藜芦胺对人胶质母细胞瘤细胞U251增殖的影响及机制  

Effects and mechanism of veratramine on the proliferation of human glioblastoma U251 cells

作　　者：曹梓珍 张琳 傅若秋 赵祎博 陈翔 陈剑鸿 CAO Zizhen;ZHANG Lin;FU Ruoqiu;ZHAO Yibo;CHEN Xiang;CHEN Jianhong(Dept.of Pharmacy,Army Medical Center of PLA,Chongqing 400042,China)

机构地区：[1]陆军特色医学中心药剂科,重庆400042

出　　处：《中国药房》2023年第22期2734-2739,共6页China Pharmacy

基　　金：重庆市研究生教育“课程思政”示范项目(No.YKCSZ23057);重庆市卫生计生委中医药科技项目(No.ZY201801004)。

摘　　要：目的 探究藜芦胺(VTM)对人胶质母细胞瘤(GBM)细胞U251增殖的影响及其潜在机制。方法 借助网络药理学方法,筛选VTM干预GBM的铁死亡相关交集靶点,并进行基因本体、京都基因和基因组百科全书富集分析。以U251细胞为对象,采用CCK-8法、细胞克隆形成实验、细胞形态变化观察、2′,7′-二氯荧光素二乙酸酯(DCFH-DA)荧光探针法、FerroOrange荧光探针法、Western blot实验对VTM抑制U251细胞增殖的作用及可能机制进行验证。结果 共筛选出VTM干预GBM的铁死亡相关交集靶点462个,富集于氧化应激、细胞凋亡等生物学过程,胞质囊泡、线粒体膜等细胞成分,影响二价铁离子(Fe^(2+))结合、DNA转录过程等分子功能,以及铁死亡、磷脂酰肌醇3激酶/蛋白激酶B信号通路等。40、60、80、100、120、140μmol/L的VTM均可显著降低细胞存活率(P<0.01);40、80、120μmol/L的VTM可使细胞萎缩、细胞核破碎,可显著抑制其克隆形成,显著提高其活性氧(ROS)、Fe^(2+)水平,并可不同程度地上调核转录因子红系2相关因子2(Nrf2)、血红素加氧酶1(HO-1)蛋白的表达,下调谷胱甘肽过氧化物酶4(GPX4)蛋白的表达(P<0.05或P<0.01)。结论 VTM可抑制U251细胞的增殖,促进细胞内ROS和Fe^(2+)的蓄积,从而诱导铁死亡,上述作用可能与调控Nrf2/HO-1/GPX4信号通路有关。OBJECTIVE To explore the effects and potential mechanism of veratramine(VTM)on the proliferation of human glioblastoma U251 cells.METHODS The network pharmacology methods were adopted to screen the targets of ferroptosis related to the effects of VTM on glioblastoma,and to conduct gene ontology and Kyoto Encyclopedia of Genes and Genosomes enrichment analysis.Using U251 cells as the object,CCK-8 assay,the observation of cell morphological changes,DCFH-DA fluorescence probe method,FerroOrange fluorescence probe method and Western blot assay were used to validate the inhibitory effects of VTM on U251 cell proliferation and its possible mechanism.RESULTS Totally 462 targets of ferroptosis related to the effects of VTM on glioblastoma were screened out;they mainly enriched in biological processes such as oxidative stress and apoptosis,and cellular components such as cytoplasmic vesicles and mitochondrial membranes;they affected molecular functions such as iron ion(Fe^(2+))binding and DNA transcription processes,as well as iron death and phosphoinositide 3-kinase/protein kinase B signaling pathways.VTM with 40,60,80,100,120 and 140μmol/L could significantly reduce the cell survival rate(P<0.01);VTM with 40,80 and 120μmol/L could cause cell atrophy and nuclear fragmentation,significantly inhibit the clone formation,increase the levels of intracellular reactive oxygen species(ROS)and Fe^(2+)levels,increase the expressions of nuclear factor-erythroid 2-related factor 2(Nrf2)and heme oxygenase 1(HO-1)protein to different extents,while down-regulate the expression of glutathione peroxidase 4(GPX4)protein(P<0.05 or P<0.01).CONCLUSIONS VTM can inhibit the proliferation of U251 cells,and promote the accumulation of intracellular ROS and Fe^(2+),thus inducing ferroptosis;its mechanism might be related to the regulation of the Nrf2/HO-1/GPX4 signaling pathway.

关 键 词：藜芦胺 胶质母细胞瘤 铁死亡 核转录因子红系2相关因子2/血红素加氧酶1/谷胱甘肽过氧化物酶4信号通路 

分 类 号：R965[医药卫生—药理学]