The Molecular Mechanism of Gypenosides-induced Apoptosis in Human Hepatoma Cells
| Autor: | Qwa-Fun Wang, 王貴芳 |
|---|---|
| Rok vydání: | 2002 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 90 Gypenosides are triterpenoid saponins contained in an extract from Gynostemma pentaphyllum Makino and reported to induce apoptosis inhuman hepatoma cells. However, the molecular mechanism underlying the Gypenosides-induced apoptotic process is unclear. To understand the molecular mechanism of Gypenosides-inducced apoptosis, we had detected the caspase cascade, Bcl-2 family, Fas/Fas Ligand system, and p53 gene regulation. In this study, we have observed that treatment with Gypenosides resulted in a dose- and time-dependent decrease in cell viability. We found that Gypenosides-induced apoptosis in human hepatoma Huh-7, Hep3B and HA22T cell lines as evidenced by morphological changes, 4’, 6’- diamidino-2-phenylindole (DAPI) staining and in situ terminal transferase-mediated dUTP-fluorescensinnick end-labeling (TUNEL) assay. In Bcl-2 family, our data demonstrated that Gypenosides -induced apoptotic cell death was accompanied by up-regulation of Bax, Bak and Bcl-XL, and down-regulation of Bcl-2 and Bad, while it had no effect on the level of Bag-1 protein. In caspase cascade, moreover, Gypenosides treatment caused the release of mitochondrial cytochrome c to cytosol and sequential activation of caspases, including caspase-1, -9 and -3, then leading to cleavage of poly-ADP-ribose polymerase. Furthermore, the Gypenosides-induced apoptosis was markedly blocked by the broad-spectrum caspase inhibitor, z-VAD-fmk. In Fas/Fas Ligand system, our results showed that caspase-8 was not activated upon Gypenosides treatment, and Gypenosides did not affect the expression levels of Fas and Fas Ligand proteins, indicating that stimulation of Fas/Fas Ligand and activation of caspase-8 did not involve in Gyp-induced caspase-3 activation and apoptosis. In p53 gene regulation, no accumulation of wild-type p53 protein was detected upon treatment of HA22T cells with Gyp. In addition, cells lacking functional p53 (Hep3B and Huh-7 cells) exhibit a similar response to Gypenosides treatment as do cells with functional p53 (HA22T cells). These results suggest that Gypenosides-mediated up-regulation of Bax protein and apoptosis occurred independence of p53 protein.Taken together, these results suggest that treatment of human hepatoma cells with Gypenosides induced apoptosis through the up-regulation of Bax and Bak, and down-regulation of Bcl-2, release of mitochondrial cytochrome c and activation of caspase cascade. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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