Gypenosides induce apoptosis in human hepatoma Huh-7 cells through a calcium/reactive oxygen species-dependent mitochondrial pathway.
We have previously reported that gypenosides induce apoptosis in human hepatocarcinoma Huh-7 cells through a mitochondria-dependent caspase-9 activation cascade. In order to further explore the critical events leading to apoptosis in gypenosides-treated cells, the following effects of gypenosides on components of the mitochondrial apoptotic pathway were examined: generation of reactive oxygen species (ROS), alteration of the mitochondrial membrane potential (MPT), and the subcellular distribution of Bcl-2 and Bax. We show that gypenosides-induced apoptosis was accompanied by the generation of intracellular ROS, disruption of MPT, and inactivation of ERK, as well as an increase in mitochondrial Bax and a decrease of mitochondrial Bcl-2 levels. Ectopic expression of Bcl-2 or treatment with furosemide attenuated gypenosides-triggered apoptosis. Treatment with ATA caused a drastic prevention of apoptosis and the gypenosides-mediated mitochondrial Bcl-2 decrease and Bax increase, but failed to inhibit ROS generation and MPT dysfunction. Incubation with antioxidants significantly inhibited gypenosides-mediated ROS generation, ERK inactivation, MPT and apoptosis. Moreover, an increase of the intracellular calcium ion (Ca(2+)) concentration rapidly occurred in gypenosides-treated Huh-7 cells. Buffering of the intracellular Ca(2+) increase with a Ca(2+) chelator BAMTA/AM blocked the gypenosides-elicited ERK inactivation, ROS generation, Bcl-2/Bax redistribution, mitochondrial dysfunction, and apoptosis. Based on these results, we propose that the rise in intracellular Ca(2+) concentration plays a pivotal role in the initiation of gypenosides-triggered apoptotic death.
Wang
Qwa-Fun
QF
School of Post-baccalaureate Chinese Medicine, China Medical University, Taichung, Taiwan, Republic of China.
Chiang
Chi-Wu
CW
Wu
Chun-Chi
CC
Cheng
Chi-Chih
CC
Hsieh
Shur-Jong
SJ
Chen
Jung-Chou
JC
Hsieh
Yun-Chih
YC
Hsu
Shih-Lan
SL
D000972:Antineoplastic Agents, Phytogenic / Q000008:administration & dosage / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects*; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy / Q000473:pathology; D045744:Cell Line, Tumor / Q000187:drug effects; D029774:Gynostemma; D006801:Humans; D008113:Liver Neoplasms / Q000188:drug therapy / Q000473:pathology; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008517:Phytotherapy; D010936:Plant Extracts / Q000008:administration & dosage / Q000494:pharmacology* / Q000627:therapeutic use; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000378:metabolism
D000972:Antineoplastic Agents, Phytogenic; D010936:Plant Extracts; D019253:Proto-Oncogene Proteins c-bcl-2; D017382:Reactive Oxygen Species; D051028:bcl-2-Associated X Protein; C474918:gypenoside