lncRNA JPX对鼻咽癌细胞增殖、侵袭能力的调控作用及其机制

Regulatory effects of lncRNA JPX on proliferation and invasion of nasopharyngeal carcinoma cells and the mechanism

目的观察长链非编码RNA(lncRNA)JPX对鼻咽癌细胞增殖、侵袭能力的调控作用并探讨其机制。方法取鼻咽癌细胞系CNE-1、5-8F、6-10B、HONE-1、C666-1及鼻咽正常上皮细胞系NP69,采用RT-qPCR法检测细胞中lncRNA JPX表达,取lncRNA JPX表达最高的C666-1细胞作为研究细胞。C666-1细胞随机分为JPX降表达组、降表达对照组、JPX高表达组、高表达对照组,分别转染JPX沉默序列sh-JPX、阴性对照序列sh-NC及JPX过表达质粒pcDNA3.1-JPX、阴性对照质粒pcDNA 3.1-NC。采用MTT法观察细胞增殖能力,Transwell实验观察细胞侵袭能力。利用StarBase数据库对lncRNA JPX进行靶向miRNA及基因的预测,发现lncRNA JPX与miR-1265存在靶向结合位点、miR-1265与MAT1存在靶向结合位点;采用双荧光素酶实验对靶向关系进行验证显示,lncRNA JPX可与miR-1265靶向结合并对miR-1265表达具有抑制作用,miR-1265可与MAT1靶向结合并对MAT1表达具有抑制作用。进一步将C666-1细胞随机分为对照组、JPX降表达组、JPX降表达+miR-1265降表达组、JPX降表达+MAT1降表达组,分别转染NC对照序列、sh-JPX序列、sh-JPX+miR-1265 inhibitor序列、sh-JPX+si-MAT1序列。采用Western blotting法检测各组细胞MAT1蛋白表达、MTT法观察细胞增殖能力,Transwell实验观察细胞侵袭能力。结果细胞活力、侵袭细胞数JPX高表达组>高表达对照组、降表达对照组>JPX降表达组(P均<0.05)。MAT1蛋白表达、细胞活力及侵袭细胞数对照组、JPX降表达+miR-1265降表达组>JPX降表达组>JPX降表达+MAT1降表达组(P均<0.05)。结论lncRNA JPX可促进鼻咽癌细胞的增殖、侵袭,其机制可能与靶向调控miR-1265/MAT1轴有关。

Objective To observe the regulatory effects of long non-coding RNA(lncRNA)JPX on the proliferation and invasion of nasopharyngeal carcinoma cells and to explore their mechanism.Methods The nasopharyngeal carcinoma cell lines CNE-1,5-8F,6-10B,HONE-1,C666-1 and nasopharyngeal normal epithelial cell line NP69 were selected,and we used RT-qPCR to detect the expression of lncRNA JPX in the cells.C666-1 cells with the highest expression of lncRNA JPX were taken as the study cells.C666-1 cells were randomly divided into the JPX down-expression group,downexpression control group,high JPX expression group,and high expression control group,respectively.They were transfected with JPX silencing sequence sh JPX,negative control sequence sh NC,JPX over-expression plasmid pcDNA3.1-JPX,and negative control plasmid pcDNA 3.1-NC,respectively.MTT assay was used to observe cell proliferation ability,and Transwell assay was used to observe cell invasion ability.We used the StarBase database to predict the targeted miRNAs and genes of lncRNA JPX,and found that lncRNA JPX had a targeted binding site with miR-1265,and miR-1265 had a targeted binding site with MAT1;the dual luciferase assay was used to verify the targeted relationship,and it showed that lncRNA JPX could bind to miR-1265 and inhibit miR-1265 expression,and miR-1265 could bind to MAT1 and inhibit MAT1 expression.Further,C666-1 cells were randomly divided into the control group,JPX down-regulation group,JPX down-regulation+miR-1265 down-regulation group,and JPX down-regulation+MAT1 down-regulation group,and were transfected with NC control sequence,sh JPX sequence,sh JPX+miR-1265 inhibitor sequence,and sh JPX+si-MAT1 sequence,respectively.Western blotting was used to detect the expression of MAT1 protein in cells of each group,MTT assay was used to observe cell proliferation ability,and Transwell assay was used to observe cell invasion ability.Results Cell viability and the number of invasive cells were as follows:the high JPX expression group>the high expression control group,and down-expression control group>JPX down-expression group(all P<0.05).The MAT1 protein expression,cell viability,the number of invasive cells were as follows:control group,JPX down-regulation+miR-1265 downregulation group>JPX down-regulation group>JPX down-regulation+MAT1 down-regulation group(all P<0.05).Conclusion LncRNA JPX can promote the proliferation and invasion of nasopharyngeal carcinoma cells,and its mechanism may be related to targeted regulation of the miR-1265/MAT1 axis.