沉默己糖激酶结构域蛋白1表达对胃癌细胞增殖、侵袭、转移和上皮间充质转化的影响  

Effect of silencing hexokinase domain containing protein 1 expression on proliferation, invasion, metastasis and epithelial-mesenchymal transformation of gastric cancer cells

作　　者：张作艳 吴林文 马家思 郑珊珊 何镭 张翀[1] 曾玲晖[1] ZHANG Zuo-yan;WU Lin-wen;MA Jia-si;ZHENG Shan-shan;HE Lei;ZHANG Chong;ZENG Ling-hui(School of Medicine,Zhejiang University City College,Hangzhou 310015,China)

机构地区：[1]浙江大学城市学院医学院,浙江杭州310015

出　　处：《中国药理学与毒理学杂志》2020年第2期112-118,共7页Chinese Journal of Pharmacology and Toxicology

基　　金：浙江大学城市学院教师科研基金(J-19006)

摘　　要：目的探讨己糖激酶结构域蛋白1(HKDC1)成为胃癌潜在治疗靶点的可能性。方法用脂质体法向胃癌细胞NCI-N87,BGC-823和HGC-27转染沉默HKDC1表达的小干扰RNA(siRNA)(siRNA-1和siRNA-2),同时设阴性对照siRNA(siRNA-NC)组,分别获得NCI-N87-siRNA-NC,NCI-N87-siRNA-1和NCI-N87-siRNA-2细胞,BGC-823-siRNA-NC,BGC-823-siRNA-1和BGC-823-siRNA-2细胞,HGC-27-siRNA-NC,HGC-27-siRNA-1和HGC-27-siRNA-2细胞,用Western印迹法检测siRNA对HKDC1表达的抑制效果,并计算抑制率。将上述细胞接种入96孔板,孵育72 h后,磺酰罗丹明B(SRB)染色法检测细胞存活,计算细胞存活率。将上述细胞分别接种入无基质胶和铺有基质胶的Transwell小室,24 h后计数小室下室细胞数,分别检测细胞迁移和侵袭能力。收集上述细胞提取蛋白质,用Western印迹法检测上皮间充质转化相关蛋白E-钙黏蛋白、N-钙黏蛋白和Snail的表达。结果分别转染siRNA-1和siRNA-2,对NCIN87-siRNA-1和NCI-N87-siRNA-2细胞HKDC1表达的抑制率分别为(20.5±0.1)%和(48.6±0.1)%,对BGC-823-siRNA-1和BGC-823-siRNA-2细胞的抑制率分别为(56.5±0.1)%和(58.9±0.1)%,对HGC-27-siRNA-1和HGC-27-siRNA-2细胞的抑制率分别为(63.5±0.1)%和(85.9±0.1)%。与对应的阴性对照组细胞相比,NCI-N87-siRNA-1和NCI-N87-siRNA-2细胞的存活率分别为(68.2±2.5)%和(70.4±2.1)%(P<0.01),BGC-823-siRNA-1和BGC-823-siRNA-2细胞的存活率分别为(77.0±0.1)%和(73.7±0.1)%(P<0.05),HGC-27-siRNA-1和HGC-27-siRNA-2细胞的存活率分别为(63.2±0.6)%和(70.4±0.1)%(P<0.01,P<0.05)。NCI-N87-siRNA-NC,NCI-N87-siRNA-1和NCI-N87-siRNA-2细胞的迁移数分别为488±43,319±27和262±37(P<0.01),侵袭数分别为143±8,68±2和30±2(P<0.01);BGC-823-siRNA-NC,BGC-823-siRNA-1和BGC-823-siRNA-2细胞的迁移数分别为1131±69,830±159和579±57(P<0.01),侵袭数分别为1127±113,781±97和565±96(P<0.01);HGC-27-siRNA-NC,HGC-27-siRNA-1和HGC-27-siRNA-2细胞的迁移数分别为453±36,190±25和57±16(P<0.01),侵袭数分别为645±53OBJECTIVE To explore the potential of hexokinase domain-containing protein 1(HKDC1)as a therapeutic target for gastric cancer.METHODS The liposome method was used to transfect small interference RNA(si RNA)(si RNA-1 and si RNA-2)that could silence HKDC1 expression into gastric cancer cells NCI-N87,BGC-823 and HGC-27,respectively,and the negative control si RNA(si RNA-NC)groups were set.Then,the NCI-N87-siRNA-NC,NCI-N87-siRNA-1 and NCI-N87-siRNA-2 cells,BGC-823-siRNA-NC,BGC-823-siRNA-1 and BGC-823-siRNA-2 cells,HGC-27-siRNA-NC,HGC-27-siRNA-1 and HGC-27-siRNA-2 cells were prepared.Western blotting was used to detect the inhibitory effect of siRNA on expressions of HKDC1 protein and calculate the inhibitory rate.Cells were seeded into the96-well plate.After 72 h,sulforhodamine B assay was used to detect the proliferation of gastric cancer cells and calculate the cell viability.Cells were planted into the upper chamber of matrigel free and matrige-transwell.The numbers of cells in the lower chamber were calculated to detect the effect of migration and invasion on gastric cancer cells.Western blotting was used to detect the expressions of epithelial-mesenchymal transition-related proteins,such as E-cadherin,N-cadherin and Snail,when the cells were harvested and lysed.RESULTS The inhibitory rates of siRNA-1 and siRNA-2 on HKDC1 expression in NCI-N87-si RNA-1 and NCI-N87-si RNA-2 cells were(20.5±0.1)%and(48.6±0.1)%respectively,those of BGC-823-si RNA-1 and BGC-823-si RNA-2 cells were(56.5±0.1)%and(58.9±0.1)%respectively,and those of HGC-27-siRNA-1 and HGC-27-siRNA-2 cells were(63.5±0.1)%and(85.7±0.1)%respectively.Compared with the corresponding si RNA-NC group,the survival rates of NCI-N87-si RNA-1 and NCI-N87-siRNA-2 cells were(68.2±2.5)%and(70.4±2.1)%(P<0.01)respectively,those of BGC-823-siRNA-1 and BGC-823-siRNA-2 cells were(77.0±0.1)%and(73.7±0.1)%(P<0.05)respectively,and those of HGC-27-si RNA-1 and HGC-27-si RNA-2 cells were(63.2±0.6)%and(70.4±0.1)%(P<0.01,P<0.05)respectively.The migration numbers of N

关 键 词：己糖激酶结构域蛋白1 胃癌 转移 增殖 

分 类 号：R735[医药卫生—肿瘤]