Induction of apoptosis in melanoma A375 cells by a chloroform fraction of Centratherum anthelminticum (L.) seeds involves NF-kappaB, p53 and Bcl-2-controlled mitochondrial signaling pathways

Looi, Chung Yeng; Moharram, Bushra; Paydar, Mohammadjavad; Wong, Yi Li; Leong, Kok Hoong; Mohamad, Khalit; Arya, Aditya; Wong, Won Fen; Mustafa, Mohd Rais

doi:10.1186/1472-6882-13-166

Cited by 54 publications

(32 citation statements)

References 34 publications

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“…1×10 4 cells per well were seeded onto 96-well plate for 16 h. Next, cells were treated with DESs (IC 50 dosage) and further incubated for 24 h. To examine plasma membrane permeability, cell permeability dye (Excitation 491/Emission 509) were added to live cells and incubated for 1 h, as previously described [ 22 ]. Cells were washed three times with PBS before fixing with 4% formaldehyde for 15 min.…”

Section: Materials and Experimental Methodologiesmentioning

confidence: 99%

In Vitro and In Vivo Toxicity Profiling of Ammonium-Based Deep Eutectic Solvents

et al. 2015

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The cytotoxic potential of ammonium-based deep eutectic solvents (DESs) with four hydrogen bond donors, namely glycerine (Gl), ethylene glycol (EG), triethylene glycol (TEG) and urea (U) were investigated. The toxicity of DESs was examined using In Vitro cell lines and In Vivo animal model. IC50 and selectivity index were determined for the DESs, their individual components and their combinations as aqueous solutions for comparison purposes. The cytotoxicity effect of DESs varied depending on cell lines. The IC50 for the GlDES, EGDES, UDES and TEGDES followed the sequence of TEGDES< GlDES< EGDES< UDES for OKF6, MCF-7, A375, HT29 and H413, respectively. GlDES was selective against MCF-7 and A375, EGDES was selective against MCF-7, PC3, HepG2 and HT29, UDES was selective against MCF-7, PC3, HepG2 and HT29, and TEGDES was selective against MCF-7 and A375. However, acute toxicity studies using ICR mice showed that these DESs were relatively toxic in comparison to their individual components. DES did not cause DNA damage, but it could enhance ROS production and induce apoptosis in treated cancer cells as evidenced by marked LDH release. Furthermore, the examined DESs showed less cytotoxicity compared with ionic liquids. To the best of our knowledge, this is the first time that combined In Vitro and In Vivo toxicity profiles of DESs were being demonstrated, raising the toxicity issue of these neoteric mixtures and their potential applicability to be used for therapeutic purposes.

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Section: Materials and Experimental Methodologiesmentioning

confidence: 99%

In Vitro and In Vivo Toxicity Profiling of Ammonium-Based Deep Eutectic Solvents

et al. 2015

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“…The Cellomics ArrayScan HCS system was used to analyze the suppressive effect of AMEAE on the nuclear translocation of NF-κB induced by TNF-α. The experiment was carried out using Cellomics nucleus factor-κB (NF-κB) activation kit (Thermo Scientific) as previously described [ 20 ]. In brief, A549 cells (1.0 × 10 4 cells/well) were treated with AMEAE at different concentrations in a 96-well plate for 3 h. The treated A549 cells were stimulated by TNF-α (1 ng/mL) for 30 min.…”

Section: Methodsmentioning

confidence: 99%

Annona muricata leaves induced apoptosis in A549 cells through mitochondrial-mediated pathway and involvement of NF-κB

Moghadamtousi

Kadir

Paydar

et al. 2014

BMC Complement Altern Med

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BackgroundAnnona muricata leaves have been reported to have antiproliferative effects against various cancer cell lines. However, the detailed mechanism has yet to be defined. The current study was designed to evaluate the molecular mechanisms of A. muricata leaves ethyl acetate extract (AMEAE) against lung cancer A549 cells.MethodsThe effect of AMEAE on cell proliferation of different cell lines was analyzed by MTT assay. High content screening (HCS) was applied to investigate the suppression of NF-κB translocation, cell membrane permeability, mitochondrial membrane potential (MMP) and cytochrome c translocation from mitochondria to cytosol. Reactive oxygen species (ROS) formation, lactate dehydrogenase (LDH) release and activation of caspase-3/7, -8 and -9 were measured while treatment. The western blot analysis also carried out to determine the protein expression of cleaved caspase-3 and -9. Flow cytometry analysis was used to determine the cell cycle distribution and phosphatidylserine externalization. Quantitative PCR analysis was performed to measure the gene expression of Bax and Bcl-2 proteins.ResultsCell viability analysis revealed the selective cytotoxic effect of AMEAE towards lung cancer cells, A549, with an IC50 value of 5.09 ± 0.41 μg/mL after 72 h of treatment. Significant LDH leakage and phosphatidylserine externalization were observed in AMEAE treated cells by fluorescence analysis. Treatment of A549 cells with AMEAE significantly elevated ROS formation, followed by attenuation of MMP via upregulation of Bax and downregulation of Bcl-2, accompanied by cytochrome c release to the cytosol. The incubation of A549 cells with superoxide dismutase and catalase significantly attenuated the cytotoxicity caused by AMEAE, indicating that intracellular ROS plays a pivotal role in cell death. The released cytochrome c triggered the activation of caspase-9 followed by caspase-3. In addition, AMEAE-induced apoptosis was accompanied by cell cycle arrest at G0/G1 phase. Moreover, AMEAE suppressed the induced translocation of NF-κB from cytoplasm to nucleus.ConclusionsOur data showed for the first time that the ethyl acetate extract of Annona muricata inhibited the proliferation of A549 cells, leading to cell cycle arrest and programmed cell death through activation of the mitochondrial-mediated signaling pathway with the involvement of the NF-kB signalling pathway.

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“…JC-1 molecules accumulate in mitochondria as aggregate, thus rendering strong red fluorescence emission. However, in apoptotic cells, JC-1 molecules exist in the cytoplasm as monomers, which can be reflected by strong green and weak red fluorescence emission [13]. The ratio of green and red fluorescence indicates depolarization percentage of mitochondria.…”

Section: Eet Regulated the Expression Of Apoptosis-related Protein Inmentioning

confidence: 99%

Ethylacetate extract from Tetrastigma hemsleyanum induces apoptosis via the mitochondrial caspase-dependent intrinsic pathway in HepG2 cells

et al. 2015

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Ethylacetate extract of Tetrastigma hemsleyanum (EET) has a potent antitumor activity in vitro and in vivo. However, the molecular mechanism underlying EET-induced apoptosis remains elusive. As part of our continuing studies, we investigated the apoptosis mechanism of HepG2 cells exposed to different concentrations of EET in vitro. Confocal laser scanning was used to detect the apoptotic morphological changes. Flow cytometer and inverted fluorescence microscope were used to detect the mitochondrial membrane potential and cytosolic Ca(2+) level. Western blotting analysis was used to evaluate the expression of the apoptosis-related proteins. Annexin V/PI staining was used to investigate cell apoptosis. Spectrophotometry was used to detect the activity of caspase family. The results showed that distinct apoptotic morphological changes occurred in HepG2 cells treated by EET. EET caused collapse of mitochondrial membrane potential, elevation of cytosolic Ca(2+) level, and evoked release of cytochrome c from mitochondria in a concentration-dependent manner. The apoptosis was accompanied by a significant activation of caspase-3, caspase-9, and the cleavage of poly (ADP-ribose) polymerase, but there was no significant change in either the activity or the expression level of caspase-8. Furthermore, EET-induced apoptosis could be inhibited by caspase-9 inhibitor Z-LEHD-FMK but not by caspase-8 inhibitor Z-IETD-FMK. Taken together, these overall results demonstrated that EET-induced apoptosis of HepG2 cells was mediated by the mitochondrial caspase-dependent intrinsic pathway rather than the death receptor/caspase-8-mediated signaling route.

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Induction of apoptosis in melanoma A375 cells by a chloroform fraction of Centratherum anthelminticum (L.) seeds involves NF-kappaB, p53 and Bcl-2-controlled mitochondrial signaling pathways

Cited by 54 publications

References 34 publications

In Vitro and In Vivo Toxicity Profiling of Ammonium-Based Deep Eutectic Solvents

In Vitro and In Vivo Toxicity Profiling of Ammonium-Based Deep Eutectic Solvents

Annona muricata leaves induced apoptosis in A549 cells through mitochondrial-mediated pathway and involvement of NF-κB

Ethylacetate extract from Tetrastigma hemsleyanum induces apoptosis via the mitochondrial caspase-dependent intrinsic pathway in HepG2 cells

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