miR-485-3p通过调控ATR信号通路增强MGC803细胞放射敏感性研究

摘要

Objective To investigate the effect of miR-485-3p on the radiosensitivity of gastric carcinoma MGC803 cells and the possible mechanism of action.Methods The MGC803 cells were transfected with miR-485-3p mimic or ATR siRNA and then treated by radiation.The MTT method,colony-forming assay,and apoptosis test were used to measure the change in radiosensitivity of such cells.RT-PCR and Western blot were used to measure the changes in the expression of miR-485-3p and ATR,and DIANA,TargetScan,and miRanda software and dual-luciferase reporter assay were used to verify the targeted effect of miR-485-3p on ATR.Results After radiation treatment,the expression of miR-485-3p in gastric carcinoma cells was downregulated.The overexpression of miR-485-3p reduced the proliferative capacity and colony-forming ability of cells,increased apoptosis rate,and thus increased radiosensitivity.The software for target gene prediction found that ATR might be the target gene of miR-485-3p,and the dual-luciferase reporter assay further confirmed that ATR was the direct target of miR-485-3p.The miR-485-3p downregulated the expression of ATR,and the inhibition of the ATR signaling pathway by transfection with ATR siRNA increased the radiosensitivity of gastric carcinoma cells.Conclusions The miR-485-3p may target at ATR and regulate the radiosensitivity of gastric carcinoma cells through inhibiting the ATR signaling pathway.%目的 研究miR-485-3p对胃癌细胞MGC803放射敏感性影响及其可能作用机制.方法 将miR-485-3p mimic或ATR siRNA转染MGC803细胞后X线照射,MTT、克隆形成实验和凋亡实验观察细胞生物效应(表达检测照射6 Gy,克隆形成实验0、2、4、6、8、10 Gy).分别用RT-PCR和蛋白印迹法检测miR-485-3p和ATR表达水平,DIANA、TargetScan和miRanda软件预测和双荧光素酶实验验证miR-485-3p对ATR靶向作用.结果 MGC803细胞放射后miR-485-3p表达下调.miR-485-3p过表达降低了细胞增殖和存活分数,增加了细胞凋亡率.经靶基因预测软件发现ATR可能是miR-485-3p靶基因,双荧光素酶实验进一步验证了ATR是miR-485-3p直接靶点.miR-485-3p负调控ATR表达,转染ATR siRNA抑制ATR信号通路后放射敏感性升高.结论 miR-485-3p可能靶向ATR,通过抑制ATR信号通路活性调节MGC803细胞放射敏感性.