等.三七皂苷R 1对D-半乳糖诱导痴呆模型小鼠学习记忆能力的影响及抗氧化应激作用研究[J].中国药房,2015,26(31):4336-4338.
[14]VILLERS A,GIESE K P,RIS L.Long-term potentiation can be induced in the CA 1region of hippocampus in the absence ofαCaMKII T 286-autophosphorylation[J].Learn Mem,2014,21(11):616-626.
[15]MORRIS K C,LIN H W,THOMPSON J W,et al.Pathways for ischemic cytoprotection:role of sirtuins in caloric restriction,resveratrol,and ischemic preconditioning[J].J Cereb Blood Flow Metab,2011,31(4):1003-1019.
[16]SUGINO T,MARUYAMA M,TANNO M,et al.Protein deacetylase SIRT 1in the cytoplasm promotes nerve growth factor-induced neurite outgrowth in PC 12cells[J].FEBS Lett,2010,584(13):2821-2826.
[17]王晓,葛勤敏.去乙酰化酶SIRT 1对氧化应激的调控[J].现代生物医学进展,2015,15(30):5974-5976.
[18]ST-PIERRE J,DRORI S,ULDRY M,et al.Suppression of reactive oxygen species and neurodegeneration by the PGC-1transcriptional coactivators[J].Cell,2006,127(2):397-408.
[19]VENDITTI P,DI STEFANO L,DI MEO S.Mitochondrial metabolism of reactive oxygen species[J].Mitochondrion,2013,13(2):71-82.
[20]CHEN Z,SHENTU T P,WEN L,et al.Regulation of SIRT 1by oxidative stress-responsive miRNAs and a systematic approach to identify its role in the endothelium[J].Antioxid Redox Signal,2013,19(13):1522-1538.
[21]黄新格,零佩东,赵利华.D-半乳糖AD模型与中医脑衰老证候病机关系的探讨及在针灸防治脑衰老中的应用[J].世界中医药,2015,10(12):1983-1986.
[22]刘顺金,李娟娟.天麻素在脑损伤治疗中的研究进展[J].神经解剖学杂志,2016,32(6):781-784.
[23]HU Y,LI C,SHEN W.Gastrodin alleviates memory deficits and reduces neuropathology in a mouse model of Alzheimers disease[J].Neuropathology,2014,34(4):370-377.
[24]吴锋,李怀斌,赵健,等.电针结合天麻素对局灶性脑缺血大鼠神经功能、额叶皮质勿动蛋白A及其受体表达的影响[J].针刺研究,2016,41(1):65-69.
[25]崔晶晶,高俊虹,王玉敏,等.针药结合增效机制研究的新思路探讨[J].针刺研究,2010,35(2):146-150.
[2]TAN M,TANG C,ZHANG Y,et al.SIRT 1/PGC-1ɑsignaling protects hepatocytes against mitochondrial oxidative stress induced by bile acids[J].Free Radic Res,2015,49(8):935-945.
[3]姚婕,张红梅,阎丽萍,等.BNDF调控SIRT 1/PGC-1ɑ通路拮抗Aβ25~35诱导的神经元细胞氧化损伤[J].中国医药导报,2016,13(20):16-21.
[4]LEUNG A W,LAM L C,KWAN A K,et al.Electroacupuncture for older adults with mild cognitive impairment:study protocol for a randomized controlled trial[J].Trials,2015,16:232.
[5]李向宇,徐乐,柳垂亮,等.电针对异氟醚引起的APPswe/PS 1dE 9小鼠学习记忆能力及海马激活型Caspase-3、Bcl-2/Bax变化的影响[J].针刺研究,2016,41(1):24-30.
[6]MENG L,QIAN S.Gastrodin protects neural progenitor cells against amyloidβ(1-42)-induced neurotoxicity and improves hippocampal neurogenesis in amyloidβ(1-42)-injected mice[J].J Mol Neurosci,2016,60(1):21-32.
[7]高健美,陆国辉,李艳茹,等.天麻素通过SIRT 1/PGC-1ɑ信号通路抗叔丁基过氧化氢诱导的HL 7702细胞氧化损伤[J].中药药理与临床,2014,30(4):32-35.
[8]李令文,陈明苍,季宇彬.老年痴呆症模型研究进展[J].亚太传统医药,2014,10(5):43-45.
[9]华兴邦,李辞蓉,周浩良,等.大鼠穴位图谱的研制[J].实验动物与动物实验,1991(1):1-5.
[10]丁见,吴锋,缪化春,等.电针对脑缺血大鼠丘脑腹后外侧核神经生长因子和生长休止特定蛋白7表达的影响[J].针刺研究,2017,42(2):125-130.
[11]CHEN J H,KE K F,LU J H,et al.Protection of TGF-β1against neuroinflammation and neurodegeneration in Aβ1-42-induced Alzheimers disease model rats[J].PLoS One,2015,10(2):e 0116549.
[12]BENILOVA I,KARRAN E,DE STROOPER B.The toxic Aβoligomer and Alzheimer’s disease:an emperor in need of clothes[J].Nat Neurosci,2012,15(3):349-357.
[13]黄金兰,秦嫦云,周楠,等.三七皂苷R 1对D-半乳糖诱导痴呆模型小鼠学习记忆能力的影响及抗氧化应激作用研究[J].中国药房,2015,26(31):4336-4338.
[14]VILLERS A,GIESE K P,RIS L.Long-term potentiation can be induced in the CA 1region of hippocampus in the absence ofαCaMKII T 286-autophosphorylation[J].Learn Mem,2014,21(11):616-626.
[15]MORRIS K C,LIN H W,THOMPSON J W,et al.Pathways for ischemic cytoprotection:role of sirtuins in caloric restriction,resveratrol,and ischemic preconditioning[J].J Cereb Blood Flow Metab,2011,31(4):1003-1019.
[16]SUGINO T,MARUYAMA M,TANNO M,et al.Protein deacetylase SIRT 1in the cytoplasm promotes nerve growth factor-induced neurite outgrowth in PC 12cells[J].FEBS Lett,2010,584(13):2821-2826.
[17]王晓,葛勤敏.去乙酰化酶SIRT 1对氧化应激的调控[J].现代生物医学进展,2015,15(30):5974-5976.
[18]ST-PIERRE J,DRORI S,ULDRY M,et al.Suppression of reactive oxygen species and neurodegeneration by the PGC-1transcriptional coactivators[J].Cell,2006,127(2):397-408.
[19]VENDITTI P,DI STEFANO L,DI MEO S.Mitochondrial metabolism of reactive oxygen species[J].Mitochondrion,2013,13(2):71-82.
[20]CHEN Z,SHENTU T P,WEN L,et al.Regulation of SIRT 1by oxidative stress-responsive miRNAs and a systematic approach to identify its role in the endothelium[J].Antioxid Redox Signal,2013,19(13):1522-1538.
[21]黄新格,零佩东,赵利华.D-半乳糖AD模型与中医脑衰老证候病机关系的探讨及在针灸防治脑衰老中的应用[J].世界中医药,2015,10(12):1983-1986.
[22]刘顺金,李娟娟.天麻素在脑损伤治疗中的研究进展[J].神经解剖学杂志,2016,32(6):781-784.
[23]HU Y,LI C,SHEN W.Gastrodin alleviates memory deficits and reduces neuropathology in a mouse model of Alzheimers disease[J].Neuropathology,2014,34(4):370-377.
[24]吴锋,李怀斌,赵健,等.电针结合天麻素对局灶性脑缺血大鼠神经功能、额叶皮质勿动蛋白A及其受体表达的影响[J].针刺研究,2016,41(1):65-69.
[25]崔晶晶,高俊虹,王玉敏,等.针药结合增效机制研究的新思路探讨[J].针刺研究,2010,35(2):146-150.