目的 通过网络药理学的预测技术和实验验证，研究木犀草素治疗脑缺血再灌注损伤（CIRI）的作用机制。方法 以“木犀草素”“脑缺血再灌注损伤”为关键词，于中药系统药理学数据库与分析平台（TCMSP）2.3数据库、GeneCards 5.19数据库检索靶点，利用Venny 2.1.0工具得到交集靶点，将交集靶点导入STRING数据库进行PPI网络构建，应用Cytoscape3.10.1软件对蛋白质-蛋白质相互作用（PPI）网络给予可视化分析，根据网络拓扑学参数挑选出关键靶点，利用Metascape数据库对交集靶点进行基因本体（GO）和京都基因与基因组百科全书（KEGG）通路富集分析。利用分子对接技术，验证木犀草素对关键靶蛋白的结合能力。SPF级雄性大鼠设置假手术组、模型组和木犀草素低、中、高剂量(30、50、70 mg·kg^-1)组，除假手术组外，均制备CIRI模型，造模后，每天18∶00时ig给药，假手术和模型组ig同等剂量0.9%氯化钠溶液，连续治疗7 d。对大鼠进行神经功能评分，利用苏木精-伊红（HE）染色观察病理改变，采用免疫组化和Western blotting检测Toll样受体4（TLR4）、核因子（NF）-κB p65和白细胞介素（IL）-6的蛋白表达。结果 网络药理学分析预测木犀草素调控IL6、NF-κB、前列腺素内过氧化物合酶2（PTGS2）等核心靶点，且结合能力较好，主要的作用途径包括NF-κB、磷脂酰肌醇3-激酶（PI3K）-蛋白激酶B（Akt）等。在体实验的结果表明，模型组神经功能评分明显高于假手术组（P<0.05），与模型组相比，木犀草素中、高剂量组的神经功能评分显著下降（P<0.05）；与假手术组相比，模型组大鼠海马区神经细胞离散、肿胀及细胞核固缩，给予木犀草素中、高剂量治疗后海马区的神经细胞排列相对紧密、细胞形态相对完整；与假手术组相比，模型组大鼠脑组织中TLR4、NF-κB p65和IL-6水平均明显升高（P<0.05），木犀草素中、高剂量组与模型组比较，能明显降低脑组织中TLR4、NF-κB p65、IL-6水平（P<0.05）。结论 木犀草素可改善CIRI引起的神经损伤，降低神经功能评分，其作用机制可能与抑制TLR4/NF-κB/IL6通路相关。

Objective The mechanism of luteolin in treating cerebral ischemia-reperfusion injury（CIRI） was investigated through network pharmacology prediction techniques and experimental verification. Methods The keywords “luteolin” and “cerebral ischemia-reperfusion injury” were used to search for targets in the TCMSP 2.3 and GeneCards 5.19 databases. The intersection targets were obtained using the Venny 2.1.0 tool and imported into the STRING database for PPI network construction. The PPI network was visualized and analyzed using Cytoscape 3.10.1 software, and key targets were selected based on network topology parameters. The Metascape database was used to perform gene ontology（GO） biological function and Kyoto Encyclopedia of Genes and Genomes（KEGG） pathway enrichment analysis on the intersection targets. Molecular docking technology was used to verify the binding ability of luteolin to key target proteins. SPF-grade male rats were divided into sham operation group, model group, and low-, medium-, and high-dose luteolin groups(30, 50, and 70 mg·kg^-1). Except for the sham operation group, CIRI models were prepared in the other groups. After modeling, the rats were ig administered at 18:00 every day. The sham operation and model groups were ig administered the same dose of 0.9% sodium chloride solution for seven consecutive days. The neurological function scores of the rats were evaluated, and pathological changes were observed by HE staining. The protein expressions of Toll-like receptor 4（TLR4）, nuclear factor（NF）-κB p65, and interleukin（IL）-6 were detected by immunohistochemistry and Western blotting. Results Network pharmacology analysis predicted that luteolin regulated core targets such as IL6, NF-κB, and prostaglandin-endoperoxide synthase 2（PTGS2）, and had good binding ability. The main action pathways included NF-κB, phosphatidylinositol 3-kinase（PI3K）-protein kinase B（AKT）, etc. The results of in vivo experiments showed that the neurological function score of the model group was significantly higher than that of the sham operation group（P < 0.05）. Compared with the model group, the neurological function scores of the medium-and high-dose luteolin groups were significantly decreased（P < 0.05）. Compared with the sham operation group, the neurons in the hippocampus of the model group rats were dispersed, swollen, and the nuclei were condensed. After treatment with medium-and highdose luteolin, the neurons in the hippocampus were relatively closely arranged and the cell morphology was relatively intact. Compared with the sham operation group, the levels of TLR4, NF-κB p65, and IL-6 in the brain tissue of the model group rats were significantly increased（P < 0.05）. Compared with the model group, the levels of TLR4, NF-κB p65, and IL-6 in the brain tissue of the medium-and high-dose luteolin groups were significantly decreased（P < 0.05）. Conclusion Luteolin can improve the neurological damage caused by CIRI and reduce the neurological function score. Its mechanism of action may be related to the inhibition of the TLR4/NF-κB/IL6 pathway.

R285.5

国家自然科学基金资助项目(82360873); 贵州中医药大学大学生创新创业项目(S2024106621492);贵州中医药大学研究生教育创新计划项目(YCXKYB2023017); 贵州省科技计划项目(黔科合基础-ZK[2022]一般471)