Zhang Lei, Liang Peng, Zhang Rui
Department of Neurosurgery, Harbin Medical University Cancer Hospital, Harbin, Heilongjiang, China (mainland).
Department of Neurosurgery, Harbin Medical University Cancer Hospital, Harbin, China (mainland).
Med Sci Monit. 2015 Nov 23;21:3629-33. doi: 10.12659/msm.894006.
BACKGROUND Most mitochondria-mediated apoptosis has some relevance to the cell cycle, but there is still a lack of investigations about U251 cell cycle in human brain glioma cells. In this study, we aimed to clarify the correlation of mitochondria-mediated apoptosis with the U251 cell cycle and its influence on apoptosis, through observing the impact of mitochondria-mediated apoptosis in U251cell specificity cycle arrest and Caspase activation. MATERIAL AND METHODS AnnexinV/PI and API were used to label the brain glioma cells for flow cytometry analysis of U251 cell apoptosis and cell cycle. RT-PCR and Western blot were performed to detect Caspase-3 and Caspase-9 activation. RESULTS Peripheral blood in stationary phase is not sensitive to apoptosis induction, but U251 cells have obvious apoptosis. Mitochondria-mediated apoptosis mainly occurs in the G1 phase of the cell cycle. Caspase-3 and Caspase-9 mRNAs and proteins expression increased significantly after the cells were treated by mitochondrial apoptosis-related gene Bax induction. CONCLUSIONS Mitochondria-mediated apoptosis is related to the U251 cell cycle with specific G1 stage arrest. Caspase activation occurs in the process of cell apoptosis.
大多数线粒体介导的细胞凋亡与细胞周期存在一定关联,但在人脑胶质瘤细胞中,关于U251细胞周期的研究仍较为缺乏。在本研究中,我们旨在通过观察线粒体介导的凋亡对U251细胞特异性周期阻滞和半胱天冬酶激活的影响,阐明线粒体介导的凋亡与U251细胞周期的相关性及其对细胞凋亡的影响。
采用膜联蛋白V/碘化丙啶(AnnexinV/PI)和细胞周期蛋白依赖性激酶抑制因子(API)标记脑胶质瘤细胞,用于U251细胞凋亡和细胞周期的流式细胞术分析。进行逆转录聚合酶链反应(RT-PCR)和蛋白质免疫印迹法(Western blot)检测半胱天冬酶-3(Caspase-3)和半胱天冬酶-9(Caspase-9)的激活情况。
静止期外周血对凋亡诱导不敏感,但U251细胞有明显凋亡。线粒体介导的凋亡主要发生在细胞周期的G1期。经线粒体凋亡相关基因Bax诱导处理细胞后,Caspase-3和Caspase-9的mRNA及蛋白表达显著增加。
线粒体介导的凋亡与U251细胞周期相关,具有特异性的G1期阻滞。半胱天冬酶激活发生在细胞凋亡过程中。