文献类型：期刊文献

中文题名：miR-335-5p过表达对胃癌基因表达谱的影响及筛选基因的验证研究

英文题名：Effect of miR-335-5p overexpression on gene expression profile and validation of selected genes in gastric cancer

作者：王宏伟[1];赵雪灵[1];王蒙[2];高夏青[2];陈凤琴[3];宁月[2];李海龙[2]

第一作者：王宏伟

机构：[1]甘肃中医药大学附属医院检验科,甘肃兰州730020;[2]甘肃中医药大学第一临床医学院,甘肃兰州730101;[3]甘肃中医药大学附属医院功能科,甘肃兰州730020

第一机构：甘肃中医药大学第二附属医院

年份：2023

卷号：44

期号：23

起止页码：2874

中文期刊名：国际检验医学杂志

外文期刊名：International Journal of Laboratory Medicine

收录：CSTPCD

基金：甘肃省自然科学基金项目(21JR7RA579);甘肃省卫生行业科研项目(GSWSKY2022-25);甘肃省中医药科研课题(GZKG-2022-53)。

语种：中文

中文关键词：微小RNA-335-5p;胃癌;信号通路;表达谱芯片

外文关键词：microRNA-335-5p;gastric cancer;signaling pathways;expression profiling chip

摘要：目的探讨上调微小RNA(miR)-335-5p对胃癌细胞增殖和侵袭能力的影响,研究并验证miR-335-5p过表达后胃癌细胞中差异表达的靶基因。方法使用miR-335-5p慢病毒载体感染胃癌细胞,通过嘌呤霉素筛选构建稳转细胞株,采用MTT法和Transwell实验观察上调miR-335-5p对胃癌细胞增殖和侵袭能力的影响,使用表达谱芯片筛选过表达miR-335-5p组和阴性对照组细胞之间差异表达的基因。使用miRSystem网站的生物信息学数据库进行miR-335-5p的靶基因预测,以至少3个软件支持的基因与芯片筛选基因取得交集,采用DAVID 6.8软件对miR-335-5p的靶基因开展京都基因与基因组百科全书信号通路富集分析,并选择部分差异表达最明显的基因进行实验验证。结果上调miR-335-5p表达可抑制胃癌细胞的增殖和侵袭能力,表达谱芯片筛选结合生物信息学预测分析共获得212个miR-335-5p靶基因,富集在20个肿瘤相关的信号通路中。选择其中的靶基因环腺苷酸反应元件结合蛋白1(CREB1)、苏氨酸激酶3(AKT3)、外被体包被蛋白β2亚基(COPB2)、芳香烃受体核易位蛋白2(ARNT2)、转化生长因子(TGF)-β2、Kirsten大鼠肉瘤病毒癌基因(KRAS)、血小板源性生长因子-β(PDGF-β)进行验证,实时荧光定量PCR和蛋白质印迹法结果表明,过表达miR-335-5p后可抑制上述靶基因的表达(P<0.05)。结论上调miR-335-5p的表达能抑制胃癌细胞的增殖和侵袭能力,其机制与抑制其靶基因CREB1、AKT3、COPB2、ARNT2、TGF-β2、KRAS、PDGF-β的表达有关。
Objective To investigate the effects of up-regulation of microRNA(miR)-335-5p on the proliferation and invasion of gastric cancer cells,and to study and verify the differentially expressed target genes in gastric cancer cells after miR-335-5p overexpression.Methods Gastric cancer cells were infected with miR-335-5p lentivirus vector and screened by puromycin to establish a stable cell line.The effects of miR-335-5p upregulation on the proliferation and invasion of gastric cancer cells were observed by MTT assay and Transwell assay.The differentially expressed genes between the miR-335-5p overexpression group and the negative control group were screened by expression microarray.The target genes of miR-335-5p were predicted by the bioinformatics database of miRSystem website,and at least three genes supported by the software were intersected with the genes screened by microarray.DAVID 6.8 software was used to perform Kgoto Encyclopedia of Genes and Genomes signaling pathway enrichment analysis of the target genes of miR-335-5p.The mos significantly differentially expressed genes were selected for experimental verification.Results Overexpression of miR-335-5p inhibited the proliferation and invasion of gastric cancer cells.A total of 212 target genes of miR-335-5p were identified,which were enriched in 20 tumor-related signaling pathways.The target genes cyclic adenylate response element binding protein 1(CREB1),threonine kinase 3(AKT3),exosome envelope protein beta 2 subunit(COPB2),aryl hydrocarbon receptor nuclear translocation protein 2(ARNT2),transforming growth factor(TGF)-β2,Kirsten rat sarcoma viral oncogene(KRAS),platelet-derived growth factor-β(PDGF-β)were selected for verification.The results of real-time fluorescent quantitative PCR and Western blot showed that overexpression of miR-335-5p could inhibit the expression of the above target genes(P<0.05).Conclusion Overexpression of miR-335-5p could inhibit the proliferation and invasion of gastric cancer cells,and the mechanism is related to the down-regulation of its target genes CREB1,AKT3,COPB2,ARNT2,TGF-Β2,KRAS and PDGF-β.