三焦针法对快速老化模型小鼠海马神经元细胞骨架重组相关蛋白的影响  

Effect of Sanjiao Acupuncture on Cytoskeleton Recombination Related Proteins in Hippocampal Neurons of Senescence Accelerated Mouse

作　　者：刘涛[1] 何佳[1] 张雪竹[2] 贾玉洁[2] 韩景献[1] 聂坤[2] LIU Tao;HE Jia;ZHANG Xue-zhu;JIA Yu-jie;HAN Jing-xian;NIE Kun(Department of Acupuncture,First Teaching Hospital of Tianjin University of Traditional Chinese Medicine,National Clinical Research Center for Chinese Medicine Acu puncture and Moxibustion,Tianjin(300193;Acupuncture Research Institute,First Teaching Hospital of Tianjin University of Traditional Chinese Medicine,Tianjin(300193))

机构地区：[1]天津中医药大学第一附属医院针灸部,国家中医针灸临床医学研究中心,天津300193 [2]天津中医药大学第一附属医院针灸研究所,天津300193

出　　处：《中国中西医结合杂志》2019年第12期1463-1468,共6页Chinese Journal of Integrated Traditional and Western Medicine

基　　金：国家自然科学基金面上项目(No.81574049)

摘　　要：目的观察三焦针法对快速老化模型小鼠(SAMP8)海马神经元细胞骨架重组相关蛋白的影响,探讨针刺治疗阿尔茨海默氏病(AD)的可能细胞生物学机制。方法以7月龄SAMP8为AD动物模型,随机分为对照组、针刺组和非穴组,每组各20只。针刺组给予三焦针法(取穴:膻中、中脘、气海、血海、足三里),非穴组以双侧胁肋下非经非穴点针刺处理,1次/日,连续6日,第7日休息,连续干预4周。采用Morris水迷宫隐蔽平台试验检测小鼠逃避潜伏期,采用空间探索实验检测小鼠原平台象限停留时间。提取小鼠海马组织脂筏,采用液质联用蛋白质组学方法检测各组小鼠海马脂筏蛋白,分析其中募集的细胞骨架相关蛋白的差异表达;采用免疫荧光共定位法检测各组β-tubulin及Cdc42表达;采用Western Blot法检测各组β-tubulin、β-actin、neurofibrillary蛋白表达。结果三组小鼠5天逃避潜伏期逐渐缩短(F=8.61,P=0.0048)。针刺组逃避潜伏期与对照组和非穴组均有显著差异(P<0.01)。与对照组和非穴组比较,针刺组第3天开始逃避潜伏期时间差异有统计学意义(P<0.01)。针刺组原平台象限停留时间[(19.14±4.32)s]明显高于对照组[(11.38±3.35)s]和非穴组[(13.51±2.46)s],差异有统计学意义(P<0.01)。与对照组比较,针刺组与非穴组海马脂筏募集的蛋白在种类和数量上存在差异。针刺组脂筏募集的细胞骨架相关蛋白和小G蛋白明显多于非穴组,且表现出同步增多的特点;免疫荧光共定位提示,针刺组β-tubulin与Cdc42在细胞核周以及轴突部位高度富集;与对照组比较,针刺对3种细胞骨架蛋白表达量的影响未显示统计学意义。结论三焦针法可调节海马神经元细胞骨架重组,这可能是其治疗AD的重要细胞生物学机制之一。Objective To observe the effect of Sanjiao acupuncture on the cytoskeleton recombination related proteins in hippocampus neurons of senescence accelerated mouse(SAMP8),and to explore the possible cellular biological mechanism of acupuncture in the treatment of Alzheimer’s disease(AD).Methods Seven-month-old SAMP8 was used as animal model of AD,and were randomly divided into control group,acupoint group and non-acupoint group,with 20 rats in each group.The acupoint group was treated with Sanjiao acupuncture [acupoints:Danzhong(CV17),Zhongwan(CV12),Qihai(CV6),Xuehai(ST36),Zusanli(SP10)].Non-acupoint group was treated with non-meridian and non-acupoint needling at bilateral subcostal.Both acupuncture groups were needled once daily,6 times a week for 4 weeks.Morris Water Maze was used for analyzing the effect of acupuncture on learning and memory ability of SAMP8 by hidden platform test and spatial probing test.The hippocampal tissue lipid rafts were extracted,the changes of cytoskeleton-related proteins in hippocampal lipid rafts recruitment of three groups were detected by HPLC MS/MS proteomics method;the intracellular co-localization of cytoskeleton and related proteins was examined by immunofluorescence staining method.Three cytoskeleton proteins(β-tubulin,3-actin,neurofibrillary)were determined by Western Blot.Results The 5-day escape latency of three groups was gradually shortened,the difference was statistically significant among groups(F=8.61,P=0.0048).The escape latency of acupuncture group was significantly different from that of control group and non-acupoint group(P <0.01).Compared with the control group and the non acupoint group,there was a statistically significant difference in the time of escape latency on the third day in the acupuncture group(P <0.01).The retention time of original platform quadrant in acupuncture group [(19.14±4.32)s]was significantly higher than that in control group [(11.38±3.35)s] and non acupoint group [(13.51 土2.46)s],the difference was statistically significant(P <0.0

关 键 词：阿尔茨海默氏病 快速老化模型小鼠 三焦针法 细胞骨架 小G蛋白 

分 类 号：R24[医药卫生—中医临床基础]