基于PI3K/Akt/mTOR通路探讨补肾通络方对血管性痴呆大鼠海马神经元突触可塑性的影响  

Effect of Bushen Tongluo Prescription on Synaptic Plasticity of Hippocampal Neurons of Vascular Dementia Model Rats Based on PI3K/Akt/mTOR Pathway

作　　者：张杰[1] 周媛[1] 马云枝[1] 赵敏[1] 代景妍 李思敏 ZHANG Jie;ZHOU Yuan;MA Yun-zhi;ZHAO Min;DAI Jing-yan;LI Si-min(The First Affiliated Hospital of Henan University of Chinese Medicine,Zhengzhou 450008,China;Henan University of Chinese Medicine,Zhengzhou 450046,China)

机构地区：[1]河南中医药大学第一附属医院,郑州450008 [2]河南中医药大学,郑州450046

出　　处：《中国实验方剂学杂志》2021年第18期25-31,共7页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：河南省中医药科学研究专项基金项目(2019ZY2142,2016ZY2050);国家自然科学基金项目(81904264)。

摘　　要：目的:探讨补肾通络方对血管性痴呆(VD)模型大鼠海马神经元突触可塑性的影响及其作用机制研究。方法:选用SPF级SD大鼠,采用双侧颈总动脉分次结扎法,建立VD大鼠模型。选取造模成功的大鼠,随机分为模型组,补肾通络方高、中、低剂量组,胰岛素样生长因子-1(IGF-1)组,并设假手术对照组,分别给予对应的药物灌胃,1次/d,连续4周;其中补肾通络方高、中、低剂量组的灌胃剂量分别为3,1.5,0.75 g·kg^(-1),IGF-1组灌胃剂量20μg·kg^(-1),模型组和假手术组给予同体积的生理盐水灌胃。末次灌胃结束后,采用Morris水迷宫法观察各组大鼠空间学习记忆能力,原位末端标记法(TUNEL)检测海马神经元细胞凋亡情况,高尔基染色法(Golgi)观察海马神经元突触形态及树突棘数量变化,蛋白免疫印迹法(Western blot)检测磷脂酰肌醇3-激酶(PI3K),蛋白激酶B(Akt),哺乳动物雷帕霉素靶蛋白(mTOR),突触素(SYP)及淀粉样前体蛋白(APP)蛋白的表达。结果:与假手术组比较,模型组大鼠逃避潜伏期、游泳路程明显延长,撤除平台后跨越原平台次数明显减少(P<0.05),细胞凋亡数目明显增多(P<0.05);树突棘数量明显减少(P<0.05);PI3K,Akt,mTOR,SYP蛋白表达明显减少,APP表达明显增多(P<0.05);与模型组比较,补肾通络方组,IGF-1组逃避潜伏期、游泳路程明显缩短,撤除平台后跨越原平台次数明显增多(P<0.05),细胞凋亡数目明显减少(P<0.05);PI3K,Akt,mTOR,SYP蛋白表达明显增多,APP表达明显减少(P<0.05);与IGF-1组比较,在大鼠逃避潜伏期、游泳路程、撤除平台后跨越原平台次数,细胞凋亡数目,树突棘数量,PI3K,Akt,mTOR,SYP,APP蛋白表达等,补肾通络方高剂量组差异均无统计学意义;补肾通络方中、低剂量组差异均有统计学意义(P<0.05)。结论:补肾通络方能够改善VD大鼠学习记忆能力,其机制可能与激活PI3K/Akt/mTOR自噬通路,改善海马神经元细胞突触可塑性有关。Objective: To explore the effect of Bushen Tongluo prescription(BSTLP)on the synaptic plasticity of hippocampal neurons in vascular dementia(VD)model rats and its mechanism. Method: SD male rats of SPF grade were selected. The rat model of VD was established by permanent bilateral ligation of the common carotid artery several times. The model rats were randomly divided into a model group,an insulin-like growth factor-1(IGF-1,20 μg·kg^(-1))group,high-dose(3 g·kg^(-1)),medium-dose(1.5 g·kg^(-1)),and low-dose(0.75 g·kg^(-1))BSTLP groups. A sham operation group was also set. Drugs were administered to rats by gavage once a day for four weeks. The model group and the sham operation group received the same volume of normal saline. After the last administration,all the rats were detected for spatial learning and memory by the Morris water maze. The apoptosis of hippocampal neurons was detected by terminal deoxynucleotidyl transferase(TdT)dUTP nick-end labeling(TUNEL)assay. The changes in synaptic morphological structure and the number of dendritic spines in hippocampal neurons were detected by Golgi’s method. The expression levels of phosphatidylinositol 3-kinase(PI3 K),protein kinase B(Akt),mammalian target of rapamycin(mTOR),synaptophysin(SYP),and amyloid precursor protein(APP)in hippocampal neurons were detected by Western blot. Result: Compared with the sham operation group,the model group showed prolonged escape latency,lengthened swimming distance,dwindled the number of times for the platform crossing after platform removal(P<0.05),increased apoptotic cells(P<0.05),declining synaptic dendritic spines(P<0.05),down-regulated expression levels of PI3 K,Akt,mTOR,and SYP proteins,and up-regulated expression level of APP protein in hippocampal neurons(P<0.05). Compared with the model group,the BSTLP groups and the IGF-1 group showed shortened escape latency and swimming distance,increased number of times for the platform crossing after platform removal(P<0.05),declining apoptotic cells(P<0.05),up-regulated expres

关 键 词：血管性痴呆 补肾通络方 磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt)/哺乳动物雷帕霉素靶蛋白(mTOR)信号通路 突触可塑性 

分 类 号：R2-0[医药卫生—中医学] R22