Ji Yu-Bin, Qu Zhong-Yuan, Zou Xiang
Engineering Research Center of Natural Antineoplastic Drugs, Ministry of Education, Harbin 150076, Heilongjiang, China.
Exp Toxicol Pathol. 2011 Jan;63(1-2):69-78. doi: 10.1016/j.etp.2009.09.010. Epub 2009 Oct 7.
This study was designed to investigate the effect of juglone on the apoptosis of human gastric cancer SGC-7901 cells. The cytotoxic activity of juglone on SGC-7901 cells was tested by the sulforhodamine B (SRB) assay. The morphological changes in the cells were observed by transmission electron microscopy (TEM). The apoptotic rate, the level of reactive oxygen species (ROS), mitochondrial transmembrane potential and the expression of cytochrome c protein were detected by flow cytometry (FCM). The expression of Bcl-2 and Bax proteins were examined by Western blot. Caspase 3 activity was determined with a microplate reader. Our results were as follows: the GI(50) values for SGC-7901 cells were 36.51 ± 1.05 μmol/L (24h) and 25.37 ± 1.19 μmol/L (48 h). After 24h of exposure to juglone (5, 10, 15 and 20 μmol/L), the cells presented the typical morphological changes of apoptosis, and the rate of apoptosis was found to increase in a dose-dependent manner. After cells were treated with juglone at the same dose for 24h, the level of ROS was significantly higher, the expression of Bcl-2 was significantly down-regulated and the expression of Bax was significantly up-regulated compared to the control. The mitochondrial transmembrane potential was significantly lower, and the expression of the cytochrome c protein was significantly higher relative to the control. Caspase 3 was activated in a concentration-dependent manner. In conclusion, juglone can induce apoptosis in SGC-7901 cells through a mitochondrial pathway that seems to be mediated by the generation of ROS and a reduction in the Bcl-2/Bax ratio.
本研究旨在探讨胡桃醌对人胃癌SGC - 7901细胞凋亡的影响。采用磺酰罗丹明B（SRB）法检测胡桃醌对SGC - 7901细胞的细胞毒活性。通过透射电子显微镜（TEM）观察细胞的形态变化。采用流式细胞术（FCM）检测细胞凋亡率、活性氧（ROS）水平、线粒体跨膜电位及细胞色素c蛋白的表达。通过蛋白质免疫印迹法检测Bcl - 2和Bax蛋白的表达。用酶标仪测定Caspase 3活性。结果如下：SGC - 7901细胞的GI(50)值在24小时为36.51±1.05μmol/L，48小时为25.37±1.19μmol/L。用胡桃醌（5、10、15和20μmol/L）处理24小时后，细胞呈现典型的凋亡形态变化，且凋亡率呈剂量依赖性增加。用相同剂量的胡桃醌处理细胞24小时后，与对照组相比，ROS水平显著升高，Bcl - 2的表达显著下调，Bax的表达显著上调。线粒体跨膜电位显著降低，细胞色素c蛋白的表达相对于对照组显著升高。Caspase 3以浓度依赖性方式被激活。综上所述，胡桃醌可通过线粒体途径诱导SGC - 7901细胞凋亡，该途径似乎由ROS的产生和Bcl - 2/Bax比值的降低介导。
