Gambogenic acid induced mitochondrial-dependent apoptosis and referred to Phospho-Erk1/2 and Phospho-p38 MAPK in human hepatoma HepG2 cells

Key Laboratory of Xin'an Medicine, Ministry of Education, Anhui Province Key Laboratory of R&D of Chinese Medicine, Anhui University of Traditional Chinese Medicine, Hefei 230038, China.
Environmental toxicology and pharmacology 03/2012; 33(2):181-90. DOI: 10.1016/j.etap.2011.12.006
Source: PubMed


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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    • "Gambogenic acid is responsible for antitumor effects, and was found in Gamboge. In recent years, potent anticancer activity has also been discovered in gambogenic acid, which can inhibit the growth of various types of human cancer cell lines in both in vitro and in vivo modes, such as human lung adenocarcinoma (A549), hepatoma (H-22), human nasopharyngeal carcinoma cell line (CNE-1) and human gastric carcinoma (BGC-823), but since some normal cells (HUVEC) are not sensitive to it (Qu et al., 1991; Yan et al., 2011; Li et al., 2010), the induction of apoptosis has been proved to be a possible mechanism (Cheng et al., 2011; Chen et al., 2012; Yan et al., 2012; Yu et al., 2012). Earlier publications have described methods for GNA determination in plasma using HPLC-UV (Huang et al., 2013; Wang et al., 2012). "
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    ABSTRACT: A highly sensitive and rapid ultra-high-performance liquid chromatography–tandem mass spectrometry method was developed and validated for the determination of gambogenic acid in dog plasma. Gambogic acid was used as an internal standard (IS). After a simple liquid–liquid extraction by ethyl acetate, the analyte and internal standard were separated on an Acquity BEH C18 (100 × 2.1 mm, 1.7 µm; Waters ) column at a flow rate of 0.2 mL/min, using 0.1% formic acid–methanol (10:90, v/v) as mobile phase. Electrospray ionization source was applied and operated in the positive ion mode. Multiple reaction monitoring mode with the transitions m/z 631.3 → 507.3 and m/z 629.1 → 573.2 was used to quantify gambogenic acid and the internal standard, respectively. The calibration curves were linear in the range of 5–1000 ng/mL, with a coefficient of determination (r) of 0.999 and good calculated accuracy and precision. The low limit of quantification was 5 ng/mL. The intra-and inter-day precisions (relative standard deviations) were <15%. The methodology recoveries were more than 66.63%. This validated method was successfully applied to a pharmacokinetic study after intravenous injection administration of gambogenic acid in dogs at a dose of 1 mg/kg. Copyright © 2014 John Wiley & Sons, Ltd.
    Full-text · Article · Dec 2014 · Biomedical Chromatography
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    • "Unfortunately, we did not continue to investigate the detail signaling pathways. Li et al. have reported that GNA can inhibit A549 cell proliferation through apoptosis by inducing the up-regulation of the p38 MAPK cascade and cell cycle arrest (Li et al., 2010; Yan et al., 2012). Chen et al. found that GNA induced time-and dose-dependent growth inhibition and apoptosis Akt pathway inactivation in U251 glioblastoma cells (Chen et al., 2012). "
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    ABSTRACT: Background: Gambogenic acid is a major active compound of gamboge which exudes from the Garcinia hanburyi tree. Gambogenic acid anti-cancer activity in vitro has been reported in several studies, including an A549 nude mouse model. However, the mechanisms of action remain unclear. Methods: We used nude mouse models to detect the effect of gambogenic acid on breast tumors, analyzing expression of apoptosis-related proteins in vivo by Western blotting. Effects on cell proliferation, apoptosis and apoptosis-related proteins in MDA-MB-231 cells were detected by MTT, flow cytometry and Western blotting. Inhibitors of caspase-3,-8,-9 were also used to detect effects on caspase family members. Results: We found that gambogenic acid suppressed breast tumor growth in vivo, in association with increased expression of Fas and cleaved caspase-3,-8,-9 and bax, as well as decrease in the anti-apoptotic protein bcl-2. Gambogenic acid inhibited cell proliferation and induced cell apoptosis in a concentration-dependent manner. Conclusion: Our observations suggested that Gambogenic acid suppressed breast cancer MDA-MB-231 cell growth by mediating apoptosis through death receptor and mitochondrial pathways in vivo and in vitro.
    Preview · Article · Dec 2013 · Asian Pacific journal of cancer prevention: APJCP
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    • "Acridine orange/ethidium bromide (AO/EB) double fluorescent dyes were used to qualitatively observe apoptotic cells. AO can emit a green fluorescence if it passes through the complete cell membrane and embeds in nuclear DNA while EB can mark nuclear DNA of damaged cells and emit a red–orange fluorescence (Yan et al., 2012 "
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    ABSTRACT: Herbacetin (HER) is a natural flavonoid compound that can be extracted from Ramose Scouring Rush Herb, and its biological and pharmacological activities lack of corresponding attention. In this study, the apoptotic effect of HER against the human hepatoma cell line (HepG2) was investigated. The results showed that HepG2 cells apoptosis occurred in a dose-dependent manner within 48h incubated with HER, which was confirmed by DNA fragmentation, nuclear shrinkage, and poly (ADP-ribose) polymerase (PARP) cleavage. HER at 25-100μM induced a mitochondria-dependent apoptotic pathway associated with Bcl-2/Bax ratio decrease, mitochondrial membrane potential (ΔΨ) collapse, cytochrome c release, and caspase-3 activation. Increasing expression of peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α) was also observed in HER-treated cells. Furthermore, the addition of a ROS inhibitor (N-Acetyl-l-cysteine, NAC) significantly attenuated the apoptosis induced by HER and also blocked the expression of PGC-1α protein. Additionally, HER effectively inhibited the phosphorylation of Akt and the phosphatidylinositol-3 kinase (PI3K) inhibitor LY294002 increased the inhibition effect of HER on Akt phosphorylation. These findings provide evidences that HER induces HepG2 apoptosis in a ROS-mediated mitochondria-dependent manner that correlate with the inactivation of the PI3K/Akt pathway.
    Full-text · Article · Oct 2012 · Food and chemical toxicology: an international journal published for the British Industrial Biological Research Association
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