Zhang Jie, Pi Jingnan, Liu Yue, Yu Jia, Feng Tao
Research Center of Molecular Medicine and Cancer, Chongqing Medical University, Chongqing 400016, China.
State Key Laboratory of Medical Molecular Biology, Chinese Academy of Medical Sciences, Beijing 100005, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2017 Dec;33(12):1628-1634.
Objective To investigate the effect of knockdown of YTH N-methyladenosine RNA binding protein 2 (YTHDF2) on cell proliferation, cell cycle and apoptosis of MGC-803 human gastric cancer cells in vitro. Methods The TCGA database was downloaded from UCSC Cancer Browser and to search for the differential expressions of YTHDF2 mRNA in gastric cancer tissues. Short hairpin RNA (shRNA) targeting YTHDF2 was designed and cloned into lentivirus expression vector pLKO.1. Furthermore, MGC-803 gastric cancer cells were transfected with pLKO.1-shRNA to knockdown the expression of YTHDF2, which was confirmed by the detection of YTHDF2 mRNA and protein expression using real-time quantitative PCR and Western blotting, respectively. Then cell proliferation was observed by CCK-8 assay, and cell cycle and apoptosis were examined by flow cytometry. Results According to the TCGA database, the expression of YTHDF2 mRNA in gastric cancer was significantly higher than that in the normal tissues. MGC-803 stably expressing YTHDF2-shRNA was successfully established. Furthermore, the proliferation capacity of YTHDF2-shRNA-expressing MGC-803 cells was significantly inhibited compared with the controls. Similarly, the percentage of YTHDF2-shRNA-expressing MGC-803 cells in G1 phase increased and in S phase decreased compared with the controls. Meanwhile, apoptosis ratio of YTHDF2-shRNA-expressing MGC-803 cells was significantly higher compared with the control groups. Conclusion Knockdown of YTHDF2 in MGC-803 cells inhibits cell proliferation and promotes apoptosis.
目的 探讨敲低YTH N-甲基腺苷RNA结合蛋白2(YTHDF2)对人胃癌MGC-803细胞体外增殖、细胞周期及凋亡的影响。方法 从UCSC癌症浏览器下载TCGA数据库,以搜索YTHDF2 mRNA在胃癌组织中的差异表达。设计靶向YTHDF2的短发夹RNA(shRNA)并克隆到慢病毒表达载体pLKO.1中。此外,用pLKO.1-shRNA转染MGC-803胃癌细胞以敲低YTHDF2的表达,分别通过实时定量PCR和蛋白质印迹法检测YTHDF2 mRNA和蛋白质表达来证实。然后通过CCK-8法观察细胞增殖情况,通过流式细胞术检测细胞周期和凋亡情况。结果 根据TCGA数据库,YTHDF2 mRNA在胃癌中的表达明显高于正常组织。成功构建了稳定表达YTHDF2-shRNA的MGC-803细胞系。此外,与对照组相比,表达YTHDF2-shRNA的MGC-803细胞的增殖能力明显受到抑制。同样,与对照组相比,表达YTHDF2-shRNA的MGC-803细胞在G1期的比例增加,在S期的比例降低。同时,表达YTHDF2-shRNA的MGC-803细胞的凋亡率明显高于对照组。结论 在MGC-803细胞中敲低YTHDF2可抑制细胞增殖并促进凋亡。