miR-135a-5p增强舌鳞状细胞癌细胞对奥沙利铂敏感性研究  

Influence of miR-135a-5p on the sensitivity of tongue squamous cell carcinoma cells to oxaliplatin

作　　者：卢娜 张强 袁荃 王晓波 LU Na;ZHANG Qiang;YUAN Quan;WANG Xiao-bo(Department of Orthodontic Restoration,Affiliated Hospital of North China University of Science and Technology»Tangshan»Hebei 063000,China)

机构地区：[1]华北理工大学附属医院口腔正畸修复,河北唐山063000

出　　处：《中华实用诊断与治疗杂志》2021年第12期1191-1197,共7页Journal of Chinese Practical Diagnosis and Therapy

基　　金：河北省医学科学研究课题计划(20200133)。

摘　　要：目的探讨miR-135a-5p增强舌鳞状细胞癌细胞对奥沙利铂(oxaliplatin,L-OHP)敏感性的作用及其可能机制。方法采用实时荧光定量PCR法检测人口腔角质形成细胞(human oral keratinocyte,HOK)和舌鳞状细胞癌细胞系CAL-27、SCC9、SCC25细胞中miR-135a-5p和B淋巴瘤莫洛尼鼠白血病病毒插入区1(B-lymphoma Moloney murine leukemia virus insertion region-1,BMI1)mRNA相对表达量,采用Western blot法检测4种细胞BMI1蛋白相对表达量。对数生长期SCC25细胞分为miR-NC组(转染miR-NC)、miR-135a-5p模拟物组(miR-135a-5p模拟物)和空白对照组(不进行转染操作),采用实时荧光定量PCR法检测3组细胞miR-135a-5p和BMI1 mRNA相对表达量,采用Western blot法检测3组细胞BMI1蛋白相对表达量,采用荧光素酶报告实验检测miR-135a-5p与BMI1的靶向关系。SCC25细胞加入不同浓度L-OHP,采用CCK-8实验检测L-OHP对细胞的半数抑制浓度,确定L-OHP在后续实验中的浓度。对数生长期SCC25细胞再分为miR-NC+pcDNA-NC组(转染miR-NC和pcDNA-NC)、miR-135a-5p模拟物+pcDNA-NC组(转染miR-135a-5p模拟物和pcDNA-NC)、miR-135a-5p模拟物+pcDNA-BMI1组(转染miR-135a-5p模拟物和pcDNA-BMI1)、对照组(不进行转染操作),采用实时荧光定量PCR法检测细胞miR-135a-5p和BMI1 mRNA相对表达量,采用Western blot法检测细胞BMI1蛋白相对表达量,采用EdU增殖实验检测细胞增殖情况,采用Transwell实验检测迁移细胞数和侵袭细胞数,采用流式细胞术检测细胞凋亡率。结果CAL-27、SCC9、SCC25细胞miR-135a-5p相对表达量(0.43±0.04、0.62±0.09、0.41±0.07)均低于HOK细胞(1.00±0.15)(P<0.05),BMI1 mRNA(4.57±0.67、2.64±0.59、5.82±0.87)和蛋白(0.61±0.22、0.34±0.28、1.00±0.24)相对表达量均高于HOK细胞(1.00±0.31、0.21±0.14)(P<0.05);与HOK细胞比较,SCC25细胞miR-135a-5p、BMI1 mRNA和蛋白相对表达量差异最大,选用SCC25细胞进行后续实验。miR-135a-5p模拟物组细胞miR-135a-5p相对表达量高于miR-NC�Objective To investigate the role of miR-135 a-5 p in increasing the sensitivity of tongue squamous cell carcinoma to oxaliplatin(L-OHP)and its possible mechanism.Methods The relative expressions of miR-135 a-5 p and B lymphoma Moloney murine leukemia virus insertion region-1(BMI1)mRNA in human oral keratinocyte HOK and tongue squamous cell carcinoma cell line(CAL-27,SCC9,SCC25)were detected by real-time fluorescence quantitative PCR(qRT-PCR).Western blot was used to detect the relative expression of BMI1 protein in those four kinds of cells.SCC25 cells in logarithmic growth phase were divided into miR-NC group(transfected with miR-NC),miR-135 a-5 p mimics group(transfected with miR-135 a-5 p mimics)and blank control group(without transfection).The relative expressions of miR-135 a-5 p and BMI1 mRNA were detected by qRT-PCR,the relative expression of BMI1 protein was detected by Western blot in three groups,and the targeting relationship between miR-135 a-5 p and BMI1 was detected by luciferase reporting assay.SCC25 cells were added with different concentrations of L-OHP,and the half maximal inhibitory concentration(IC50)of L-OHP on cells was detected by CCK-8 assay to determine the concentration of L-OHP in the subsequent experiments.SCC25 cells in logarithmic growth phase were subdivided into miR-NC+pcDNA-NC group(transfected with miR-NC and pcDNA-NC),miR-135 a-5 p mimics+pcDNA-NC group(transfected with miR-135 a-5 p mimics and pcDNA-NC),miR-135 a-5 p mimics+pcDNA-BMI1 group(transfected with miR-135 a-5 p mimics+pcDNA-BMI1)and control group(without transfection).The relative expressions of miR-135 a-5 p and BMI1 mRNA were detected by qRT-PCR,the relative expression of BMI1 protein was detected by Western blot,the cell proliferation rate was detected by EdU proliferation assay,the number of migrating cells and the number of invading cells were detected by Transwell assay,and the apoptosis rate was detected by flow cytometry.Results The relative expression of miR-135 a-5 p was lower in CAL-27(0.43±0.04),SCC9(0.6

关 键 词：舌鳞状细胞癌 奥沙利铂 miR-135a-5p B淋巴瘤莫洛尼鼠白血病病毒插入区1 敏感性 

分 类 号：R739.86[医药卫生—肿瘤]