链球菌蛋白诱导胶质母细胞瘤BT325细胞周期阻滞及凋亡的机制研究

doi:10.3969/j.issn.1673-8640.2014.04.015

检验医学 ›› 2014, Vol. 29 ›› Issue (4): 363-368.DOI: 10.3969/j.issn.1673-8640.2014.04.015

链球菌蛋白诱导胶质母细胞瘤BT325细胞周期阻滞及凋亡的机制研究

王艳¹，张力平²，陈辉²，曹宁²，朱俊萍²，平国玲²，李卫红³

1.北京积水潭医院检验科，北京 100035；2.首都医科大学病原生物学系，北京 100069；3. 首都医科大学细胞生物学系，北京 100069

收稿日期:2013-03-16 出版日期:2014-04-30 发布日期:2014-06-06
通讯作者: 张力平，联系电话:010-83911742。
作者简介:王艳，女，1981年生，硕士，医师，主要从事临床微生物学及免疫学检验工作。
基金资助:
北京市教育委员会科技发展计划项目(KM2004 10025008)

Research on the mechanisms of streptococcal protein inducing cycle arrest and apoptosis in glioblastoma BT325 cell

WANG Yan¹, ZHANG Liping², CHEN Hui², CAO Ning², ZHU Junping², PING Guoling², LI Weihong³

1. Department of Clinical Laboratory, Beijing Jishuitan Hospital, Beijing 100035,China;
2. Department of Pathogenic Biology, Capital Medical University, Beijing 100069,China;
3. Department of Cell Biology, Capital Medical University, Beijing 100069,China

Received:2013-03-16 Online:2014-04-30 Published:2014-06-06

摘要/Abstract

摘要：

目的　探讨链球菌蛋白诱导人脑多形性胶质母细胞瘤BT325细胞周期阻滞及凋亡的相关作用机制。方法　体外培养人脑多形性胶质母细胞瘤BT325细胞，噻唑蓝（MTT）法检测细胞增殖活性，倒置显微镜下观察细胞形态变化，原位染色后荧光显微镜下观察凋亡细胞，流式细胞术检测细胞周期、细胞凋亡率及线粒体膜电位（MMP），蛋白质印迹法（Western blot）检测细胞凋亡相关蛋白的表达。结果　链球菌蛋白（10～100 mg/L）可显著抑制细胞增殖，使细胞周期阻滞于G2/M期，呈时间和剂量依赖性；链球菌蛋白作用细胞24 h后，MMP显著下降（P＜0.01），凋亡细胞逐渐增多，48 h时各实验组凋亡率明显高于对照组（P＜0.01）；链球菌蛋白（50 mg/L）呈时间依赖性增强细胞P53蛋白表达，抑制B淋巴细胞瘤-2（Bcl-2）蛋白表达，促使细胞色素C（CytC）从线粒体进入胞质，从而激活半胱氨酸天冬氨酸蛋白酶3前体（Procaspase-3）。结论　链球菌蛋白可显著抑制人脑多形性胶质母细胞瘤BT325细胞增殖，影响细胞周期分布，通过线粒体途径诱导细胞发生凋亡，由此发挥其抗肿瘤活性作用。

关键词: 链球菌, 人脑多形性胶质母细胞瘤BT325细胞, 细胞周期, 细胞凋亡, 线粒体膜电位

Abstract:

Objective　To investigate the mechanisms of streptococcal protein inducing cycle arrest and apoptosis in human cerebral glioblastoma BT325 cell. Methods　Human cerebral glioblastoma BT325 cells were cultured in vitro， and methlcyclopentadienyl manganese tricarbonyl（MTT） assay was used to determine the proliferation activity of BT325 cells. The morphological changes of BT325 cells were observed by the phase-contrast microscopy. The apoptotic cells were observed after fluorescence staining. Flow cytometry was used to determine the cell cycle， the rate of cell apoptosis and the change of mitochondrial membrane potential（MMP）. The expression of apoptosis-related proteins was determined by Western blot. Results　Streptococcal protein（10-100 mg/L） inhibited the proliferation of BT325 cells significantly and induced G2/M phase arrest in time- and dose-dependent manners. MMP declined（P＜0.01）， and apoptotic cells gradually increased after BT325 cells were treated with streptococcal protein for 24 h. The rate of cell apoptosis in determination group was significantly higher than that in control group at 48h（P＜0.01）. In a time-dependent manner， streptococcal protein（50 mg/L） up-regulated the expression of cell P53 protein， down-regulated the expression of B-cell lymphoma-2（Bcl-2）， led to the release of mitochondrial cytochrome C（CytC） from mitochondria into cytoplasm， and then caused the activation of the precursor of cysteinyl aspartate specific proteinase 3（Procaspase-3）. Conclusions　Streptococcal protein may exert its anticancer activity through inhibiting the proliferation of human cerebral glioblastoma BT325 cells， effecting the distribution of cell cycle and inducing cell apoptosis by mitochondrial-dependent pathway.

Key words: Streptococcus, Human cerebral glioblastoma BT325 cell, Cell cycle, Cell apoptosis, Mitochondrial membrane potential

中图分类号:

R446.62

王艳，张力平，陈辉，曹宁，朱俊萍，平国玲，李卫红. 链球菌蛋白诱导胶质母细胞瘤BT325细胞周期阻滞及凋亡的机制研究[J]. 检验医学, 2014, 29(4): 363-368.

WANG Yan，ZHANG Liping，CHEN Hui，CAO Ning，ZHU Junping，PING Guoling，LI Weihong. Research on the mechanisms of streptococcal protein inducing cycle arrest and apoptosis in glioblastoma BT325 cell[J]. , 2014, 29(4): 363-368.

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链球菌蛋白诱导胶质母细胞瘤BT325细胞周期阻滞及凋亡的机制研究

Research on the mechanisms of streptococcal protein inducing cycle arrest and apoptosis in glioblastoma BT325 cell

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