葡萄糖转运蛋白1影响食管癌细胞对2-脱氧葡萄糖敏感性的研究  

Study of glucose transporter type 1 enhances esophageal cancer cell sensitivity to 2-deoxyglucose treatment

作　　者：程遥[1] 李少民[1] 党诚学[2] 周斌[1] 宋永春[2] 刁冬梅[2] CHEN Yao;LI Shao-min;DANG Chen-xue;ZHOU Bin;SONG Yong-chun;DIAO Dong-mei(Department of Thoracic Surgery,Second Affiliated Hospital of Xi’an Jiaotong University,Xi’an 710004,Shannxi Province,China;Department of Surgical Oncology,First Affiliated Hospital of Xi’an Jiaotong University,Xi’an 710061,Shannxi Province,China)

机构地区：[1]西安交通大学第二附属医院胸外科,陕西西安710004 [2]西安交通大学第一附属医院肿瘤外科,陕西西安710061

出　　处：《世界临床药物》2020年第4期253-260,共8页World Clinical Drug

基　　金：国家自然科学基金(81501826);陕西省自然科学基础研究计划(2020-JQ-540&2020-JQ-504)。

摘　　要：目的糖酵解抑制剂2-脱氧葡萄糖(2-deoxyglucose,2DG)可以特异性阻断癌细胞Warburg效应发挥抗肿瘤作用,其靶点是葡萄糖转运蛋白(glucose transporter type 1,GLUT-1)。但GLUT-1的表达是否影响2DG的作用效率尚不得知,因此,笔者拟在食管癌细胞中证明GLUT-1的表达是否影响食管癌对2DG的敏感性及其可能的作用机制。方法本研究通过体外试验,使用细胞转染技术过表达及敲除食管癌细胞GLUT-1蛋白,将细胞行2DG干预后,使用噻唑蓝比色[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide,MTT]法测定细胞活性,酶联免疫吸附试验(enzyme linked immunosorbent assay,ELISA)法测定上清葡萄糖含量。并进一步使用免疫蛋白印迹(western blot,WB)法测定细胞内磷酸化蛋白激酶B(phosphor-protein kinase B,p-Akt)及LC3蛋白家族B(LC3-B)表达水平。结果Eca-109和TE-1细胞中,GLUT-1高表达组72 h细胞数是空白对照组的1.44倍和1.46倍,葡萄糖消耗是空白对照组的1.62倍和1.39倍,GLUT-1低表达组的细胞数分别是相应空白对照组的59.4%和58.2%,葡萄糖消耗分别是空白对照组的33.1%和27.6%,组间差异均有统计学意义(P<0.05);加入2DG干预后,GLUT-1高表达组72 h细胞抑制率分别为56%及71%,葡萄糖消耗减少50%及49%,空白对照组的抑制率分别为43%及34%,葡萄糖消耗减少15%及27%,GLUT-1低表达组分别为28%及17%,葡萄糖消耗减少7%及9%,组间差异均有统计学意义(P<0.05)。使用LY294002阻断蛋白激酶B(protein kinase B,Akt)通路后,两株细胞的组间差异均无统计学意义(P>0.05)。WB法检测显示GLUT-1表达与p-Akt呈正相关,高表达GLUT-1的细胞LC3-B水平较低,使用2DG干预后,所有细胞LC3-B水平均明显增高,p-Akt表达显著降低。结论食管癌细胞中,GLUT-1是2DG的作用靶点,其表达上调增强细胞对2DG的敏感性,Akt信号通路可能参与其相关机制。同时,2DG还可能通过Akt通路影响食管癌细胞的自噬水平,值得临床进一步�Objective Glycolysis inhibitor 2-deoxyglucose(2DG)can block the Warburg effect of tumor and play an anti-tumor role,and its target is glucose transporter type 1(GLUT-1).However,which factors took part in influencing 2DG sensitivity is still unknown.In this study,we planed to unveil whether GLUT-1 expression was a key factor of esophageal react to 2DG treatment and the possible mechanism.Methods In vitro experiments,cell transfection technique was used to overexpress and knockout the GLUT-1 protein of esophageal cancer cells.After 2DG incubation,cell activity was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and glucose content in the medium was measured by enzyme linked immunosorbent assay.Then we used western blot(WB)to detect cell Akt,LC3-B expression level,wondering whether 2DG in esophageal cancer may participate in the mechanism of action.Results In Eca-109 and TE-1 cells,the number of cells in the high GLUT-1 expression group was 1.44 and 1.46 times of the blank control group in 72 hours,and the glucose consumption in the high GLUT-1 expression group was 1.62 and 1.39 times of the blank control group,the number of cells in GLUT-1 low expression group was 59.4%and 58.2%of the corresponding blank control group,and the glucose consumption was 33.1%and 27.6%of the blank control group respectively,the difference between the two groups was statistically significant(P<0.05).After the treatment with 2DG,the cell inhibition rate of GLUT-1 high expression group was 56%and 71%,the glucose consumption was reduced by 50%and 49%,the inhibition rate of blank control group was 43%and 34%,the glucose consumption was reduced by 15%and 27%respectively,GLUT-1 low expression group was 28%and 17%,glucose consumption decreased by 7%and 9%,the difference between the two groups was statistically significant(P<0.05).After LY294002 blocked the protein kinase B(Akt)pathway,there was no significant difference between the two cell lines(P>0.05).WB method showed that the expression of GLUT-1 was positivel

关 键 词：食管癌 Warburg效应 葡萄糖转运蛋白1 2-脱氧葡萄糖 

分 类 号：R459.9[医药卫生—治疗学]