中药新药与临床药理2026，Vol.37Issue(5)：782-796,15.DOI:10.19378/j.issn.1003-9783.2026.05.002
基于BMAL1介导抗氧化应激探讨葛根素减轻Aβ25-35诱导海马神经元凋亡的分子机制
Exploring the Molecular Mechanism of Puerarin Attenuating Aβ25-35-Induced Apoptosis in Hippocampal Neurons Based on BMAL1-Mediated Antioxidative Stress
摘要
Abstract
Objective To investigate the molecular mechanism by which puerarin alleviates Aβ25-35-induced apoptosis in hippocampal neurons,focusing on Brain and Muscle Arnt-Like-1(BMAL1)-mediated antioxidative stress.Methods(1)Network pharmacology analysis was employed to screen the potential targets of puerarin in ameliorating circadian rhythm disorder in Alzheimer's disease(AD).A protein-protein interaction(PPI)network was constructed,followed by GO functional enrichment and KEGG pathway analyses.Molecular docking,molecular dynamics simulation,and Drug Affinity Responsive Target Stability(DARTS)assays were conducted to validate the binding stability of puerarin with the core circadian regulator BMAL1 protein.(2)Primary mouse hippocampal neurons were extracted and identified via immunofluorescence,with purity assessed by flow cytometry.An AD cell model was established by treating primary hippocampal neurons with Aβ25-35 for 47 hours.Cell grouping and interventions were as follows:normal control group,AD model group,and low-,medium-,and high-dose puerarin groups(primary cells were pretreated with 12,24,and 48 µmol·L-1 puerarin for 1 hour,followed by Aβ25-35-induced injury for 47 hours).Western Blot was used to detect the expression levels of relevant proteins;cellular levels of glutathione(GSH),total superoxide dismutase(T-SOD),and catalase(CAT)were measured;mitochondrial membrane potential was assessed using the TMRE fluorescent probe;apoptosis rate and reactive oxygen species(ROS)levels were determined by flow cytometry.Results(1)A total of 86 potential targets of puerarin for AD-related circadian rhythm disorder were identified;20 core targets were closely associated with the occurrence and regulation of oxidative stress.The potential targets were involved in multiple signaling pathways related to circadian rhythm,oxidative stress,and neurodegeneration.Biological processes were primarily enriched in reactive oxygen species metabolic process,response to oxidative stress,circadian rhythm,and cognition;cellular components were mainly concentrated in the synaptic membrane,postsynaptic membrane,and presynaptic membrane;molecular functions were primarily enriched in ubiquitin-protein ligase binding,antioxidant activity,and protein phosphatase binding.Molecular simulation,molecular docking,and DARTS assays consistently demonstrated stable binding between puerarin and BMAL1 protein.(2)Compared with the normal control group,the AD model group exhibited significantly reduced BMAL1 protein expression(P＜0.01);significantly elevated ROS levels(P＜0.01);significantly decreased levels of GSH,T-SOD,CAT,and mitochondrial membrane potential(P＜0.01);and a significantly increased total apoptosis rate(P＜0.01),with Bax protein expression significantly upregulated(P＜0.01)and Bcl-2,PSD95,and SYN protein expression significantly downregulated(P＜0.01).Compared with the AD model group,the medium-and high-dose puerarin groups showed significantly increased BMAL1 protein expression(P＜0.01),significantly decreased ROS levels(P＜0.01),significantly elevated GSH,CAT levels,and mitochondrial membrane potential(P＜0.01),and a significantly reduced total apoptosis rate(P＜0.01).Bax expression was significantly downregulated(P＜0.01),while Bcl-2,PSD95,SYN expression were significantly upregulated(P＜0.01),NRF2 and HO-1 expression were significantly upregulated(P＜0.05,P＜0.01).T-SOD levels were significantly increased in the high-dose puerarin group(P＜0.01).Conclusion Puerarin attenuates Aβ25-35-induced apoptosis in hippocampal neurons by mediating antioxidative stress via BMAL1,and its mechanism may involve activation of the BMAL1-NRF2/HO-1 pathway.关键词
葛根素/阿尔茨海默病/昼夜节律/BMAL1/氧化应激/NRF2/HO-1通路/小鼠原代海马神经元Key words
puerarin/Alzheimer's disease/circadian rhythm/BMAL1/oxidative stress/NRF2/HO-1 pathway/mice primary hippocampal neurons分类
医药卫生引用本文复制引用
马国重,苗益恺,颜涛,孟云,刘玉莲,康桂琼,蒋晴,车峰远..基于BMAL1介导抗氧化应激探讨葛根素减轻Aβ25-35诱导海马神经元凋亡的分子机制[J].中药新药与临床药理,2026,37(5):782-796,15.基金项目
中国博士后科学基金面上项目(2023M732121) （2023M732121）
山东省自然科学基金青年项目(ZR2023QH090) （ZR2023QH090）
徐州医科大学附属医院发展基金优秀人才基金项目(XYFY202343). （XYFY202343）
