Open AccessApoptosis initiated by carbon tetrachloride in mitochondria of rat primary cultured hepatocytes
Author(s) -
CAI Yan,
GONG Likun,
QI Xinming,
LI Xianghong,
REN Jin
Publication year - 2005
Publication title -
acta pharmacologica sinica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.514
H-Index - 90
eISSN - 1745-7254
pISSN - 1671-4083
DOI - 10.1111/j.1745-7254.2005.00143.x
Subject(s) - dna fragmentation , apoptosis , malondialdehyde , carbon tetrachloride , cytochrome c , glutathione , thiobarbituric acid , chemistry , viability assay , fragmentation (computing) , lipid peroxidation , oxidative stress , biochemistry , mitochondrion , ccl4 , microbiology and biotechnology , lactate dehydrogenase , biology , programmed cell death , enzyme , ecology , organic chemistry
Aim: To investigate the mitochondria‐initiated apoptosis pathway involved in Carbon tetrachloride (CCl4) hepatotoxicity in vitro . Methods: Several cytotoxicity endpoints, including WST‐8 metabolism, lactate dehydrogenase leakage and morphological changes, were examined. The 5,5′‐dithio‐bis(2‐nitrobenzoic acid) reaction was used to measure reduced glutathione level, and the malondialdehyde level was determined using the thiobarbituric acid assay. The release of cytochrome c and Bcl‐XL was detected by Western blot. Caspase‐3 activity was measured using the fluorogenic substrate Ac‐DEVD‐AMC. DNA fragmentation was used to evaluate cell apoptosis. Results: A time‐and dose‐dependent decrease in cellular glutathione content was observed, along with a concomitant increase in malondialdehyde levels following the application of CCl 4 . Caspase 3 activity was stimulated at all doses of CCl 4 , with the most significant activation at 3 mmol/L. Cytochrome c was released obviously after CCl 4 treatment. A time‐dependent decrease in Bcl‐XL expression was observed. DNA fragmentation results revealed apoptosis and necrosis following CCl 4 treatment. Conclusion: Oxidative damage is one of the essential mechanisms of CCl 4 hepatotoxicity, which triggers apoptosis via the mitochondria‐initiated pathway.