Tetrandrine-induced apoptosis in rat primary hepatocytes is initiated from mitochondria: caspases and endonuclease G (Endo G) pathway.
Tetrandrine, a bisbenylisoquinoline alkaloid isolated from the dried root of Stephenia tetrandra (S Moore), possesses a remarkable pharmacological profile. However, the mechanisms of tetrandrine hepatotoxicity remain to be elucidated. In this study, we first proved apoptosis and mitochondrial dysfunction induced by tetrandrine in Sprague-Dawley rat liver in vivo. By further assuming apoptosis as an important mechanism in tetrandrine-induced hepatotoxicity, we focused on mitochondria-initiated apoptosis in primary hepatocytes isolated from Sprague-Dawley male rats. Tetrandrine treatment led to significant release of cytochrome c and downregulation of Bcl-X(L) accompanied by caspase 3 activation, and ultimately, DNA fragmentation. Caspase 3 activation was markedly inhibited by cyclosporin A (CsA) and Ac-DEVD-CHO. Furthermore, Endo G, a caspase-independent apoptotic protein, was detected for its expression and DNase activity. CsA blocked the release both of Endo G and cytochrome c significantly. Additionally, the generation of reactive oxygen species (ROS) increased in a time-dependent manner corresponding with a fall in intracellular GSH content after 10 microM tetrandrine treatment in 4h. Tetrandrine also induced mitochondrial dysfunction indicated by transition of mitochondrial transmembrane potential and decrease of intracellular ATP level. The findings indicated that the caspase-dependent mitochondrial apoptosis pathway was primarily involved in tetrandrine-induced apoptosis in rat primary hepatocytes. In addition, a caspase-independent pathway indicated by Endo G also contributed to apoptosis caused by tetrandrine. Meanwhile, ROS was proved an important inducer in this apoptosis process.
Yan
Cai
C
State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 201203, China.
Xin-Ming
Qi
Q
Li-Kun
Gong
G
Lin-Lin
Liu
L
Fang-Ping
Chen
C
Ying
Xiao
X
Xiong-Fei
Wu
W
Xiang-Hong
Li
L
Jin
Ren
R
D000470:Alkaloids / Q000009:adverse effects*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D044182:Benzylisoquinolines / Q000009:adverse effects*; D015153:Blotting, Western; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D045304:Cytochromes c / Q000096:biosynthesis; D004365:Drugs, Chinese Herbal / Q000009:adverse effects; D004591:Electrophoresis, Polyacrylamide Gel; D004706:Endodeoxyribonucleases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects*; D020287:In Situ Nick-End Labeling; D007425:Intracellular Membranes / Q000187:drug effects; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D008930:Mitochondria, Liver / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D008933:Mitochondrial Swelling / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D051020:bcl-X Protein / Q000096:biosynthesis
D000470:Alkaloids; C494813:Bcl2l1 protein, rat; D044182:Benzylisoquinolines; D061945:Caspase Inhibitors; D004365:Drugs, Chinese Herbal; D004791:Enzyme Inhibitors; D017382:Reactive Oxygen Species; D051020:bcl-X Protein; C009438:tetrandrine; D045304:Cytochromes c; D004706:Endodeoxyribonucleases; C052391:endonuclease G; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases