Jiang Yuan, Sun Yan, Yuan Yuan
Department of Head and Neck Surgery, Jiangsu Cancer Hospital and Research Institute, Nanjing 210009, China.
Zhonghua Zhong Liu Za Zhi. 2012 Oct;34(10):734-8. doi: 10.3760/cma.j.issn.0253-3766.2012.10.004.
To investigate the new mechanism of temozolomide (TMZ) induced anti-tumor effects on glioblastoma cells in vitro.
Grade IV glioma cell lines SHG44 and U251 cells were treated with TMZ. MTT test was used to determine the proliferation of glioma cells. Hoechst 33342 assay was used to detect apoptosis in the tumor cells. The cell cycle progression was assessed by flow cytometry. The level of intracellular reactive oxygen species (ROS) was detected using DCFH-DA probe. real-time PCR assay and Western blotting were used to analyze the expression of SIRT1.
Treatment with TMZ for 72 hours inhibited cell proliferation (P < 0.05) and induced apoptosis in the two cell lines in a concentration-dependent manner. TMZ at 100 µmol/L significantly resulted in G(2)/M cell cycle arrest (66.16%, 69.65%), and triggered a robust increase in cell apoptosis [(33.4 ± 1.8)% and (26.8 ± 3.2)%]. TMZ remarkably increased reactive oxygen species (ROS) production (P < 0.05), indicating an overexpression of signal for SIRT1 activation.
Our findings suggest that temozolomide mediates anti-tumor effects on glioma cells in vitro via ROS-dependent SIRT1 signaling pathway, therefore, provide a theoretical evidence for a new approach to improve the treatment of glioma in future.
探讨替莫唑胺（TMZ）体外诱导胶质母细胞瘤细胞抗肿瘤作用的新机制。
用TMZ处理IV级胶质瘤细胞系SHG44和U251细胞。采用MTT试验测定胶质瘤细胞的增殖情况。用Hoechst 33342检测法检测肿瘤细胞凋亡。通过流式细胞术评估细胞周期进程。使用DCFH-DA探针检测细胞内活性氧（ROS）水平。采用实时PCR检测法和蛋白质免疫印迹法分析SIRT1的表达。
TMZ处理72小时可抑制两种细胞系的细胞增殖（P < 0.05），并以浓度依赖性方式诱导细胞凋亡。100 μmol/L的TMZ显著导致G(2)/M期细胞周期阻滞（66.16%，69.65%），并引发细胞凋亡显著增加[（33.4 ± 1.8）%和（26.8 ± 3.2）%]。TMZ显著增加活性氧（ROS）的产生（P < 0.05），表明SIRT1激活信号的过表达。
我们的研究结果表明，替莫唑胺通过ROS依赖的SIRT1信号通路介导体外对胶质瘤细胞的抗肿瘤作用，因此，为未来改善胶质瘤治疗的新方法提供了理论依据。
