百里醌对人胶质瘤细胞凋亡的作用及其机制研究

目的 分析百里醌对胶质瘤U87细胞生长抑制和凋亡诱导的功能.方法 体外胶质瘤细胞株U87以及人星形胶质细胞株NHA中添加不同浓度百里醌后,CCK-8法检测细胞活力;克隆形成实验观察U87细胞形成细胞克隆的能力;流式细胞术检测细胞周期和ROS含量;Hoechst染色法和流式细胞术测定细胞凋亡;流式细胞术(JC-1荧光染色...

Apoptosis induction and underlying mechanisms of thymoquinone on human glioma cells

Objective To explore the apoptosis induction and underlying molecular mechanisms of thymoquinone on human glioma cells.Methods Human glioma cell line U87 and normal human astrocytes NHA were treated with thymoquinone in vitro.CCK-8 and cell clone formation assays were employed to detect the cell viability and proliferation activity in vitro.The cell cycle and production of ROS were analyzed by flow cytometry.Hoechst 33342 staining and Annexin V/PI apoptosis assays were utilized to explore the apoptosis of U87 cells.The mitochondrial membrane potential of U87 cell was observed by JC-1 staining.The cell cycle regulating proteins(Cyclin B1,Cdc2,Cdc25 c,p53 and p73),cell apoptosis related factors(Fas-L,Fas,Bax,tBid,PARP,caspase 9 and caspase 3) as well as DNA damage signaling factors(p-ATM,p-ATR,γ-H2 AX,p-Chk1,p-Chk2 and p-p53) were determined by western blot.Results Thymoquinone treatment suppressed the viability and proliferation of U87 cells in a concentration-dependent manner,and the levels of p-ATM,p-ATR,γ-H2 AX,p-Chk1,p-Chk2 and p-p53 were all increased by the treatment of thymoquinone.Thymoquinone also increased cell arrest in the G2/M phase by decreasing Cyclin B1,Cdc2,and Cdc25 c,and up-regulating p53 and p73.Thymoquinone induced U87 cell apoptosis through the up-regulation of Fas-L,Fas,and Bax as well as the proteolysis of PARP,caspase 9 and caspase 3.Thymoquinone also contributed to the loss of mitochondrial membrane potential.The apoptotic cell death induced by thymoquinone was inhibited by pretreatment with Z-VAD-FMK,a pan-caspase inhibitor.Moreover,the apoptotic effect of thymoquinone was reactive oxygen species(ROS)-dependent,as evidenced by the inhibition of thymoquinone-induced PARP cleavage and ROS generation via N-acetylcysteine(NAC)-induced ROS scavenging.Conclusion Thymoquinone exerts a promotive effect on the apoptosis of human glioma cells through the ROS pathway.