Acceleration of chrysin on apoptosis of hepatoma cell lines HepG2 induced by TNF-α
- Author:
Xin LI
1
Author Information
1. Institute of Toxicology
- Publication Type:Journal Article
- Keywords:
Apoptosis;
Chrysin;
Tumor necrosis factor (TNF)
- From:
Chinese Traditional and Herbal Drugs
2010;41(11):1828-1834
- CountryChina
- Language:Chinese
-
Abstract:
Objective: To research whether chrysin can sensitize the cell death of hepatoma (HepG2) cell lines induced by TNF-α and explore the molecular mechanism of this sensitization. Methods: HepG2 Cells were pretreated with designed dose of chrysin (10, 20, and 40 μmol/L) for 2 h, then followed TNF-α (10 ng/mL) for 24 h, the morphologic changes were observed under inversed microscope and the percentage of sub-G1 was measured using flow cytometry, the proprotein and cleavage of caspase-3, caspase-8, and PARP, regarded as the protein mark of apoptosis induced by TNF-α, were determined by Western blotting; After treating with chrysin for different times, the time course of apoptosis inhibitory protein, such as Bcl-xL, and cIAPs, xIAP, and cFLIP, were also detected using Western blotting. Results: The cell death increasing was observed in the group with combination of chrysin and TNF-α, but no obvious cell death could be found in chrysin, TNF-α alone, and control groups. The data of sub-G1 supported this result that, with the enhancement of pretreatment dose of chrysin, the percentage of sub-G1 reached (27.84±0.54)%. There was significant difference between the combined group and the control one (1.52±0.13)% (P<0.05) and obvious dose-dependent effect could be found. But the percentages of subG1 in chrysin and TNF-α alone groups were all under 5%, and there were no significant differences compared to the control group (P>0.05). Chromatin condensation which was the indication of apoptosis could be observed when the cells were stained by Hochest 33342; The proprotein of caspase-3inverted commascaspase-8, and PARP degraded, their cleavage appeared, and there were dose-dependent effects; The pan-caspase inhibitor z-VAD-fmk could inhibit the apoptosis of HepG2 cells which were treated with the combination of chrysin and TNF-α, according to the percentage of sub-G1 and the activation of caspase-3, caspase-8, and PARP; The apoptosis inhibitory protein cFLIP-L could be observed the down-regulation compared to the control group in a time-dependent way. Other inhibitory protein, such as Bcl-xL, xIAP did not change too much. Conclusion: Chrysin could accelerate the apoptosis induced by TNF-α, the down-regulation of apoptosis inhibitory protein cFLIP-L, which is regulated by NF-κB and augmented by TNF-α, and plays an important role in the acceleration of apoptosis.
