miR-126在食管癌组织中低表达并抑制食管癌EC109细胞的增殖和迁移  

Down-regulation of miR-126 in esophageal carcinoma tissues and its inhibition effect on proliferation and migration of esophageal carcinoma cell line EC109

作　　者：聂正超 翁文浩[1] 李静[1] 许焱婷 李智[2] 

机构地区：[1]同济大学附属第十人民医院检验科,上海200072 [2]同济大学附属杨浦医院检验科,上海200090

出　　处：《肿瘤》2015年第1期55-64,共10页Tumor

摘　　要：目的:探讨微小RNA(microRNA,miRNA,miR)-126在食管癌组织中的表达及mi R-126对食管癌EC109细胞增殖和迁移的影响。方法:应用实时荧光定量PCR法检测24例食管癌组织及其相应癌旁组织中mi R-126及SOX2(sex determining region Y-box 2)m RNA的表达。将miR-126模拟物(mimics)或其阴性对照(miR-negative control,miR-NC)转染至EC109细胞后,应用实时荧光定量PCR法检测EC109细胞中mi R-126的表达水平,MTT法和Transwell小室法分别检测细胞的增殖能力和迁移能力。构建野生型和突变型SOX2基因3’-端非翻译区(3’-untranslated region,3’-UTR)双荧光素酶报告系统,检测萤火虫荧光素酶的相对活性,以验证mi R-126对SOX2基因的作用靶点。应用实时荧光定量PCR和蛋白质印迹法分别检测转染mi R-126 mimics后,EC109细胞中SOX2 m RNA及其蛋白的表达水平。免疫组织化学法检测食管癌组织及其相应癌旁组织中SOX2蛋白的表达。结果:食管癌组织中miR-126的表达水平明显低于相应的癌旁组织(P<0.01),SOX2 m RNA的表达水平明显高于相应的癌旁组织(P<0.05),且mi R-126与SOX2的表达呈负相关(r=-0.837,P<0.001)。mi R-126mimics转染后,EC109细胞中mi R-126的表达水平明显高于阴性对照组(P<0.01),细胞的增殖能力和迁移能力明显低于阴性对照组(P<0.05,P<0.01)。mi R-126与SOX 2基因的2个预测结合靶点都能特异性结合。转染mi R-126 mimics后,EC109细胞中SOX2 m RNA及蛋白的表达水平明显低于阴性对照组(P<0.01)。食管癌组织中SOX2蛋白的阳性表达率明显高于相应的癌旁组织(P<0.01)。结论:mi R-126在食管癌组织中呈低表达,且对食管癌EC109细胞的增殖和迁移能力具有抑制作用。SOX2可能是mi R-126的靶基因之一。Objective：To investigate the expression of microRNA（miRNA,miR）-126 in esophageal carcinoma tissues and the effect of miR-126 on the proliferation and migration of esophageal cell line EC109.Methods：The expressions of miR-1 26 and sex determining region Y-box 2（SOX2） mRNA in esophageal cancer tissues and their paracancerous tissues from 24 patients were detected by real-time fluorescence-based quantitative PCR.The expression of miR-1 26 in EC1 09 cells after transfection with miR-1 26 mimics or miR-negative control（NC） was detected by realtime fluorescence-based quantitative PCR.The abilities of cell proliferation and migration of EC1 09 cells transfected with miR-1 26 mimics were measured by MTT and Transwell assays,respectively.The dual luciferase reporter vectors containing 3＇-untranslated region（3＇-UTR） with miR-1 26 binding site of wild type or mutant SOX2 gene were constructed by using Dual-Luciferase Reporter Assay System,then the relative activity of firefly luciferase was detected to confirm the binding site of miR-1 26 on S0X2 gene.The expression levels of SOX2 mRNA and protein in EC109 cells transfected with miR-1 26 mimics were detected by real-time fluorescence-based quantitative PCR and Western blotting,respectively.The expression of SOX2 protein in esophageal cancer tissues and their paracancerous tissues was examined by immunohistochemistry.Results：The expression level of miR-1 26 in esophageal cancer tissues was lower than that in paracancerous tissues（P〈0.01）,but the expression level of SOX2 mRNA was opposite（P〈0.05）.The expression of miR-1 26 was negatively correlated with the expression of SOX2（r =- 0.837,P〈0.001）.After transfection with miR-1 26 mimics,the expression level of miR-1 26 in EC1 09 cells was higher than that in miR-NC group（P〈0.01）,but the abilities of cell proliferation and migration were opposite（P〈0.05,P〈0.01）.miR-1 26 can directly target the S0X2 3＇-UTR through two predicted binding sites.The expression levels of SOX2

关 键 词：食管肿瘤 微RNAs 细胞增殖 细胞运动 MIR-126 基因 SOX2 

分 类 号：R735.1[医药卫生—肿瘤]