目的 借助网络药理学、分子对接技术和细胞功能实验探究漏芦抗结肠癌的潜在作用机制。方法 通过中药系统药理学数据库与分析平台（TCMSP）、GeneCards数据库获取结肠癌靶点。将漏芦活性成分靶点与结肠癌靶点取交集获得共同作用靶点，借助微生信平台绘制交集靶点韦恩（Venn）图。将交集靶点导入STRING数据库构建蛋白质-蛋白质相互作用（PPI）网络，利用Cytoscape软件进行可视化及核心靶点筛选。通过DAVID数据库进行基因本体（GO）和京都基因与基因组百科全书（KEGG）富集分析。借助Autodock Vina、Pymol软件对活性成分与核心通路靶点进行分子对接及可视化。细胞增殖与活性检测（CCK-8）实验验证漏对结肠癌细胞的影响，对数据处理后计算出半数抑制浓度（IC₅₀）用于后续实验验证。通过平板克隆实验检测细胞增殖能力，明场拍照对细胞形态进行观察，蛋白免疫印迹法（Western blotting）验证核心靶点。结果共获得包括木犀草素、槲皮素、蜕皮甾酮等在内的10个活性成分，162个“漏芦-结肠癌”共同作用靶点；利用CytoNCA、MCODE插件对PPI网络筛选得到3个抗结肠癌核心靶点，分别是白细胞介素-1β（Il1B）、蛋白激酶B（AKT1）、肿瘤坏死因子（TNF）； GO富集分析主要包括对外源刺激的反应、基因表达的正调控、凋亡过程的负调控、炎症反应、DNA模板化转录的正调控等； KEGG富集分析得到乙型肝炎信号通路、脂质和动脉粥样硬化、化学致癌-受体激活、蛋白多糖在癌症、磷脂酰肌醇-3-激酶-蛋白激酶B（PI3K-Akt）、丝裂原活化蛋白激酶（MAPK）等信号通路；分析对接结果表明，IL1B、AKT1、TNF与对应小分子结合力均较稳定。细胞功能实验结果表明，木犀草素在HCT-15和HCT-116细胞中以剂量-时间相关性抑制细胞增殖、克隆，下调Bcl-2/Bax水平，上调凋亡蛋白clealved-PARP蛋白表达水平，下调p-Akt（Ser473）、p-ERK蛋白表达水平，而pan-Akt、pan-ERK蛋白表达量无明显变化。结论 木犀草素可以通过PI3K/Akt信号通路和MAPK/ERK信号通路抑制结肠癌细胞增殖水平并诱导细胞凋亡，为今后结肠癌的治疗提供了新的研究思路。

Objective To explore the potential mechanism of Rhapontici Radix’s anti-colon cancer effect by means of network pharmacology, molecular docking technology and cell function experiments. Method The main active components of Aconitum lucidum were collected through the Systematic Pharmacology Database and Analysis Platform for Traditional Chinese Medicine (TCMSP) and the HERB database. Colon cancer targets were obtained through the databases of Drugbank, TTD, OMIM and GeneCards. The intersection of the active component target of Rhapontici Radix and the target of colon cancer was taken to obtain the co-action target, and the Venn map of the intersection target was drawn with the help of the microbioinformatics platform. The intersection targets were imported into the STRING database to construct the protein-protein interaction (PPI) network, and the Cytoscape software was used for visualization and core target screening. Gene ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were conducted through the DAVID database. Molecular docking and visualization of the active ingredients and core pathway targets were carried out with the aid of Autodock Vina and Pymol software. The cell proliferation and activity assay (CCK-8) experiment verified the effect of leakage on colon cancer cells. After processing the data, the half maximal inhibitory concentration (IC50) was calculated for subsequent experimental verification. The cell proliferation ability was detected by plate cloning assay, the cell morphology was observed by bright field photography, and the core targets were verified by Western blotting. Results A total of 10 active components including luteolin, quercetin, ecdymolone, etc. were obtained, and 162 co-action targets of “Rhapontici Radix-colon cancer” were obtained. Three core anti-colon cancer targets were obtained by screening the PPI network using CytoNCA and MCODE plugins, namely Interleukin-1β (IL1B), threonine kinase 1 (AKT1) and tumor necrosis factor (TNF). GO enrichment analysis mainly includes responses to exogenous stimuli, positive regulation of gene expression, negative regulation of the apoptotic process, inflammatory responses, and positive regulation of DNA templating transcription, etc. KEGG enrichment analysis obtained signaling pathways such as the hepatitis B signaling pathway, lipids and atherosclerosis, chemical carcinogene-receptor activation, proteoglycans in cancer, PI3K-Akt signaling pathway, and MAPK signaling pathway; The analysis of the docking results indicated that the binding forces of IL1B, AKT1, TNF and the corresponding small molecules were all relatively stable. Cell experiments show. Conclusion Luteolin can inhibit the proliferation level of colon cancer cells and induce cell apoptosis through the PI3K/Akt signaling pathway and the MAPK/ERK signaling pathway, providing new research ideas for the treatment of colon cancer in the future.

R285.5

山东省中医药科技项目（M20241844）；山东中医药学会临床药学科研专项基金项目（SDACM202204）；山东省药学会医院药学科研专项项目（yyyx2021ms-04）