五脏温阳化瘀汤抑制周期蛋白依赖激酶-5过表达对SAMP8小鼠脑神经纤维缠结的影响
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摘要
目的:探讨五脏温阳化瘀汤改善SAMP8小鼠认知能力的周期蛋白依赖性激酶-5(CDK-5)相关脑神经纤维缠结机制。方法:10只SAMR1小鼠为正常组,40只SAMP8小鼠随机分为模型组,多奈哌齐组(0.4 mg·kg~(-1)·d~(-1)),五脏温阳化瘀汤高、低剂量组(5,1.25 g·kg~(-1)·d~(-1)),连续灌胃给药4周。Morris水迷宫测试定位航行与空间探索能力;实时荧光定量聚合酶链式反应(Real-time PCR)检测脑神经组织CDK-5 mRNA表达水平;蛋白免疫印迹法(Western blot)检测脑神经组织CDK-5蛋白表达与Tau蛋白磷酸化水平;银染法检测脑神经纤维缠结。结果:与正常组比较,模型组脑神经组织CDK-5与Tau蛋白磷酸化表达水平均显著升高(P<0.01),脑神经纤维缠结明显增多,定位航行与空间探索能力显著减弱(P<0.01);与模型组比较,五脏温阳化瘀汤高、低剂量组脑神经组织CDK-5与Tau蛋白磷酸化表达水平均明显降低(P<0.05,P<0.01),脑神经纤维缠结减少,小鼠定位航行并空间探索能力明显改善(P<0.05,P<0.01)。结论:五脏温阳化瘀汤可显著改善SAMP8小鼠认知能力,可能与其抑制CDK-5过表达进而抑制Tau蛋白过磷酸化导致脑神经纤维缠结减少有关。
Objective: To explore the mechanism of Wuzang Wenyang Huayu decoction in improving the cognitive competence and the pharmacological mechanism for neurofibrillary tangles related to cyclin-dependent kinase-5( CDK-5). Method: The 10 SAMR1 mice were used as normal group,40 SAMP8 mice were randomly divided into model group,donepezil group( 0. 4 mg·kg~(-1)·d~(-1)),high and low dose Wuzang Wenyang Huayu decoction groups( 5,1. 25 g·kg~(-1)·d~(-1)). Drugs were administered by gastric lavage for 4 continuous weeks.Directional navigation and space exploration ability were evaluated with Morris amaze. Real-time PCR was used to measure the mRNA expression of CDK-5 in brain nerve tissues. Western blot was used to detect the protein expression of CDK-5 and phosphorylation of Tau protein. Meanwhile,neurofibrillary tangles in brain tissue were detected with silver staining method. Result: As compared with normal group,both CDK-5 expression and Tau protein phosphorylation in brain nerve tissues were remarkably increased in model group( P < 0. 01),with significantly increased neurofibrillary tangles and significantly decreased directional navigation ability and space exploration ability( P < 0. 01). As compared with model group,CDK-5 expression and Tau protein phosphorylation were lower in Wuzang Wenyang Huayu decoction treated groups( P < 0. 05, P < 0. 01),accompanied by decreased number of neurofibrillary tangles as well as improved directional navigation and space exploration ability( P < 0. 05,P < 0. 01). Conclusion: Wuzang Wenyang Huayu decoction can markedly improve the cognitive competence of SAMP8 mice,and the mechanism may be related to its inhibition on CDK-5 over-expression,and down-regulation of Tau protein phosphorylation and neurofibrillary tangles in brain tissue.
Objective: To explore the mechanism of Wuzang Wenyang Huayu decoction in improving the cognitive competence and the pharmacological mechanism for neurofibrillary tangles related to cyclin-dependent kinase-5( CDK-5). Method: The 10 SAMR1 mice were used as normal group,40 SAMP8 mice were randomly divided into model group,donepezil group( 0. 4 mg·kg~(-1)·d~(-1)),high and low dose Wuzang Wenyang Huayu decoction groups( 5,1. 25 g·kg~(-1)·d~(-1)). Drugs were administered by gastric lavage for 4 continuous weeks.Directional navigation and space exploration ability were evaluated with Morris amaze. Real-time PCR was used to measure the mRNA expression of CDK-5 in brain nerve tissues. Western blot was used to detect the protein expression of CDK-5 and phosphorylation of Tau protein. Meanwhile,neurofibrillary tangles in brain tissue were detected with silver staining method. Result: As compared with normal group,both CDK-5 expression and Tau protein phosphorylation in brain nerve tissues were remarkably increased in model group( P < 0. 01),with significantly increased neurofibrillary tangles and significantly decreased directional navigation ability and space exploration ability( P < 0. 01). As compared with model group,CDK-5 expression and Tau protein phosphorylation were lower in Wuzang Wenyang Huayu decoction treated groups( P < 0. 05, P < 0. 01),accompanied by decreased number of neurofibrillary tangles as well as improved directional navigation and space exploration ability( P < 0. 05,P < 0. 01). Conclusion: Wuzang Wenyang Huayu decoction can markedly improve the cognitive competence of SAMP8 mice,and the mechanism may be related to its inhibition on CDK-5 over-expression,and down-regulation of Tau protein phosphorylation and neurofibrillary tangles in brain tissue.
引文
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[2]XIA C J,Dickerson B C.Tau PET:the next frontier in molecular imaging of dementia[J].Int Psychogeriatr,2016,28(9):1403-1406.
[3]SHEN X Y,LUO T,LI S,et al.Quercetin inhibits okadaic acid-induced tau protein hyperphosphorylation through the Ca2+calpain p25 CDK5 pathway in HT22cells[J].Int J Mol Med,2018,41(2):1138-1146.
[4]许小泰,谢炜.中医补肾法联合西药治疗老年性痴呆的Meta分析[J].中国实验方剂学杂志,2015,21(3):205-208.
[5]王若丹,李国春.健脑灵片联合复方银杏通脉口服液治疗血管性认知障碍45例[J].中国实验方剂学杂志,2015,21(19):181-184.
[6]王晋平,唐农,刘布谷,等.基于《内经》重阳思想的痴呆病病因病机及治疗思路探讨[J].时珍国医国药,2016,27(1):150-151.
[7]翟阳,唐农,黎军宏,等.五脏温阳化瘀胶囊治疗血管性痴呆的随机对照临床研究[J].辽宁中医杂志,2017,44(6):1212-1214.
[8]唐农,古联,严雁.五脏温阳化瘀汤治疗动脉硬化性血管性痴呆87例临床疗效观察[J].时珍国医国药,2015,26(6):1421-1422.
[9]陈炜,吴林,胡跃强,等.五脏温阳化瘀汤治疗肾虚血瘀型老年性痴呆30例临床研究[J].湖南中医杂志,2018,34(9):4-6,15.
[10]章元沛.药理学实验[M].2版.北京:人民卫生出版社,1996:238.
[11]孔小广.五脏温阳化瘀汤治疗动脉硬化性血管性痴呆的临床疗效观察[J].实用医技杂志,2018,25(6):659-660.
[12]张海燕,唐农,葛金文,等.五脏温阳化瘀汤对动脉粥样硬化血管性痴呆大鼠海马组织Nrf2蛋白通路的影响[J].云南中医学院学报,2013,36(3):8-11,20.
[13]ZHANG J H,YU L J,YANG H,et al.Huatuo Zaizao pill ameliorates cognitive impairment of APP/PS1 transgenic mice by improving synaptic plasticity and reducing Aβdeposition[J].BMC Complement Altern Med,2018,18(1):167.
[14]Das S,Basu S.Multi-targeting strategies for alzheimer's disease therapeutics:pros and cons[J].Curr Top Med Chem,2017,17(27):3017-3061.
[15]孙岩,马晨铭,刘锐,等.中西医治疗老年性痴呆研究述评[J].中医学报,2016,31(5):730-733.
[16]黄丽萍,熊玉洁,赵梦岚,等.覆盆子有效部位改善肾阳虚型痴呆大鼠学习记忆作用机制研究[J].中国实验方剂学杂志,2013,19(19):192-196.
[17]王飞,王民集.补肾活血汤加减结合五神针治疗肝肾不足型血管性痴呆症临床观察[J].中国实验方剂学杂志,2017,23(12):168-172.
[18]刘根,贺文彬,赵子强,等.基于中医传承辅助平台对老年性痴呆防治方剂核心药物组合的筛选研究[J].中国实验方剂学杂志,2016,22(7):223-228.
[19]易亚乔,方锐,葛金文,等.基于数据挖掘的历代医家痴呆相关病症用药规律分析[J].中国中药杂志,2018,43(16):3376-3381.
[20]宗鑫,季旭明,魏凤琴,等.基于中医传承辅助系统的治疗老年痴呆组方规律分析[J].中国中药杂志,2014,39(4):640-643.
[21]胡跃强,唐农,吴林,等.中医综合疗法联合西药治疗血管性痴呆103例临床观察[J].中医杂志,2014,55(23):2003-2006.
[22]杜娟,滕晶,王高峰.中医学对老年性痴呆的病机理论探讨[J].中华中医药学刊,2013,31(1):54-56.
[23]MA R H,ZHANG Y,HONG X Y,et al.Role of microtubule-associated protein tau phosphorylation in Alzheimer's disease[J].J Huazhong Univ Sci Technolog Med Sci,2017,37(3):307-312.
[24]Bakhtiari M,Panahi Y,Ameli J,et al.Protective effects of flavonoids against Alzheimer's disease-related neural dysfunctions[J].Biomed Pharmacother,2017,93(6):218-229.