miR-18a对结直肠癌SW116细胞辐射敏感性的影响

Effect of miR-18a on irradiation sensitivity of colorectal cancer SW116 cells

目的:探讨miR-18a与结直肠癌SW116细胞辐射敏感性之间的关系,阐明miR-18a影响细胞凋亡和自噬的可能机制。方法:人结直肠癌SW116细胞分为miR-18aNC组、miR-18amimic组、miR-18aNC+4Gy组和miR-18amimic+4Gy组。qRT-PCR法检测照射前后SW116细胞中miR-18a的表达;集落形成实验观察miR-18a对SW116细胞辐射敏感性的影响;GFPLC3形态学方法检测SW116细胞自噬率;流式细胞术检测SW116细胞凋亡率;采用生物信息学方法预测miR-18a的靶基因,以荧光素酶报告基因验证miR-18a与靶基因3′UTR结合;Western blotting法检测SW116细胞中共济失调-毛细血管扩张突变基因(ATM)蛋白表达。结果:与照射前比较,照射后SW116细胞中miR-18a表达水平明显降低(P<0.05)。集落形成实验,与miR-18aNC组比较,miR-18amimic组SW116细胞辐射敏感性增强(P<0.05),细胞凋亡率和自噬率明显增加(P<0.05)。与miR-18aNC+4Gy组比较,miR-18amimic+4Gy组SW116细胞中ATM蛋白表达减少。结论:miR-18a的靶基因为ATM;miR-18amimic可促进辐射诱导的细胞凋亡和自噬,且能增加结直肠癌SW116细胞辐射敏感性。

Objective To discuss the relationship between miR-18 aand the irradiation sensitivity of the colorectal cancer SW116 cells,and to elucidate the possible mechanism of the effects of miR-18 aon the apoptosis and autophagy of the cells.Methods The SW116 cells were divided into miR-18 aNC group,miR-18 amimic group,miR-18aNC＋4Gy group,and miR-18 a mimic＋4Gy group.The expression of miR-18 ain SW116cells was detected by qRT-PCR before and after irradition;colony-forming assay was used to detect the effect of miR-18 aon the irradiation sensitivity of the SW116cells;GFPLC3morphological assay was used to detect the autophagy rate of SW116cells;the apoptotic rate of SW116 cells was analyzed by flow cytometry;the target gene of miR-18 awas predicted by bioinformatics methods and dual-luciferase reporter assay was used to demonstrate the binding of miR-18 aand 3＇UTR targets;the expression of ataxia-telangiectasia mutated（ATM）protein in SW116 cells was measured by Western blotting method.Results compared with before irradation,the expression level of miR-18 a in the SW116 cells after irradation was decreased obviously（P〈0.05）.The colony-forming assay results showed that the irradiation sensitivity of the SW116 cells in miR-18 amimic group was increased compared with miR-18 aNC group（P〈0.05）,and the apoptotic rate and autophagy rate of the SW116 cells in miR-18 a mimic group were increased（P〈0.05）.Compared with miR-18aNC＋4Gy group,the expression level of ATM protein in miR-18 a mimic＋4Gy group was decreased.Conclusion ATM is the target gene of miR-18 ain the SW116 cells.miR-18 a mimic can increase the apoptosis and autophagy induced by irradition,and can increase the radiosensitivity of colorectal cancer SW116 cells.