目的 采用生物信息学与实验验证探究重楼皂苷 I 调控上皮-间充质转化 （EMT） 抑制胶质母细胞瘤（GBM） 侵袭、迁移的作用机制。方法 通过肿瘤基因组图谱 （TCGA） 数据库筛选胶质母细胞瘤的差异表达基因（DEGs） 并进行富集分析，探究其发病相关的作用机制。将预测得到的重楼皂苷 I 靶点与疾病的 DEGs 取交集，构建蛋白互作网络，筛选药物发挥治疗作用的关键靶点进行富集分析并通过分子对接进行验证，以明确重楼皂苷 I 治疗胶质母细胞瘤的潜在分子机制。基于以上生物信息学结果，进行体外实验验证。以不同浓度的重楼皂苷 I 分别干预人胶质母细胞瘤细胞系 LN229 和 U251 后，细胞增殖与细胞毒性检测法 （CCK-8） 检测细胞活性；Transwell实验检测细胞侵袭能力；划痕实验检测细胞迁移能力；Immunoblotting法检测转化生长因子β1 （TGFβ1） 、钙黏蛋白 1 （CDH1） 、钙黏蛋白 2 （CDH2） 、波形蛋白 （VIM） 的蛋白表达水平。通过基因表达谱交互分析数据库（GEPIA） 分析胶质母细胞瘤与正常组织中 CDH2、TGFβ1、VIM 的表达差异，预测其对患者生存情况的影响。结果 基于生物信息学蛋白互作分析，筛选出 CDH1、CDH2 和 TGFβ1 等 7 个关键靶点，富集分析发现与EMT 生物过程及 TGFβ 信号通路关系密切。不同浓度的重楼皂苷 I 分别干预 LN229 与 U251 后细胞活性明显降低 （P＜0.05，P＜0.01） 。与对照组比较，重楼皂苷 I 可明显抑制 LN229 和 U251 细胞的侵袭 （P＜0.01） 及迁移能力 （P＜0.05，P＜0.01） ，下调 TGFβ1、CDH2、VIM 蛋白并上调 CDH1 蛋白表达水平 （P＜0.05，P＜0.01） ，呈剂量依赖效应 （P＜0.01） 。GEPIA 分析发现胶质母细胞瘤组织中 CDH2、TGFβ1 和 VIM 表达水平明显高于正常组织 （P＜0.05） ，其中 TGFβ1 和 VIM 的高表达可能是影响患者无病生存期的关键因素 （P＜0.05） 。结论 重楼皂苷 I 可通过 TGFβ1 介导的 EMT 进程，抑制 LN229 与 U251 细胞的侵袭与迁移能力，为临床精准治疗和中药小分子开发提供了新的研究方向与证据支持。

Objective Bioinformatics and experimental validation were used to explore the mechanism of action of polyphyllin Ⅰ on regulating epithelial-mesenchymal transition （EMT） to inhibit the invasion and migration of glioblastoma （GBM）. Methods Firstly，differentially expressed genes （DEGs） of glioblastoma were screened by The Cancer Genome Atlas Program（TCGA）database，and then they were used for enrichment analysis to explore the pathogenesis-related mechanism. Subsequently，the predicted targets of polyphyllinⅠwere taken to intersect with the DEGs of the disease to construct a protein interaction network. The key targets for the therapeutic effects of the drug were screened and used for enrichment analysis. Furthermore，molecular docking was applied for validation to clarify the potential molecular mechanism of polyphyllinⅠfor the treatment of glioblastoma，which was then validated by in vitro experiments based on the above bioinformatics results. After the intervention of concentration-gradient polyphyllinⅠagainst human glioblastoma cell lines LN229 and U251，cell activity was detected by Cell Counting Kit-8（CCK-8），invasive ability of glioblastoma cells was detected by Transwell assay，and migratory ability of glioblastoma cells was detected by scratch assay. Immunoblotting assay was performed to detect protein expression of transforming growth factor β1 （TGFβ1）， cadherin 1 （CDH1）， cadherin 2 （CDH2） and vimentin （VIM）. Finally， differences in the expression of CDH2，TGFβ1，and VIM in glioblastoma and normal tissues were analyzed by Gene Expression Profiling Interaction Analysis database （GEPIA） to predict their effects on patient survival. Results Seven key targets，including CDH1， CDH2 and TGFβ1， were screened based on bioinformatics analysis， and enrichment analysis revealed that the above targets were closely related to the EMT biological process and TGF β signaling pathway. The cell viability was significantly reduced after the intervention of concentration-gradient polyphyllinⅠagainst LN229 and U251（P＜0.05，P＜0.01）. Compared with the control group，polyphyllinⅠsignificantly inhibited the invasion（P＜0.01）and migration ability （P＜0.05，P＜0.01） of LN229 and U251 cells， up-regulated the protein expression of CDH1 and down-regulated the protein expression of TGFβ1，CDH2 and VIM （P＜0.05，P＜0.01） in a dose-dependent effect （P＜0.01）. GEPIA analysis revealed that the expression of CDH2，TGFβ1 and VIM in glioblastoma tissues were significantly higher than those in normal tissues（P＜0.05） ，and the high expression of TGFβ1 and VIM might be the key factors affecting the disease-free survival of patients （P＜0.05）. Conclusion PolyphyllinⅠcan inhibit the invasion and migration ability of LN229 and U251 cells through TGFβ1-mediated EMT process. This study provides a new research direction and evidence support for the clinical precision treatment and the development of traditional Chinese medicine small molecules.

R285.5

山东省医药卫生科技发展计划项目 （202004090876）；山东省医药卫生科技面上项目 （202304040722）