Gambogenic acid induced mitochondrial-dependent apoptosis and referred to phospho-Erk1/2 and phospho-p38 MAPK in human hepatoma HepG2 cells.
ABSTRACT Gambogenic acid, identified from Gamboge, is responsible for anti-tumor effects, and has been shown to be a potential molecule against human cancers. In this study, the molecular mechanism of gambogenic acid-induced apoptosis in HepG2 cells was investigated. Gambogenic acid significantly inhibited cell proliferation and induced apoptosis. Acridine orange/ethidium bromide (AO/EB) staining was used to observe apoptosis, and then confirmed by transmission electron microscopy. Gambogenic acid induced apoptosis and morphological changes in mitochondria, and intracellular reactive oxygen species (ROS) and mitochondrial membrane permeabilization (MMP) in mitochondrial apoptosis pathway were also examined. Results showed that the levels of phospho-p38 and its downstream phospho-Erk1/2 of HepG2 cells increased in time- and concentration-dependent manners after gambogenic acid treatments. Additionally, gambogenic acid increased expression ratio of Bcl-2/Bax in mRNA levels, Western blotting analysis also further confirmed the reduced level of Bcl-2 and increase the expression level of Bax in HepG2 cells. These results indicated that gambogenic acid induced mitochondrial oxidative stress and activated caspases through a caspase-3 and caspase-9-dependent apoptosis pathway. Moreover, gambogenic acid mediated apoptosis and was involved in the phospho-Erk1/2 and phospho-p38 MAPK proteins expression changes in HepG2 cells.
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ABSTRACT: OBJECTIVE: Gambogic acid (GBA) targeted Heat shock protein 90 (Hsp90) and prohibited TNF-α/NF-κB signaling pathway. It can be inferred that the anti-inflammatory activity of GBA results from inhibiting the cytokine production via NF-κB signaling pathway. We used the RAW264.7 cell line and the endotoxin shock mouse model to confirm the hypothesis that GBA protects mice from endotoxin shock by suppressing cytokine synthesis. METHOD: RAW264.7 cells were cultured and the endotoxin shocked mice model was constructed. ELISA was employed to evaluate the change of cytokine secretion levels. The effects of GBA on the activation of NF-κB signaling pathway were also determined by western blot and immune-fluorescent analysis. Cell viability was determined by MTT assay, and the cell migration was tested by wound healing assay. RESULT: Our results demonstrated that GBA significantly inhibited the LPS-induced release of pro-inflammatory factors both in cell lines and mice serum, thereby protecting mice from endotoxin shock. Furthermore, we observed that the reduction of inflammatory cytokines interleukin 1-beta, interleukin 6 and TNF-α resulted from the Hsp90's client protein IKK degradation and the suppression of NF-κB pathway. Moreover, GBA suppressed the migration of LPS-induced RAW264.7 cells. CONCLUSION: Our results indicate that GBA has a potential both as an antitumor and anti-inflammatory therapeutic agent.Agents and Actions 10/2012; · 1.59 Impact Factor