c-Myc调控lncRNA KCTD13-DT对口腔鳞癌的影响  

Effects of c-Myc regulation of lncRNA KCTD13-DT on oral squamous carcinoma

作　　者：李文丽 朱友明[1,2] 何家才 Li Wenli;Zhu Youming;He Jiacai(College&Hospital of Stomatology,Anhui Medical University,Hefei 230032;Key Lab.of Oral Diseases Research of Anhui Province,Hefei 230032)

机构地区：[1]安徽医科大学口腔医学院,安徽医科大学附属口腔医院,合肥230032 [2]安徽省口腔疾病研究中心实验室,合肥230032

出　　处：《安徽医科大学学报》2024年第10期1759-1768,共10页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金项目(编号:81771117);安徽省自然科学基金项目(编号:2308085Y23)。

摘　　要：目的探讨长链非编码RNA(lncRNAs)lncRNA KCTD13-DT在口腔鳞癌(OSCC)中的作用机制及与转录因子c-Myc的潜在相互作用,为OSCC患者提供新的治疗靶点和途径。方法用qRT-PCR检测OSCC及癌旁组织中lncRNA KCTD13-DT的表达。检测敲低和过表达人舌鳞癌细胞HN6、CAL27中c-Myc后lncRNA KCTD13-DT的表达。采用荧光原位杂交实验(FISH)检测lncRNA KCTD13-DT在细胞内的定位。采用双荧光素酶报告基因分析c-Myc与lncRNA KCTD13-DT启动子区域的靶向结合作用。通过慢病毒感染的方式在人OSCC细胞系HN6、CAL27中构建lncRNA KCTD13-DT敲低和过表达的稳转细胞系,通过qRT-PCR检测lncRNA KCTD13-DT的敲低和过表达效率;采用生长曲线实验、CCK-8法、克隆形成实验检测细胞增殖变化,采用划痕实验和Transwell检测细胞迁移情况。结果lncRNA KCTD13-DT在OSCC组织和OSCC细胞系(HN6、CAL27)中表达水平均降低,Western blot验证在HN6、CAL27中敲低和过表达c-Myc的效果,实验表明过表达c-Myc显著下调lncRNA KCTD13-DT水平,敲低c-Myc明显上调lncRNA KCTD13-DT水平。双荧光素酶报告基因显示c-Myc可靶向调控lncRNA KCTD13-DT,c-Myc可以参与调节并且抑制lncRNA KCTD13-DT的转录活性。FISH显示lncRNA KCTD13-DT主要存在于细胞核中。生长曲线实验、CCK-8法、细胞划痕实验、Transwell、克隆形成等实验显示,敲低lncRNA KCTD13-DT促进OSCC细胞的生长增殖,过表达lncRNA KCTD13-DT明显抑制OSCC细胞增殖与迁移能力。结论LncRNA KCTD13-DT受c-Myc负性调控,敲低lncRNA KCTD13-DT会促进细胞增殖与迁移,反之则抑制细胞增殖与迁移。Objective To explore the action mechanism of long non-coding RNA(lncRNAs)lncRNA KCTD13-DT in oral squamous cell carcinoma(OSCC)and its potential interaction with transcription factor c-Myc,providing a potential diagnostic and therapeutic target for patients with OSCC.Methods The expression of lncRNA KCTD13-DT in OSCC and paracancerous tissues was detected by qRT-PCR.The effects of c-Myc overexpression and knockdown on human tongue squamous carcinoma cells HN6 and CAL27 were detected by qRT-PCR.Fluorescence in situ hybridization(FISH)assessed the localization of lncRNA KCTD13-DT in cells.A dual luciferase reporter gene was used to analyze the role of c-Myc in target binding to the promoter region of lncRNA KCTD13-DT.Stable cell lines with knockdown or overexpression of lncRNA KCTD13-DT were constructed in human OSCC cell lines HN6 and CAL27 by lentiviral infection,and the knockdown and overexpression efficiencies of lncRNA KCTD13-DT were detected by qRT-PCR.Cell proliferation changes were detected by growth curve assay,CCK-8 assay,colony formation assay,and cell migration was detected by scratch assay and Transwell.Results lncRNA KCTD13-DT expression level was reduced in OSCC tissues and OSCC cells(HN6,CAL27),and Western blot verified that after knocking down and overexpression of c-Myc in HN6 and CAL27,the qRT-PCR experiments showed that c-Myc negatively regulated lncRNA KCTD13-DT,and overexpression of c-Myc significantly down-regulated lncRNA KCTD13-DT;knockdown of c-Myc significantly up-regulated lncRNA KCTD13-DT levels.Dual luciferase reporter gene showed that c-Myc could target lncRNA KCTD13-DT,and c-Myc could be involved in regulating and repressing the transcriptional activity of lncRNA KCTD13-DT.FISH showed that lncRNA KCTD13-DT mainly existed in the nucleus.Growth curve assay,CCK-8 assay,cell scratch assay,Transwell,and colony formation assay showed that knockdown of lncRNA KCTD13-DT promoted the growth and proliferation of OSCC cells,and overexpression of lncRNA KCTD13-DT significantly inhibited the prol

关 键 词：长链非编码RNA C-MYC 口腔鳞癌 细胞增殖 细胞迁移 

分 类 号：R739.86[医药卫生—肿瘤]