Background: It has been proven that circular RNAs (circRNAs) play an important role on the process of many types cancer and circUBAP2 was a cancer-promoting circRNA, however, the role and mechanism in lung cancer was not clear. The aim of this study is to investigate the effects of circUBAP2 on cell proliferation and invasion of human lung cancer A549 cells.
Methods: CCK-8 assay was employed to detect the effect of circUBAP2 sliencing on cell proliferation of A549 cells. Fow cytometry was applied to detect the impact of circUBAP2 sliencing on cell cycle and cell anoikis, and Transwell invasion assay was applied to determine cell invasion of A549 cells. We also employed Western blot and Real-time PCR to determine the expressions of CDK6, cyclin D1, p27 and c-IAP1, Bcl-2, Survivin, Bax, FAK, Rac1 and MMP2, and the activities of JNK and ERK1/2, luciferase report gene assay was used to detect the targets.
Results: CCK-8 assay showed that the inhibition of cell proliferation in the circUBAP2-siRNA group compared to untreated group and siRNA control group. Results of cell cycle detected by flow cytometry showed that cell cycle arrestd at G0/G1 after circUBAP2 silencing, cell apoptosis rate increased also. We also found that after circUBAP2 silencing, cell invasion of A549 cells was significantly inhibited. Western blot and Real-time PCR results showed that expression of CDK6, cyclin D1, c-IAP1, Bcl-2, Survivin, FAK, Rac1 and MMP2 were down-regulated, and the expression of p27 and Bax were up-regulated. Moreover, the activities of JNK and ERK1/2 were inhibited because of circUBAP2 silencing, the target genes were miR-339-5p, miR-96-3p and miR-135b-3p.
Conclusions: CircUBAP2 plays an important role in the proliferation and invasion of human lung cancer. Silencing of circUBAP2 might be a novel target for molecular targeted therapy of patients with lung cancer. .
背景与目的 已有的研究证明环状RNA（circular RNAs, circRNAs）在多种肿瘤发生发展中起重要作用，circUBAP2是一种促癌的circRNA，但是其在肺癌中的作用和机制尚不明确。本研究旨在探讨circular RNA UBAP2（circUBAP2）对人肺癌A549细胞体外增殖和侵袭能力的影响并初步探讨其机制。方法 应用CCK-8法检测circUBAP2表达沉默对A549细胞生长的抑制作用，采用流式细胞仪技术检测circUBAP2表达沉默对A549细胞周期和细胞失巢凋亡的影响，Transwell实验检测circUBAP2表达沉默对A549细胞侵袭能力的影响，Western blot和Real-time PCR检测CDK6、cyclin D1、p27、c-IAP1、Bcl-2、Survivin、Bax、FAK、Rac1、MMP2的表达变化以及JNK和ERK1/2的活性，荧光素酶报告技术检测circUBAP2直接靶基因。结果 CCK-8实验结果显示circUBAP2表达沉默后，A549细胞株增殖能力受抑制；流式细胞结果显示，circUBAP2表达沉默后，细胞周期停滞在G0期/G1期，且细胞凋亡率显著增加；Transwell实验结果显示，细胞体外侵袭能力明显受到抑制。Western blot和Real-time PCR结果显示，circUBAP2表达沉默后，CDK6、cyclin D1、c-IAP1、Bcl-2、Survivin、FAK、Rac1、MMP2等基因的表达水平显著下降，而p27和Bax的表达显著上升，而且JNK、ERK1/2的活性也受到明显的抑制，circUBAP2直接靶基因为miR-339-5p、miR-96-3p和miR-135b-3p。结论 circUBAP2在肺癌的体外增殖和侵袭中发挥重要的作用，抑制其表达，有望成为肺癌分子靶向治疗的新靶点。.
Keywords: A549 cells; Invasion; Lung neoplasms; circUBAP2.