摘要
目的探讨毛蕊花糖苷(VB)对胶质母细胞瘤细胞上皮间质转化的作用及分子机制。方法(1)实验1:常规培养胶质母细胞瘤T98和U251细胞并各自分为4组,依次添加0、20、40、80μmol/L的VB处理24 h。采用CCK-8实验检测4组细胞存活率;采用Transwell实验检测4组细胞迁移、侵袭情况;采用RT-PCR实验及Western blotting实验检测4组细胞转化生长因子-β(TGF-β)、波形蛋白、Snail蛋白mRNA及蛋白表达情况。(2)实验2:T98、U251细胞各分为4组,依次为对照组、VB组、TGF-β组和TGF-β+VB组。对照组及VB组细胞转染空载质粒、TGF-β组及TGF-β+VB组细胞转染TGF-β质粒;转染24 h后VB组、TGF-β+VB组细胞均加入40μmol/L VB继续培养24 h,随后进行后续实验(迁移和侵袭实验、Western blotting实验,实验方法及检测指标同实验1)。(3)体内实验:裸鼠皮下种植U251细胞后分为2组,每组8只。实验组每天腹腔注射100 mg/kg的VB,对照组注射等量的PBS。记录小鼠肿瘤体积大小和小鼠体质量的变化;实验终点(移植后21 d)时,检测小鼠肿瘤重量并采用Western blotting实验检测肿瘤中TGF-β、波形蛋白、Snail的表达。结果(1)实验1:与0μmol/L VB组相比较,20μmol/L、40μmol/L VB组的细胞存活率差异无统计学意义(P>0.05),而80μmol/L VB组细胞存活率下降,差异有统计学意义(P<0.05);与0μmol/L VB组比较,20和40μmol/L VB组迁移和侵袭到下室的细胞随浓度的增高而减少,TGF-β、波形蛋白、Snail蛋白的mRNA和蛋白表达水平也依次降低,差异有统计学意义(P<0.05)。(2)实验2:与对照组相比较,VB组细胞迁移和侵袭的细胞数量减少,TGF-β、波形蛋白及Snail蛋白的表达下调,差异有统计学意义(P<0.05);而TGF-β组细胞迁移和侵袭的细胞数量增加,TGF-β、波形蛋白及Snail蛋白的表达上调,差异有统计学意义(P<0.05);与VB组比较,TGF-β组及TGF-β+VB组细胞迁移和侵袭的细胞数量增加,TGF-β、波形蛋白及Snail蛋白的�
Objective To investigate the effect of verbascoside(VB)on epithelial-mesenchymal transformation and its potential molecular mechanism in glioblastoma cells.Methods(1)Glioblastoma cell lines T98 and U251 were routinely cultured in vitro.Cells were divided into 4 groups and treated with 0μmol/L(controls),20μmol/L,40μmol/L,and 80μmol/L VB for 24 h,respectively;CCK-8 was used to detect the cell viability;Transwell assay was used to evaluate the migration and invasion abilities;reverse transcription(RT)-PCR and Western blotting were conducted to examine the mRNA and protein expressions of transforming growth factor(TGF)-β,vimentin and Snail,respectively.(2)T98 and U251 cells were divided into 4 groups:control group(cells being transfected with empty plasmid),VB group(cells being transfected with empty plasmid and treated by 40μmol/L VB),TGF-βgroup(cells being transfected with TGF-βoverexpression plasmid),TGF-β+VB group(cells being transfected with TGF-βoverexpression plasmid and treated by 40μmol/L VB);CCK-8 was used to detect the cell viability;Transwell assay was used to evaluate the migration and invasion abilities;RT-PCR and Western blotting were conducted to examine the mRNA and protein expressions of TGF-β,vimentin and Snail,respectively.(3)U251 cells were implanted subcutaneously into the nude mice;the mice were divided into VB group and control group(n=8).The mice in the VB group were intraperitoneally injected with 100 mg/kg/d VB,and mice in the control group were injected with the same volume of PBS.Tumor size and mice body mass were recorded.At the end of the experiment,mice were sacrificed;tumors were removed and weighed.The protein expressions of TGF-β,vimentin and Snail in tumor tissues were detected by Western blotting.Results(1)As compared with that in the 0μmol/L VB group,the cell survival rate in the 20μmol/L and 40μmol/L VB group showed no significant difference(P>0.05),while that in the 80μmol/L VB group showed significant difference(P<0.05).As compared with that in the 0μmol/L VB
作者
张时真
王业忠
钱程
肖仕印
Zhang Shizhen;Wang Yezong;Qian Cheng;Xiao Shiyin(Department of Neurosurgery,Second Affiliated Hospital of Guangzhou Medical University,Guangzhou 501260,China)
出处
《中华神经医学杂志》
CAS
CSCD
北大核心
2021年第1期40-48,共9页
Chinese Journal of Neuromedicine
基金
国家自然科学基金青年项目(81901117)。
关键词
胶质母细胞瘤
毛蕊花糖苷
上皮间质转化
迁移
侵袭
转化生长因子-Β
Glioblastoma
Verbascoside
Epithelial-mesenchymal transition
Migration
Invasion
Transforming growth factorβ
