摘要
研究杜仲提取物对APP/PS1双转基因小鼠空间学习障碍的改善作用及探讨其与雌激素受体β(ERβ)/c-Jun氨基末端激酶(JNK)信号通路的关系。将60只3月龄雄性APP/PS1小鼠随机分为模型组、抗脑衰胶囊组(0.585 g·kg^(-1))、盐酸多奈哌齐组(0.65 mg·kg^(-1))、杜仲提取物组(1.3 g·kg^(-1));15只相同遗传背景的C57BL/6小鼠为野生(WT)对照组。通过水迷宫实验(MWM)、避暗实验(PAT)及新物体识别实验(NOR)评估小鼠学习记忆能力;苏木精-伊红(HE)染色及透射电镜(TEM)观察小鼠海马区组织形态和细胞超微结构特征;将关键活性成分与关键靶点利用AutoDock Vina软件进行分子对接验证;免疫组化方法(IHC)检测小鼠海马齿状回(DG)区中ERβ表达;蛋白免疫印迹法(WB)检测小鼠海马区ERβ、p-JNK、JNK的表达情况。与WT对照组相比,模型组小鼠在MWM中逃避潜伏期显著延长、穿越平台次数和目标象限停留时间显著减少,PAT中逃避潜伏期显著减少、错误次数显著增加,NOR中新物体识别指数显著降低;HE染色结果表明,模型组小鼠海马区出现神经元数目减少,锥体细胞排列紊乱,细胞核固缩等变化;TEM检测结果表明,模型组海马区部分神经元细胞核有固缩状态,核膜略微增厚和畸变,胞质内尼氏体较少;IHC结果表明,模型组小鼠海马DG区ERβ表达显著降低;WB结果表明,海马组织ERβ的表达降低,p-JNK/JNK水平升高。与模型组相比,杜仲提取物组小鼠在MWM中逃避潜伏期显著缩短、穿越平台次数和目标象限停留时间显著增加,PAT中逃避潜伏期显著增加、错误次数显著减少,NOR中新物体识别指数显著提高;小鼠海马区出现神经元数目增多,锥体细胞排列趋于有序;海马区神经元细胞核状态较好;小鼠海马DG区ERβ表达显著升高;海马组织ERβ的表达显著升高,p-JNK/JNK水平显著降低。盐酸多奈哌齐和抗脑衰胶囊对APP/PS1小鼠在行为学、HE、TEM实验指标中的作用与杜仲提取物作用相似,在IHC和WB实验中,盐酸多奈哌齐组与模型组没有明显差异。分子对接结果表明,杜仲提取物中雌激素类成分与ERβ结合紧密。综上所述,杜仲提取物与雌激素受体结合,调节ERβ的表达,激活ERβ/JNK信号通路可能是其改善APP/PS1小鼠学习记忆能力的关键机制之一。
To study the ameliorative effect of Eucommiae Cortex extract on spatial learning disabilities in APP/PS1 doubletransgenic mice and explore its relationship with estrogen receptorβ(ERβ)/c-Jun N-terminal kinase(JNK)signaling pathway,sixty 3-month-old male APP/PS1 mice were randomly divided into a model group,an anti-brain failure capsule group(0.585 g·kg^(-1)),a donepezil hydrochloride group(0.65 mg·kg^(-1)),and a Eucommiae Cortex extract group(1.3 g·kg^(-1)),and 15 C57BL/6 mice of the same genetic background were set as WT control group.The learning and memory ability of mice was assessed by the Morris water maze test(MWM),the passive avoidance test(PAT),and the novel object recognition test(NOR).The histomorphological and cellular ultrastructural features of the hippocampal region of the mice were observed by hematoxylin-eosin(HE)staining and transmission electron microscopy(TEM);the molecular docking validation of the key active ingredients and the key targets was performed by using AutoDock Vina software,and the immunohistochemical method(IHC)was used to detect the ERβexpression in the dentate gyrus(DG)area of mouse hippocampus.Western blot(WB)was utilized to detect the expression of ERβ,p-JNK,and JNK in mouse hippocampal area.Compared with those in the WT control group,the results of behavioral experiments showed that the latency of the mice in the model group was significantly increased,the number of platform traversals,and the target quadrant residence time were significantly decreased in the MWM.The evasion latency was significantly reduced,and the number of errors was significantly increased in the PAT.The index of recognition of novel objects was significantly reduced in the NOR.The results of HE staining indicated that the hippocampal area of mice in the model group showed a decrease in the number of neurons,disorganization of pyramidal cell arrangement,nucleus consolidation,and other changes.TEM results showed that some neuronal nuclei in the hippocampal area had a consolidated state,slightly thickened and aberrant nuclear membranes,and fewer intracytoplasmic nidus bodies;the IHC results showed that the expression of ERβin the hippocampal DG area of the mice was reduced.The WB results showed that the ERβexpression in the hippocampal tissue was decreased,and the p-JNK/JNK level was elevated.Compared with the model group,the Eucommiae Cortex extract group showed a significant decrease in latency,and increase in number of platform traversals and target quadrant residence time in the MWM,a significant increase in evasion latency and decrease in number of errors in the PAT,and a significant increase in the index of recognition of novel objects in the NOR.In addition,there was an increase in the number of neurons in the hippocampal area of mice.The pyramidal cells tended to be arranged in an orderly manner;the nuclei of neurons in the hippocampal area were in a better state;the expression of ERβin the hippocampal DG area of the mice was elevated;the expression of ERβin the hippocampal tissue was elevated,and the level of p-JNK/JNK was reduced.The effects of donepezil hydrochloride group and anti-brain failure capsule on APP/PS1 mice in terms of behavioral,HE,and TEM indexes were similar to those of Eucommiae Cortex extract,and there was no significant difference between donepezil hydrochloride group and the model group in IHC and WB experiments,and the results of molecular docking indicated that the estrogen-like components in Eucommiae Cortex extract were tightly bound to ERβ.In conclusion,the binding of Eucommiae Cortex extract to estrogen receptors,regulation of ERβexpression,and activation of ERβ/JNK signaling pathway may be one of the key mechanisms by which it improves the learning and memory ability of APP/PS1 mice.
作者
李月
张莉莉
赵灿
赵红梅
王堰
付金蕾
张杰
张宁
徐红丹
LI Yue;ZHANG Li-li;ZHAO Can;ZHAO Hong-mei;WANG Yan;FU Jin-lei;ZHANG Jie;ZHANG Ning;XU Hong-dan(Heilongjiang University of Chinese Medicine,Harbin 150040,China;Wuxi Higher Health Vocational Technology School,Wuxi 214028,China)
出处
《中国中药杂志》
北大核心
2025年第2期285-292,共8页
China Journal of Chinese Materia Medica
基金
国家自然科学基金面上项目(82174007,81673581)
中央支持地方高校改革发展资金人才培养项目
无锡市卫生健康“三名”战略人才项目——中青年医疗卫生拔尖人才培养计划项目(BJ2023094)
2022江苏高校“青蓝工程”优秀青年骨干教师培养项目。
