Biochemical and Biophysical Research Communications
Molecular mechanisms of echinocystic acid-induced apoptosis in HepG2 cells
Section snippets
Materials and methods
Materials and cell culture. Fetal bovine serum (FBS) was from Equitech-Bio. 3,3′-Dihexyloxacarbocyanine iodide (DiOC6(3)) was from Molecular Probes. The antibodies against poly(ADP)ribose polymerase (PARP), Bid, caspase-3, -8, and -9, JNK, p38, Phos-SAPK/JNK, Phos-p38, Phos-c-Jun, and α-tubulin were from Cell Signaling Technology. Cytochrome c and Bcl-2 antibodies were obtained from Santa Cruz. Caspase-8 (IETD-CHO) and caspase-9 (LEHD-CHO) inhibitors were from Calbiochem. Caspase-3
EA inhibits the proliferation of HepG2 cells through the induction of apoptosis
To test the effect of EA on the proliferation of HepG2 cells, the cells were treated with different concentrations of EA. After 24 h culture, the survival rate of HepG2 cells was investigated by a MTT assay. As shown in Fig. 1, EA inhibited cell proliferation in a dose-dependent manner with an IC50 (50% inhibitory concentration) value of 45.4 μM at 24 h treatment. To elucidate whether EA inhibits the proliferation of HepG2 cells by inducing apoptosis, we confirmed the apoptotic characterizations
Discussion
Apoptosis is a tightly regulated progress under the control of several signaling pathways, such as caspase and mitochondrial pathways [2], [3], [32]. In the present study, EA treatment caused activation of caspase-3, -8, and -9 in a time–response manner that is consistent with the results of PARP inactivation and DNA fragmentation. Moreover, a time-dependent release in mitochondrial cytochrome c and a concomitant increase in cytosolic cytochrome c were also observed in EA-treated HepG2 cells.
Acknowledgment
This work was partly supported by Cooperation of Innovative Technology and Advanced Research in Evolutional Area (CITY AREA).
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