CUDC-907诱导胶质瘤细胞DNA损伤与自噬的实验研究

CUDC-907 induces DNA damage and autophagy in glioma cells

目的:探讨新型组蛋白去乙酰化酶(histone deacetylase,HDAC)/磷脂酰肌醇3-激酶(phosphatidyl-inositol 3-kinase,PI3K)双靶点抑制剂CUDC-907对人胶质瘤U251细胞DNA损伤、细胞周期及自噬的影响。方法:采用不同浓度的CUDC-907处理U251细胞24 h,MTT法检测细胞活力的变化;激光共聚焦显微镜观察DNA损伤标志物γ-H2AX在细胞内的分布;流式细胞术分析CUDC-907对细胞周期的影响;Western blot实验检测细胞内相关蛋白表达水平的变化。结果:CUDC-907能够抑制U251细胞活力。在CUDC-907处理的细胞中,蛋白激酶B(PKB)/Akt和p70核糖体蛋白S6激酶(p70s6K)的磷酸化水平降低,γ-H2AX的焦点数量与蛋白表达显著升高(P <0.05);U251细胞经CUDC-907作用后,G2/M期细胞数量增多;Western blot实验结果表明,CUDC-907促进p21的表达,同时抑制细胞周期素B1(cyclin B1)的蛋白表达和细胞分裂周期蛋白2(Cdc2)的磷酸化水平(P <0.05);另外,CUDC-907能够诱导细胞自噬,抑制自噬可促进CUDC-907诱导的DNA损伤。结论:CUDC-907能够显著抑制PI3K/Akt信号通路,诱导胶质瘤细胞发生DNA损伤,并阻滞细胞于G_2/M期,同时可诱导胶质瘤细胞发生保护性自噬。

AIM:To investigate the effect of CUDC-907,a dual histone deacetylase(HDAC)and phosphatidylinositol 3-kinase(PI3K)inhibitor,on the DNA damage,cell cycle distribution and autophagy in human glioma U251 cells.METHODS:U251 cells were treated with CUDC-907 of different concentrations,and the cell viability was detected by MTT assay.The quantitative γ-H2AX foci were determined by laser scanning confocal microscopy.The cell cycle distribution of U251 cells was examined by flow cytometry.The protein expression was determined by Western blot analysis.RESULTS:CUDC-907 inhibited the cell viability and the phosphorylation of Akt and p70 ribosomal protein S6 kinase(p70s6K)in the U251 cells(P<0.05).In CUDC-907-treated cells,the number of γ-H2AX foci and protein expression of γ-H2AX were increased significantly(P<0.05).CUDC-907 also induced cell arrest in the G2/M phase by up-regulating the expression of p21,and inhibiting the protein level of cyclin B1 and the phosphorylation of cell division cycle protein 2(Cdc2).In addition,CUDC-907 triggered cell autophagy,and inhibition of autophagy increased CUDC-907-induced DNA damage of U251 cells.CONCLUSION:CUDC-907 significantly inhibits PI3K/Akt signaling pathway,induces DNA damage and arrests cell cycle in G2/M phase.Blockage of autophagy promotes CUDC-907-induced DNA damage of U251 cells.