胃癌细胞Cullin1基因表达与上皮-间充质转化的关系  

Relationship between Cullin1gene expression and epithelial mesenchymal transformation in gastric cancer cells

作　　者：杨沛刚[1] 田园[1] 檀碧波[1] 丁平安 郭洪海[1] 刘洋[1] 张泽 李勇[1] 赵群[1] Yang Peigang;Tian Yuan;Tan Bibo;Ding Pingan;Guo Honghai;Liu Yang;Zhang Ze;Li Yong;Zhao Qun(Department of Surgery,the Fourth Hospital of Hebei Medical University,Shijiazhuang 050011,China)

机构地区：[1]河北医科大学第四医院外三科,石家庄050011

出　　处：《中华实验外科杂志》2021年第10期1926-1930,共5页Chinese Journal of Experimental Surgery

基　　金：河北省医学科学研究重点课题计划(20180547);政府资助临床医学优秀人才培养项目(2019012);河北省高等学校科学技术研究项目(ZD2019139)。

摘　　要：目的检测Cullin1基因在胃癌组织和细胞株中的表达,并探讨其参与肿瘤上皮-间充质转化(EMT)的机制。方法选取2020年1月至2020年3月于河北医科大学第四医院行胃癌根治术12例患者离体标本癌及癌旁组织,应用实时定量反转录聚合酶链反应(RT-qPCR)和蛋白质印迹法(Western blot)技术分别检测12例胃癌与配对癌旁组织中Cullin1 mRNA和蛋白的表达;合成抑制内源性Cullin1表达的小干扰RNA(siRNA)转染胃癌细胞为实验组,非特异性siRNA转染(NS-siRNA)为对照组;用噻唑蓝(MTT)实验检测肿瘤细胞的增殖活性;应用划痕实验及Transwell小室侵袭实验检测细胞的侵袭迁移能力;通过RT-qPCR及Western blot检测转染前后细胞中EMT相关基因E-钙黏蛋白(E-cadherin)、N-钙黏蛋白(N-cadherin)、波形蛋白(Vimentin)、锌指转录因子(Snail)、基质金属蛋白酶-9(MMP-9)的表达。组间比较采用t检验或χ^(2)检验。结果胃癌组织中Cullin1 mRNA和蛋白的表达水平高于癌旁正常组织(4.19±0.50比1.19±0.28,t=-0.583,P<0.05;0.99±0.07比0.40±0.11,t=16.102,P<0.05)。MTT显示转染细胞48 h后,Cullin1-siRNA组细胞活性低于NS-siRNA组和空白组(0.55±0.06比1.16±0.10,t=1.061,P<0.05;0.55±0.06比1.22±0.10,t=13.963,P<0.05)。Cullin1-siRNA转染后细胞的侵袭能力低于NS-siRNA组和空白组(39.17±2.23比83.67±5.79,F=720.65,P<0.05;39.17±2.23比88.00±4.29,F=115.34,P<0.05);Cullin1-siRNA组细胞迁移能力低于NS-siRNA组和空白组[(0.50±0.09) mm比(0.91±0.05) mm,t=-10.064,P<0.05;(0.50±0.09) mm比(0.89±0.07) mm,t=-8.369,P<0.05]。Cullin1-siRNA转染后SGC7901细胞后EMT相关基因N-cadherin、Snail、MMP-9表达均低于空白组表达(1.04±0.05比0.55±0.04,t=10.845,P<0.05;1.05±0.07比0.44±0.04,t=10.493,P<0.05;1.04±0.05比0.37±0.03,t=15.882,P<0.05)。结论 Cullin1基因可能通过调节部分EMT基因而促进了胃癌侵袭转移。Objective Cullin1 gene expression was detected in gastric cancer tissues and cell lines,and the mechanism of Cullin1 gene′s involvement in tumor epithelial mesenchymal transformation(EMT)was discussed.Methods 12 tumor specimens and paracancerous tissues were collected from patients who underwent radical gastrectomy in the Fourth Hospital of Hebei Medical University from January 2020 to March 2020.Real-time quantitative reverse transcription polymerase chain reaction(RT-qPCR)and Western blotting were used to detect the expression of Cullin1 mRNA and protein in 12 gastric cancer patients and their paired adjacent tissues,respectively.Small interfering RNA(siRNA)was synthesized to inhibit endogenous Cullin1 expression and transfected into SGC7901 cells.methyl thiazolyl tetrazolium assay(MTT),assay was used to detect the proliferative activity of tumor cells.The invasion and migration ability of cells was detected by Wound Healing test and Transwell intraventral invasion test.The expressions of EMT-related genes E-cadherin,N-cadherin,Vimentin,Snail and matrix metalloproteinase-9(MMP-9)in the cells before and after transfection were detected by RT-qPCR and Western blotting.In this study,SPSS 21.0 was used for statistical analysis.Quantitative data were tested by Student-t test,and quantitative data were tested byχ^(2) test.P<0.05 was considered statistically significant.Results Cullin1 mRNA and protein expression levels in gastric cancer tissues were significantly higher than those in para-carcinoma normal tissues(4.19±0.50 vs1.19±0.28,t=-0.583,P<0.05;0.99±0.07 vs.0.40±0.11,t=16.102,P<0.05).MTT showed that 48h after transfection,the cell activity of Cullin1-siRNA group was lower than that of NS-siRNA group and blank group(0.55±0.06 vs.1.16±0.10,t=1.061,P<0.05;0.55±0.06 vs.1.22±0.10,t=13.963,P<0.05).The invasion ability of Cullin1-siRNA transfected cells was lower than that of NS-siRNA group and blank group(39.17±2.23 vs.83.67±5.79,F=720.65,P<0.05;39.17±2.23 vs.88.00±4.29,F=115.34,P<0.05);Cell migration

关 键 词：胃癌 Cullin1 基因干扰 上皮-间充质转化 

分 类 号：R735.2[医药卫生—肿瘤]