上调miR-27a对胃癌干细胞干性特征的影响

Effect of upregulation of miR-27a on the characteristics of stemness in gastric cancer stem cells

目的:探讨上调miR-27a对胃癌干细胞(gastric cancer stem cells, GCSCs)干性特征的影响。方法:选择人胃癌细胞株MKN-45,在含有表皮生长因子(epidermal growth factor, EGF)及成纤维生长因子(basic fibroblast growth factor, bFGF)的无血清培养液中培养,制备成球细胞,并分成实验组(miR-27a mimics组)与对照组(negative control, NC组)。实验组转染miR-27a mimics,对照组转染阴性序列质粒,采用实时定量聚合酶链式反应(quantitative real-time polymerase chain reaction,q RT-PCR)检测两组细胞miR-27a的表达变化,qRT-PCR和Western Blot检测两组细胞干性因子八聚体结合转录因子4(octamer-binding transcription factor 4, Oct-4)和性别决定区Y框蛋白2(sex determining region Y box 2, Sox2)的表达变化,成球实验检测两组细胞的成球能力。结果:与对照组比较,实验组miR-27a与干性因子Oct-4和Sox2蛋白及mRNA表达均显著上调(P<0.05),实验组成球能力明显增强(P<0.05)。结论:miR-27a在维持GCSCs的干性特征中发挥重要作用。

Objective : To investigate the effect of upregulation of miR-27a on the characteristics of stemness in gastric cancer stem cells(GCSCs). Methods : Spheroid body-forming cells were derived by placing the MKN-45 cells into serum-free 1640 culture medium supplemented with epidermal growth factor(EGF) and basic fibroblast growth factor(bFGF), and then were tranfected with miR-27a mimics(miR-27a mimics group) and negative sequence(NC group), respectively. The expression of miR-27a was detected using quantitative real-time polymerase chain reaction(qRT-PCR), and the expression of octamerbinding transcription factor 4(Oct-4) and sex determining region Y box 2(Sox2) was detected using qRT-PCR and Western Blot. Furthermore, the ability of forming spheroid body was detected by using spheroid body-forming experiment. Results : Compared with NC group, the expression levels of miR-27a, Oct-4 and Sox2 were significantly increaseded in the miR-27a mimics group(P<0.05),and the ability of forming spheroid body was also enhanced obviously(P<0.05). Conclusions : MiR-27a plays an important role in maintaining the stemness characteristics of GCSCs.