MiR-551b-3p靶向USP9X对人肺癌A549细胞凋亡和放射敏感性的影响

目的:探讨miR-551b-3p对肺癌A549细胞凋亡和放射敏感性的影响及分子机制。方法:将miRNC,miR-551b-3p,anti-miR-NC,anti-miR-551b-3p,si-NC,si-USP9X分别转染至A549中,记为miR-NC组、miR-551b-3p组、anti-miR-NC组、anti-miR-551b-3p组、si-NC组、si-USP9X组;将miR-551b-3p分别与pc DNA和pc DNA-USP9X共转染至A549中,记为miR-551b-3p+pc DNA组、miR-551b-3p+pc DNA-USP9X组。采用实时荧光定量PCR(RT-q PCR)检测miR-551b-3p和USP9X的表达水平;蛋白质印迹法检测X染色体连锁的泛素特定蛋白水解酶9(ubiquitin-specific peptidase 9 X-linked,USP9X)、 B细胞淋巴瘤/白血病-2 (B-cell lymphoma/leukemia-2, Bcl-2)、 Bcl-2相关X蛋白(Bcl-2 related X protein,Bax)的表达;荧光素酶报告实验检测miR-551b-3p和USP9X的靶向关系;流式细胞术检测细胞凋亡;细胞克隆集落形成实验检测放射敏感性。结果:肺癌组织和肺癌细胞A549中miR-551b-3p表达水平显著降低,USP9X mRNA和蛋白质表达水平显著升高(均P<0.001)。miR-551b-3p靶向调控USP9X,过表达miR-551b-3p和抑制USP9X表达,细胞凋亡率显著升高,Bcl-2表达水平显著降低,Bax表达水平显著升高,细胞的存活曲线明显下移(均P<0.001)。USP9X过表达逆转了miR-551b-3p过表达对肺癌A549细胞凋亡和放射敏感性的作用。结论:过表达miR-551b-3p促进肺癌A549细胞凋亡,增强肺癌细胞放射敏感性,其机制可能与USP9X有关。

Effect of miR-551b-3p targeting USP9X on apoptosis and radiosensitivity of human lung cancer A549 cells

Objective:To investigate the effect of miR-551b-3p on apoptosis and radiosensitivity of human lung cancer A549 cells and its molecular mechanism.Methods:miR-NC,miR-551b-3p,anti-miR-NC,anti-miR-551b-3p,si-NC,and si-USP9X were transfected into A549,respectively,and recorded as miR-NC group,miR-551b-3p group,antimiR-NC group,anti-miR-551b-3p group,si-NC group and si-USP9X group;miR-551b-3p was co-transfected with pcDNA and pcDNA-USP9X into A549,respectively,recorded as miR-551b-3p+pcDNA group and miR-551b-3p+pcDNA-USP9X group.Real-time quantitative PCR(RT-qPCR)was used to detect the expressions of miR-551b-3p and USP9X;Western blotting was used to detect ubiquitin-specific peptidase 9 X-linked(USP9X),B-cell lymphoma/leukemia-2(Bcl-2),Bcl-2 related X protein(Bax)protein expression;luciferase reporter assay detects the targeting relationship of miR-551b-3p and USP9X;flow cytometry detects apoptosis;cell clone colony formation experiments detect radiosensitivity.Results:The expression of miR-551b-3p was significantly decreased in lung cancer tissues and lung cancer cell A549,and the expressions of USP9X mRNA and protein were significantly increased(all P<0.001).MiR-551b-3p targeted USP9X,overexpressed miR-551b-3p and inhibited USP9X expression,apoptosis rate was significantly increased,Bcl-2 expression was significantly decreased,Bax expression was significantly increased,and cell survival curve was significantly shifted down(all P<0.05).Overexpression of USP9X reversed the effect of miR-551b-3p overexpression on apoptosis and radiosensitivity of lung cancer A549 cells.Conclusion:Overexpression of miR-551b-3p promotes apoptosis of lung cancer A549 cells and enhances radiosensitivity of lung cancer cells.The mechanism may be related to the USP9X.