miR-124抑制口腔鳞状细胞癌CAL27细胞的增殖活性及其对EZH2的靶向调控

口腔医学研究 ›› 2015, Vol. 31 ›› Issue (7): 676-680.

miR-124抑制口腔鳞状细胞癌CAL27细胞的增殖活性及其对EZH2的靶向调控

韩哲^1*,马会强²,彭诚¹

1. 天津医科大学第二医院口腔科天津 300210;
2. 山东省济宁市第一人民医院口腔科

收稿日期:2014-12-04 出版日期:2015-07-28 发布日期:2015-07-28
通讯作者: 韩哲,电话:13920835621
作者简介:韩哲(1986~ ),男,硕士,主治医师,主要从事口腔颌面外科的基础及临床研究。

MiR-124 Inhibits Proliferation of OSCC Cells by Targeting EZH2

HAN Zhe, MA Hui-qiang, PENG Cheng

Dept. Stomatology, The Second Hospital of Tianjin Medical University, Tianjin 300210

Received:2014-12-04 Online:2015-07-28 Published:2015-07-28

摘要/Abstract

摘要： 目的:探讨miR-124对口腔鳞状细胞癌CAL27细胞增殖活性的影响以及其与EZH2的靶向关系。方法:构建pre-miR-124、野生型EZH2(EZH2-WT)和突变型EZH2(EZH2-MT)真核表达载体,采用荧光实时定量PCR(qRT-PCR)检测pre-miR-124的转染效率,MTT法检测miR-124过表达对CAL27细胞增殖活性的影响;另外,利用双荧光素酶报告基因系统、qRT-PCR和Western blot验证miR-124与EZH2的靶向关系。结果:采用NCBI BLAST软件比对载体测序,结果显示pcDNATM6.2-GW-pre-miR-124、pmirGLO- EZH2-WT和pmirGLO- EZH2-MT表达载体构建成功,qRT-PCR显示转染pre-miR-124后CAL27细胞内miR-124的表达水平显著升高(P＜0.001)。MTT分析结果显示miR-124过表达能够明显抑制CAL27细胞的增殖活性(P＜0.05),双光素酶报告基因显示共转染pre-miR-124和EZH2-WT的CAL27细胞荧光活性显著下降(P＜0.001),且qRT-PCR的结果也显示转染pre-miR-124的CAL27细胞内EZH2的表达水平明显低于对照组(P＜0.001),Western blot显示miR-124的过表达能够显著抑制EZH2蛋白水平的表达。结论:miR-124的过表达能够显著抑制口腔鳞状细胞癌CAL27细胞的增殖,并且与EZH2之间存在具有良好的靶向关系。

关键词: 口腔鳞状细胞癌, miR-124, EZH2, 细胞增殖

Abstract: Objective: To assess the effect of miR-124 on proliferation of human OSCC cells and target relationship between miR-124 and EZH2. Methods: Pre-miR-124 sequence was synthesized and cloned into pcDNATM6.2-GW to construct recombinant plasmid pcDNATM6.2-GW-pre-miR-124 and identified by qRT-PCR. EZH2-WT and EZH2-MT sequences were respectively cloned into pmirGLO plasmid. The potential proliferation function of miR-124 on CAL27 cells was analyzed by MTT. The target relationship between miR-124 and EZH2 was identified by dual-luciferase assay system, qRT-PCR and western blot. Results: We constructed successfully the recombinant plasmids, including pcDNATM6. 2-GW-pre-miR-124, pmirGLO- EZH2-WT and pmirGLO- EZH2-MT, and the transfection efficiency of pre-miR-124 was increased in CAL27 cells compared to control group(P＜0.001). In addition, MTT assays showed that miR-124 could suppress the proliferation of CAL27 cells(P＜0.05). Furthermore, a significant reduction of luciferase activities of CAL27 cells cotransfected by pre-miR-124 and EZH2-WT(P＜0.001). The mRNA(P＜0.001) and protein expression levels of EZH2 were decreased in CAL27 cells transfected by pre-miR-124. Conclusion: Overexpression of miR-124 could suppress cell proliferation by targeting EZH2 in CAL27 cells.

Key words: OSCC, miR-124, EZH2, Cell proliferation

中图分类号:

R739.8

韩哲,马会强,彭诚. miR-124抑制口腔鳞状细胞癌CAL27细胞的增殖活性及其对EZH2的靶向调控[J]. 口腔医学研究, 2015, 31(7): 676-680.

HAN Zhe, MA Hui-qiang, PENG Cheng. MiR-124 Inhibits Proliferation of OSCC Cells by Targeting EZH2[J]. Journal of Oral Science Research, 2015, 31(7): 676-680.

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