miR-144-3p靶向E2F8抑制肺腺癌细胞的增殖和侵袭并阻滞细胞周期

miR-144-3p Inhibits Proliferation and Invasion of Lung Adenocarcinoma Cells and Arrests Cell Cycle by Targeting E2F8

摘要

为了探究miR-144-3p在肺腺癌(lung adenocarcinoma, LUAD)中的作用机制, 本研究基于多个数据库对miR-144-3p和其下游靶基因在肺腺癌组织中的表达水平进行了预测; 通过生物信息学方法和文献调研, 确定了E2F转录因子8 (E2F transcription factor 8, E2F8)为miR-144-3p的下游靶基因, 并通过双荧光素酶实验对miR-144-3p和E2F8的靶向结合关系进行了验证; 然后, 通过实时定量聚合酶链反应(real-time quantitative PCR, qRT-PCR)、免疫印迹分析(Western-blot)、细胞毒性检测(Cell Counting Kit-8 assay, CCK-8 assay)、克隆形成、侵袭能力测定和流式细胞术等实验, 对miR-144-3p和E2F8在肺腺癌中的调控机制进行了探究。结果显示, miR-144-3p在肺腺癌组织和细胞中呈显著低表达, 而E2F8则显著高表达, 两者存在靶向关系; 过表达miR-144-3p或敲低E2F8能抑制肺腺癌细胞的增殖和侵袭并阻滞细胞周期于G1期。上述结果表明, miR-144-3p通过负调控E2F8抑制肺腺癌细胞的增殖和侵袭能力, 并阻滞细胞周期于G1期, 从而抑制肺腺癌细胞的恶性进展。这为肺癌新检测、治疗手段的开发提供了参考依据。

Abstract

To investigate the mechanism of miR-144-3p in lung adenocarcinoma (LUAD), the expression le-vels of miR-144-3p and its downstream target gene in LUAD tissues were predicted based on multiple databases. Through bioinformatics methods and based on the related literature, E2F transcription factor 8 (E2F8) was identified as the target gene of miR-144-3p, and the dual-luciferase reporter gene assay was conducted to verify the target binding relationship between miR-144-3p and E2F8. The regulatory mecha-nisms of miR-144-3p and E2F8 in LUAD were explored by real-time quantitative PCR (qRT-PCR), Western-blot, Cell Counting Kit-8 (CCK-8) assay, colony formation assay, invasion assay and flow cytometry. The re-sults showed that miR-144-3p expression was significantly low in LUAD tissues and cells, while E2F8 ex-pression was significantly high. There was a targeting relationship between miR-144-3p and E2F8. Overex-pression of miR-144-3p or knockdown of E2F8 inhibited the proliferation and invasion of LUAD cells and arrested the cell cycle at G1 phase. In summary, miR-144-3p could inhibit the proliferation and invasion of LUAD cells and arrest the cell cycle at G1 phase by negatively regulating E2F8 expression, thereby inhibi-ting the malignant progression of LUAD cells. The results may be helpful for developing new methods in lung cancer detection and treatment.

关键词

miR-144-3p / E2F转录因子8 (E2F8) / 肺腺癌(LUAD) / 增殖 / 侵袭

Key words

miR-144-3p / E2F transcription factor 8 (E2F8) / lung adenocarcinoma (LUAD) / proliferation / in-vasion