目的 利用网络药理学、分子对接技术和细胞实验验证，探讨藏红花素改善阿尔茨海默病（Alzheimer’s disease，AD）的作用机制。方法 运用Super-PRED、DisGenet、Genecards、Uniprot、STRING数据库构建藏红花素与AD的相互作用网络，导入Cytoscape软件根据连接程度筛选枢纽基因，并进行基因本体（gene ontology，GO）功能及京都基因与基因组百科全书（Kyoto encyclopedia of genes and genomes，KEGG）通路富集分析，应用Autodock vina软件进行分子对接验证。体外采用脂多糖（lipopolysaccharide，LPS）诱导BV2小胶质细胞炎症模型，给予藏红花素干预后，采用CCK-8法检测细胞活力，qRT-PCR检测肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）、白细胞介素-1β（interleukin-1β，IL-1β）、IL-6 mRNA表达，Western blotting检测磷脂酰肌醇3-激酶（phosphatidylinositol 3-kinase，PI3K）/蛋白激酶B（protein kinase B，Akt）/糖原合成酶激酶-3β（glycogen synthase kinase-3β，GSK-3β）通路相关蛋白表达。结果 网络药理学结果显示，藏红花素可能通过调控PI3K-Akt信号通路、缺氧诱导因子-1（hypoxia inducible factor-1，HIF-1）信号通路等，作用于热休克蛋白90AA1（heat shock protein 90 alpha family class A member 1，HSP90AA1）、核因子-κB（nuclear factor-κB，NF-κB）、Toll样受体4（Toll-like receptor 4，TLR4）、磷脂酰肌醇3-激酶调节亚基1（phosphatidylinositol 3-kinase regulatory subunit 1，PIK3R1）、磷脂酰肌醇3-激酶催化亚基α（phosphatidylinositol 3-kinase catalytic subunit α，PIK3CA）、E1A结合蛋白p300（E1A-binding protein p300，EP300）等靶点，发挥治疗AD的作用。分子对接结果显示藏红花素与PIK3R1靶点对接效果良好。细胞实验结果显示，藏红花素显著抑制LPS诱导的BV2细胞中炎症因子表达（P＜0.05、0.01），促进PI3K、Akt、GSK-3β的磷酸化（P＜0.05）。结论 藏红花素具有改善小胶质细胞极化、抗神经炎症的作用，其作用机制与激活PI3K/Akt/GSK-3β通路有关。

Objective To investigate the mechanism of crocin in improving Alzheimer’s disease (AD) by network pharmacology, molecular docking and cell experiments. Methods The interaction network between crocin and AD was constructed using Super PRED, DisGenet, Genecards, Uniprot and STRING databases. Hub genes were screened based on connectivity using Cytoscape software, and gene ontology (GO) function and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis were performed. Molecular docking validation was performed using Autodock vina software. In vitro, a BV2 microglial inflammation model was induced by lipopolysaccharide (LPS). After intervention with crocin, cell viability was detected by CCK-8 assay, mRNA expressions of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and IL-6 was detected by qRT-PCR, Western blotting was used to detect protein expressions related to the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/glycogen synthase kinase-3β (GSK-3β) pathway. Results The results of network pharmacology showed that crocin may act on heat shock protein 90AA1 (HSP90AA1), nuclear factor-κB (NF-κB), Toll-like receptor 4 (TLR4), phosphatidylinositol 3-kinase regulatory subunit 1 (PIK3R1), and phosphatidylinositol 3-kinase catalytic subunit α (PIK3CA) and E1A binding protein p300 (EP300), regulate PI3K-Akt signaling pathway, hypoxia inducible factor-1 (HIF-1) signaling pathway, thereby exert therapeutic effects on AD. The molecular docking results showed that crocin had a good docking effect with PIK3R1. The cell experiment results showed that crocin significantly inhibited the expressions of inflammatory factors in LPS-induced BV2 cells (P < 0.05, 0.01), and promoted the phosphorylation of PI3K, Akt and GSK-3β (P < 0.05). Conclusion Crocin has the effects of improving microglial polarization and anti-neuroinflammation, and its mechanism is related to the activation of PI3K/Akt/GSK-3β pathway.

R285.5

江苏省自然科学基金青年项目（BK20240740）；南京中医药大学养老服务与管理学院专项研究项目（2024YLFWYGL012）