基于AMPK/PGC-1α信号通路探讨通脉开窍丸对血管性认知障碍大鼠海马神经元线粒体生物发生的影响  

Effect of Tongmai Kaiqiao Pills on Mitochondrial Biogenesis of Hippocampal Neurons in Rats with Vascular Cognitive Impairment Based on AMPK/PGC-1αSignaling Pathway

作　　者：马璐瑶 李彦杰[2] 刘昊源 白艳杰[3] 邢若星[2] MA Luyao;LI Yanjie;LIU Haoyuan;BAI Yanjie;XING Ruoxing(School of Rehabilitation Medicine,Henan University of Chinese Medicine,Zhengzhou 450046,China;Henan Province Hospital of Traditional Chinese Medicine,Zhengzhou 450053,China;The First Affiliated Hospital of Henan University of Chinese Medicine,Zhengzhou 450003,China)

机构地区：[1]河南中医药大学康复医学院,郑州450046 [2]河南省中医院,郑州450053 [3]河南中医药大学第一附属医院,郑州450003

出　　处：《中国实验方剂学杂志》2025年第8期125-134,共10页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家中医药管理局“张仲景传承与创新专项”(GZY-KJS-2022-043-3);河南省自然科学基金项目(242300420111)。

摘　　要：目的:观察通脉开窍丸对血管性认知障碍(VCI)大鼠海马组织AMP活化蛋白激酶(AMPK)/过氧化物酶体增殖物激活受体γ辅激活子-1α(PGC-1α)信号通路及线粒体生物发生的影响,探究通脉开窍丸改善VCI大鼠认知障碍的潜在机制。方法:从72只雄性SD大鼠中选取12只用作假手术组,剩余大鼠采用改良2VO法造模,按随机数字表法将造模成功的大鼠分为模型组,通脉开窍丸高、低剂量组(27.6、13.8 g·kg^(-1)),联合组(通脉开窍丸27.6 g·kg^(-1)+Dorsomorphin 25 mg·kg^(-1))、盐酸多奈哌齐组(0.45 g·kg^(-1)),经过4周持续腹腔注射相应药物后,使用Morris水迷宫测试来检测大鼠的学习和记忆能力;苏木素-伊红(HE)染色及尼氏(Nissl)染色检测大鼠海马组织病理学变化;比色法检测大脑海马组织线粒体三磷酸腺苷(ATP)含量;线粒体超氧化物红色荧光探针法(MitoSOX Red)检测大鼠线粒体活性氧(ROS)水平;实时荧光定量聚合酶链式反应(Real-time PCR)检测线粒体DNA拷贝数;透射电镜(TEM)观察线粒体的病理变化;蛋白免疫印迹法(Western blot)检测大鼠海马AMPK、PGC-1α、磷酸化(p)-AMPK、核呼吸因子1(Nrf1)、线粒体转录因子A(TFAM)蛋白的表达量。结果:与假手术组比较,模型组大鼠穿越平台次数显著减少(P<0.01),逃避潜伏期显著延长(P<0.01),海马区神经元排列混乱,间隙增宽,核膜核仁界限模糊,可见坏死细胞出现,尼氏小体颜色浅且数量减少,严重丢失,线粒体萎缩,嵴溶解消失,损伤严重,ROS含量增加、ATP水平降低、mtDNA拷贝数显著下降(P<0.01),p-AMPK、PGC-1α、Nrf1、TFAM蛋白表达明显减少(P<0.05,P<0.01)。与模型组比较,通脉开窍丸高剂量组、盐酸多奈哌齐组大鼠找寻平台时间显著缩短(P<0.01),穿越平台次数显著增加(P<0.01),海马神经元线粒体形态结构有所恢复,神经元死亡情况缓解,尼氏小体数量增加,损伤程度减轻,ROS含量降低、ATP水平和mtDNA拷贝数明显增Objective:To observe the effects of Tongmai Kaiqiao pills on AMP-activated protein kinase(AMPK)/peroxisome proliferator-activated receptor gamma coactivator-1α(PGC-1α)signaling pathway and mitochondrial biogenesis in hippocampal tissue of rats with vascular cognitive impairment(VCI)and to investigate the potential mechanism of Tongmai Kaiqiao pills in improving cognitive impairment in rats with VCI.Methods:Twelve of 72 male SD rats were selected as the sham operation group,and the remaining rats were modelled using the modified 2VO method.The rats that were successfully modelled were divided into the model group,the high-dose group of Tongmai Kaiqiao pills(27.6 g·kg^(-1)),the low-dose group of Tongmai Kaiqiao pills(13.8 g·kg^(-1)),the combination group(27.6 g·kg^(-1) Tongmai Kaiqiao pills+25 mg·kg^(-1) dorsomorphin),and the donepezil hydrochloride group(0.45 g·kg^(-1))according to the random number table method.After four weeks of continuous intraperitoneal injection of the corresponding drugs,the Morris water maze test was used to test the learning and memory ability of rats.Hematoxylin-eosin(HE)staining and Nissl staining were used to detect pathological changes in the hippocampus of the rats.The content of mitochondrial adenosine triphosphate(ATP)in the brain hippocampus was detected by colorimetry,and reactive oxygen species(ROS)level was detected in rat mitochondria by MitoSOX Red assay.Mitochondrial DNA copy number was detected by real-time fluorescent quantitative PCR(Real-time PCR).Pathological changes in mitochondria were observed by transmission electron microscopy(TEM),and AMPK,PGC-1α,phosphorylated AMP-activated protein kinase(p-AMPK),nuclear respiratory factor 1(Nrf1),and mitochondrial transcription factor A(TFAM)protein expression in the hippocampus of the rats were detected by Western blot.Results:Compared with those in the sham operation group,rats in the model group had a reduced number of platform crossings(P<0.01),significantly prolonged evasion latency(P<0.01),disorganized neuronal arr

关 键 词：通脉开窍丸 血管性认知障碍 线粒体生物发生 AMP活化蛋白激酶(AMPK)/过氧化物酶体增殖物激活受体γ辅激活子-1α(PGC-1α)信号通路 

分 类 号：R338[医药卫生—人体生理学] R282[医药卫生—基础医学] R289