基于细胞凋亡机制探讨黑逍遥散对阿尔茨海默病大鼠学习记忆能力的改善作用  

Hei Xiaoyaosan Improves Learning and Memory Abilities in Alzheimer's Disease Rats by Regulating Cell Apoptosis

作　　者：王虎平[1,2,3] 杨娇 陈怡琴 孟志鹏 吕育洁 胡韵韵 裴文丽 韩玉梅 WANG Huping;YANG Jiao;CHEN Yiqin;MENG Zhipeng;LYU Yujie;HU Yunyun;PEI Wenli;HAN Yumei(Gansu University of Chinese Medicine,Lanzhou 730000,China;Gansu Key Laboratory of TCM Excavation and Innovative Transformation,Lanzhou 730000,China;Gansu Engineering Laboratory for New Products of TCM,Lanzhou 730000,China)

机构地区：[1]甘肃中医药大学,兰州730000 [2]甘肃省中医方药挖掘与创新转化重点实验室,兰州730000 [3]甘肃省中药新产品创制工程实验室,兰州730000

出　　处：《中国实验方剂学杂志》2025年第9期108-115,共8页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金项目(82160862,81960828);甘肃省自然科学基金项目(22JR11RA113);第五批全国中医临床优秀人才研修项目(国中医药人教函〔2022〕239号);首批陇原青年英才项目(中共甘肃省委人才工作领导小组〔2022〕5号)。

摘　　要：目的:从磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt)/核转录因子-κB(NF-κB)信号通路介导的细胞凋亡机制探讨黑逍遥散改善阿尔茨海默病(AD)认知功能的影响。方法:体内(动物)实验将雄性4月龄SD大鼠随机分为空白组,假手术组,模型组,盐酸多奈哌齐组(0.45 mg·kg^(-1)),黑逍遥散高、中、低剂量组(15.30、7.65、3.82 g·kg^(-1)),每组10只。假手术组双侧海马注射生理盐水1μL,模型组、盐酸多奈哌齐组及黑逍遥散各剂量组均采用双侧海马注射β淀粉样蛋白1-42(Aβ_(1-42))溶液1μL制备AD模型。每天灌胃给药1次,连续42 d。Morris水迷宫实验检测大鼠学习记忆能力,苏木素-伊红(HE)染色观察大鼠海马区病理改变,酶联免疫吸附测定法(ELISA)检测胱天蛋白酶-3(Caspase-3)、B细胞淋巴瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)含量,蛋白免疫印迹法(Western blot)检测PI3K、Akt和NF-κB蛋白表达。体外(细胞)采用Aβ_(1-42)与PC12细胞共同培养建立AD的细胞模型,采用细胞增殖活性检测(CCK-8)试剂盒检测细胞活力,流式细胞术(FC)检测细胞凋亡。结果:动物实验表明,与空白组比较,模型组逃避潜伏期显著增加(P<0.01),穿越平台次数显著减少(P<0.01),海马细胞排列无序,神经元数目明显减少,Bax和Caspase-3蛋白表达显著升高,而Bcl-2蛋白表达量显著降低(P<0.01),p-PI3K/PI3K、p-Akt/Akt蛋白表达显著降低,p-NF-κB/NF-κB蛋白表达显著增加(P<0.01);与模型组比较,盐酸多奈哌齐组及黑逍遥散高、中剂量组逃避潜伏期缩短而穿越平台次数明显增加(P<0.05,P<0.01),盐酸多奈哌齐组及黑逍遥散剂量组大鼠海马区细胞排序较整齐,且神经元数目增加,盐酸多奈哌齐组及黑逍遥散给药组中Bax和Caspase-3蛋白表达明显降低,而Bcl-2蛋白表达明显升高(P<0.05,P<0.01),且其p-PI3K/PI3K、p-Akt/Akt的蛋白表达明显增加(P<0.05,P<0.01),p-NF-κB/NF-κB蛋白表达明显降低(P<0.05,P<0.01)。细胞实验表明,与空�Objective:To explore the mechanism of Hei Xiaoyaosan in improving the cognitive function in Alzheimer's disease(AD)from cell apoptosis mediated by the phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)/nuclear factor kappa B(NF-κB)signaling pathway.Methods:Four-month-old SD male rats were randomly assigned into a blank group,a sham group,a model group,a donepezil hydrochloride(0.45 mg·kg^(-1))group,and high-,medium-,and low-dose(15.30,7.65,and 3.82 g·kg^(-1),respectively)Hei Xiaoyaosan groups,with 10 rats in each group.The sham group received bilateral hippocampal injection of 1μL normal saline,while the other groups received bilateral hippocampal injection of 1μL beta-amyloid 1-42(Aβ_(1-42))solution for the modeling of AD.Rats were administrated with corresponding agents once a day for 42 consecutive days.The Morris water maze test was carried out to assess the learning and memory abilities of rats.Hematoxylin-eosin staining was employed to observe pathological changes in the hippocampus of rats.Enzyme-linked immunosorbent assay was employed to measure the levels of cysteinyl aspartate-specific proteinase-3(Caspase-3),B-cell lymphoma-2(Bcl-2),and Bcl-2-associated X protein(Bax).Western blot was employed to determine the protein levels of PI3K,Akt,and NF-κB.A cell model of AD was established by co-culturing Aβ_(1-42) and PC12 cells in vitro.Cell viability and apoptosis were detected by the cell-counting kit 8(CCK-8)assay and flow cytometry(FC),respectively.Results:Animal experiments showed that compared with the blank group,the model group had a prolonged escape latency(P<0.01),a reduced number of crossing platforms(P<0.01),disarrangement and a reduced number of hippocampal neurons,up-regulated expression of Bax and Caspase-3,down-regulated expression of Bcl-2(P<0.01),decreased p-PI3K/PI3K and p-Akt/Akt levels,and an increased p-NF-κB/NF-κB level(P<0.01).Compared with the model group,donepezil hydrochloride and high-and medium-dose Hei Xiaoyaosan shortened the escape latency and increased the num

关 键 词：阿尔茨海默病 黑逍遥散 细胞凋亡 磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt)/核转录因子-κB(NF-κB)信号通路 

分 类 号：R742[医药卫生—神经病学与精神病学] R285[医药卫生—临床医学] R259