双氢青蒿素对肝癌HepG2细胞VEGF及HUVECs血管新生的影响  

Effects of Dihydroartemisinin on VEGF in Hepatoma HepG2 Cells and Angiogenesis inHUVECs

作　　者：刘妍君 刘付红 李笑[1] 刘炬 LIU Yanjun;LIU Fuhong;LI Xiao;LIU Ju(Shandong University of Traditional Chinese Medicine,Jinan 250355,China;First Affiliated Hospital of Shandong First Medical University,Jinan 250013,China)

机构地区：[1]山东中医药大学,山东济南250355 [2]山东第一医科大学第一附属医院,山东济南250013

出　　处：《山东中医药大学学报》2022年第4期524-532,548,共10页Journal of Shandong University of Traditional Chinese Medicine

基　　金：山东省中医药科技发展计划项目(编号:2019-0370)。

摘　　要：目的:探究双氢青蒿素(DHA)对肝癌HepG2细胞血管内皮生长因子(VEGF)的表达及DHA+肿瘤条件培养基(TCM)对内皮细胞血管生成的影响。方法:将乏氧条件培养的人肝癌细胞系HepG2分为空白组、不同浓度(10μmol/L、25μmol/L、50μmol/L和100μmol/L)DHA组,通过实时荧光定量聚合酶链反应(RT-qPCR)、蛋白质印迹法(Western blot)和酶联免疫吸附测定(ELISA)分别检测DHA对VEGF mRNA、蛋白和分泌水平及缺氧诱导因子-1α(HIF-1α)蛋白的影响。选取具有有效抑制作用的最低浓度25μmol/L DHA进行后续实验,将细胞分为4组:空白组、DHA+TCM组、PX-478+TCM组和PX-478+DHA+TCM组。收集各组培养肝癌HepG2细胞的TCM,各组TCM用于人脐静脉内皮细胞(HUVECs)共培养,用四甲基偶氮唑蓝(MTT)实验、划痕实验和成管实验分别检测DHA+TCM对HUVECs增殖、迁移和血管形成的影响,揭示DHA对VEGF作用的机制。结果:DHA呈剂量依赖性地抑制肝癌HepG2细胞VEGF mRNA、蛋白及分泌水平的表达;DHA+TCM可有效抑制HUVECs增殖、迁移及血管生成;此外,DHA对肝癌HepG2细胞中HIF-1α蛋白表达具有抑制作用。DHA+TCM对HUVECs血管生成的抑制作用可通过DHA与PX-478共处理而消除。结论:DHA可以抑制肝癌HepG2细胞VEGF表达及HUVECs血管生成,其机制可能是通过减少肝癌HepG2细胞转录因子HIF-1α的表达。Objective:To investigate the mechanism of dihydroartemisinin(DHA)on the expression of vascular endothelial growth factor(VEGF)in hepatoma HepG2 cells and the effect of DHA tumor conditioned medium(TCM)on endothelial angiogenesis.Methods:hepatoma HepG2 cells cultured under hypoxic conditions were divided into blank group,DHA groups with different concentrations(10μmol/L,25μmol/L,50μmol/L and 100μmol/L).The effects of DHA on VEGF mRNA,protein and secretion levels,and hypoxia-inducible factor-1α(HIF-1α)protein were detected by real-time quantitative polymerase chain reaction(RT-qPCR),Western blot and enzyme-linked immunosorbent assay(ELISA)respectively.The minimum concentration of DHA with effective inhibitory effect of 25μmol/L was selected for the follow-up test.The cells were divided into 4 groups:blank group,DHA+TCM group,PX-478+TCM group and PX-478+DHA+TCM group.TCM of hepatoma HepG2 cells cultured in each group was collected and used for co-culture of human umbilical vein endothelial cells(HUVECs).The effects of DHA+TCM on proliferation,migration and angiogenesis of HUVECs were detected by methyl thiazolyl tetrazolium(MTT)assay,scratch test and tube formation test,respectively,to reveal the mechanism of DHA on VEGF.Results:DHA inhibited the expression of VEGF mRNA,protein and secretion levels in hepatoma HepG2 cells in a dose-dependent manner.DHA+TCM can effectively inhibit the proliferation,migration and angiogenesis of HUVECs.In addition,DHA inhibited the expression of HIF-1αin hepatoma HepG2 cells.The inhibitory effect of DHA+TCM on angiogenesis of HUVECs could be eliminated by DHA and PX-478 co-treatment.Conclusions:DHA can inhibit VEGF expression and angiogenesis of HUVECs in hepatoma HepG2 cells,possibly by reducing the expression of transcription factor HIF-1αin hepatoma HepG2 cells.

关 键 词：双氢青蒿素 血管新生 肝癌HEPG2细胞 缺氧诱导因子-1Α 血管内皮生长因子 四甲基偶氮唑蓝实验 划痕实验 成管实验 

分 类 号：R285.5[医药卫生—中药学]