circ_0001946靶向miR-1270/Cyclin T2轴调控胃癌细胞恶性表型的分子机制研究

Molecular Mechanism of Circ_0001946 Targeting MiR-1270/Cyclin T2 Axis in Regulating Malignant Phenotype of Gastric Cancer Cells

目的探讨circ_0001946对胃癌细胞恶性表型的影响及可能机制。方法采用qRT-PCR法检测胃癌组织和细胞系(HGC-27、AGS、MKN45)中circ_0001946和miR-1270表达情况,并采用Pearson相关性分析探讨胃癌组织中circ_0001946和miR-1270表达的相关性。分别转染circ_0001946小干扰RNA、miR-1270模拟物或共转染circ_0001946小干扰RNA与miR-1270抑制剂至胃癌HGC-27细胞,采用CCK-8法和克隆形成实验检测细胞增殖情况,采用流式细胞术检测细胞凋亡情况,采用Transwell法检测细胞侵袭情况,采用蛋白免疫印迹法检测细胞周期蛋白T2(Cyclin T2)表达情况。行双荧光素酶报告基因实验验证circ_0001946、miR-1270和Cyclin T2调控的关系。结果胃癌组织和细胞系中circ_0001946表达升高(P<0.05),而miR-1270表达降低(P<0.05),且胃癌组织中circ_0001946与miR-1270表达呈负相关(r=-0.752,P<0.05)。敲减circ_0001946过表达后HGC-27细胞光密度(OD)值、克隆形成数和侵袭数降低(P<0.05),而凋亡率升高(P<0.05)。敲减miR-1270过表达后可升高HGC-27细胞OD值、克隆形成数和侵袭数(P<0.05),降低凋亡率(P<0.05)。circ_0001946可竞争性结合miR-1270,Cyclin T2是miR-1270的靶基因。结论circ_0001946在胃癌组织和细胞系中表达升高,其可能通过靶向miR-1270/Cyclin T2轴促进胃癌细胞增殖和侵袭,并诱导胃癌细胞凋亡,有可能成为胃癌治疗的分子靶点。

Objective To explore the effect of circ_0001946 on malignant phenotype of gastric cancer cells and its potential mechanism.Methods Expressions of circ_0001946 and miR-1270 in gastric cancer tissues and cell lines(HGC-27,AGS,MKN45)were detected by quantitative reverse transcription-polymerase chain reaction(qRT-PCR),and the correlation between circ_0001946 with miR-1270 expressions in gastric cancer tissues was detected by Pearson correlation analysis.The respective transfection of circ_0001946 siRNA and miR-1270 mimics or cotransfection of circ_0001946 siRNA and miR-1270 inhibitor were performed into gastric cancer HGC-27 cells.Condition of cell proliferation was detected by CCK-8 method and colony formation assay,and apoptosis condition was detected by flow cytometry.Condition of cell invasion was detected by Transwell method,and the Cyclin T2 expression was detected by Western blot method.Double luciferase reporter gene assay was used to verify regulated correlations among circ_0001946 and miR-1270 with Cyclin T2.Results The circ_0001946 expressions were significantly increased(P<0.05),while miR-1270 expressions were significantly decreased in gastric cancer tissues and cell lines(P<0.05),And circ_0001946 expression was negatively correlated with the expression of miR-1270 in gastric cancer tissues(r=-0.752,P<0.05).After knockdown of overexpressed circ_0001946 miR-1270,values of optical density(OD)value,clone formation number and invasion number of HGC-27 cells were decreased(P<0.05),while apoptosis rate was significantly increased(P<0.05).After knockdown of overexpressed miR-1270,values of cell value,clone formation number and invasion number of HGC-27 cells were significantly increased(P<0.05),while apoptosis rate was significantly decreased(P<0.05).The circ_0001946 could competitively adsorb miR-1270,and Cyclin T2 was the target gene of miR-1270.Conclusion The circ_0001946 expressions are increased in gastric cancer tissues and cell lines.It may promote the proliferation and invasion of gastric cancer cells by targeting miR-1270/Cyclin T2 axis,and induce apoptosis of gastric cancer cells.Therefore,circ_0001946 may become a molecular target for the treatment of gastric cancer.