• Journal of Life Science
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• v.28 no.11
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• pp.1268-1276
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• 2018

The cytidine analog decitabine (DEC) acts as a nucleic acid synthesis inhibitor, whereas ammonium pyrrolidine dithiocarbamate (PDTC) is an inhibitor of nuclear factor-${\kappa}B$. The aim of this study was to investigate the possible synergistic inhibitory effect of these two inhibitors on proliferation of human gastric cancer AGS cells. The inhibitory effect of PDTC on AGS cell proliferation was significantly increased by DEC in a concentration-dependent manner, and this inhibition was associated with cell cycle arrest at the G2/M phase and the induction of apoptosis. This induction of apoptosis by the co-treatment with PDTC and DEC was related to the induction of DNA damage, as assessed by H2AX phosphorylation. Further studies demonstrated that co-treatment with PDTC and DEC induced the disruption of mitochondrial membrane potential, increased the generation of intracellular reactive oxygen species (ROS) and the expression of pro-apoptotic Bax, and down-regulated the expression of anti-apoptotic Bcl-2, ultimately resulting in the release of cytochrome c from the mitochondria into the cytoplasm. Co-treatment with PDTC and DEC also activated caspase-8 and caspase-9, which are representative caspases of the extrinsic and intrinsic apoptosis pathways. Co-treatment also activated caspase-3, which was accompanied by proteolytic degradation of poly (ADP-ribose) polymerase. Taken together, these data clearly indicated that co-treatment with PDTC and DEC suppressed the proliferation of AGS cells by increasing DNA damage and activating the ROS-mediated extrinsic and intrinsic apoptosis pathways.

• Journal of Life Science
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• v.29 no.10
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• pp.1062-1070
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• 2019

A components of garlic (Allium sativum) have anti-proliferative effects against various types of cancer. We aimed to investigate the capacity of garlic compounds to anti-tumor on a various cancer cell lines. Fractionation of garlic extract, guided by antiproliferative activity against human gastric cancer (AGS) cells, has resulted in the isolation of N-benzyl-N-methyldecan-1-amine (NBNMA). We investigated the effect of newly isolated NBNMA from garlic cloves on the inhibition of the growth of CT-26, AGS, HepG2, HCT-116, MCF7, B16F10, and Sarcoma-180 cells for in vitro and CT-26 colon carcinoma cells in vivo. NBNMA exhibited an antiproliferative effect in CT-26 cells by apoptotic cell death. NBNMA exhibited down-regulation of anti-apoptotic Bcl-2 proteins and up-regulation of apoptotic Bad protein expression in western blot analyses. In addition, NBNMA meagre activated caspase 3 and caspase 9, initiator caspases of the extrinsic and intrinsic pathways of apoptosis. NBNMA treatment at a dose of 10 mg/kg for 21 days in experimental mice implanted with tumors resulted in significant reduction of the tumor weight (43%). NBNMA exhibited both in vitro and in vivo anticancer activity. These results indicate that NBNMA has promising potential to become a novel anticancer agent from garlic cloves for the treatment of colon carcinoma cancer.

• Journal of Life Science
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• v.19 no.11
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• pp.1529-1537
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• 2009

To elucidate further the antitumor effects of a natural L-arginine analogue, L-canavanine, the mechanism underlying apoptogenic activity of L-canavanine and its modulation by protein tyrosine kinase $p56^{lck}$ was investigated in human Jurkat T cells. When the cells were treated with 1.25 to 2.5 mM L-canavanine for 36 h, several apoptotic events including mitochondrial membrane potential (${\Delta\Psi}m$) loss, activation of caspase-9, -3, -8, and -7, poly (ADP-ribose) polymerase (PARP) degradation, and DNA fragmentation were induced without alteration in the levels of Fas or FasL. These apoptotic changes were more significant in $p56^{lck}$-deficient Jurkat clone JCaM1.6 than in $p56^{lck}$-positive Jurkat clone E6.1. The L-canavanine-induced apoptosis observed in $p56^{lck}$-deficient JCaM1.6 cells was significantly reduced by introducing $p56^{lck}$ gene into JCaM1.6 cells by stable transfection. Treatment of JCaM1.6/lck cells with L-canavanine caused a transient 1.6-fold increase in the kinase activity of $p56^{lck}$. Both FADD-positive wild-type Jurkat T cell clone A3 and FADD-deficient Jurkat T cell clone I2.1 exhibited a similar susceptibility to the cytotoxicity of L-canavanine, excluding involvement of Fas/FasL system in triggering L-canavanine-induced apoptosis. The L-canavanine-induced apoptotic sub-$G_1$ peak and activation of caspase-3, -8, and -7 were abrogated by pan-caspase inhibitor (z-VAD-fmk), whereas L-canavanine-induced activation of caspase-9 was not affected. These results demonstrated that L-canavanine caused apoptosis of Jurkat T cells via the loss of ${\Delta\Psi}m$, and the activation of caspase-9, -3, -8, and -7, leading to PARP degradation, and that the $p56^{lck}$ kinase attenuated the ${\Delta\Psi}m$ loss and activation of caspases, and thus contributed as a negative regulator to L-canavanine-induced apoptosis.

• The korean journal of orthodontics
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• v.33 no.6 s.101
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• pp.453-463
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• 2003

Mammalian cell is critically dependent on a continuous supply of oxygen. Even brief periods of oxygen deprivation can result in profound cellular damage. The aim of this study was to examine the possible mechanism of apoptosis in response to hypoxia in MC3T3E1 osteoblasts. MC3T3El osteoblasts under hypoxic conditions ($2\%$ oxygen) resulted in apoptosis in a time-dependent manner, determined by DNA fragmentation assay and nuclear morphology, stained with fluorescent dye (Hoechst 33258) Pretreatment with Z-VAD-FMK, a pancaspase inhibitor, or Z-DEVD-CHO, a specific caspase-3 inhibitor, suppressed the DNA ladder in response to hypoxia in a concentration dependent manner. An increase in caspase-3-like protease (DEVDase) activity was observed during apoptosis, but no caspase-l activity (YVADase) was detected. To confirm what caspases were involved in apoptosis, western blot analysis was performed using an anticaspase-3 or 6 antibody. The 17-kDa protein, that corresponds to the active products of caspase-3 and the 20-kDa protein of the active protein of caspase-6 were generated in hypoxia-challenged lysates, in which the full length forms of caspase-3 and 6 were evident. With a time course similar to caspase-3 and 6 activation, hypoxic stress also caused the cleavage of Lamin A, typical of caspase-6 activity. In addition, the hypoxic stress elicited the release of cytochrome c into the cytosol during apoptosis. These findings suggested that the activation of caspases accompanied by a cytochrome c release in response to hypoxia was involved in apoptotic cell death in MC3T3E1 osteoblasts.

• Journal of Life Science
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• v.19 no.9
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• pp.1209-1217
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• 2009

Many cancer cells are sensitive to the TNF-related apoptosis-inducing ligand (TRAIL)-induced apoptosis. However, some cancer cells show either partial or complete resistance to TRAIL. Human colon carcinoma KM12 cells have been shown to be insensitive to TRAIL-induced apoptosis. To overcome TRAIL resistance in KM12 cells, we targeted key anti-apoptotic molecules involved in the modulation of TRAIL resistance in the cells, and evaluated the effects of quercetin as a TRAIL sensitizer in the cells. We found that quercetin acted in synergy with TRAIL to enhance TRAIL-induced apoptosis in KM12 cells by the down-regulation of c-FLIP and DNA-PKcs/Akt and up-regulation of death receptors (DR4/DR5), which led to the enhancement of TRAIL-mediated activation of caspases and subsequent cleavage of PARP, as well as up-regulation of Bax. These findings suggest that the DNA-PKcs/Akt signaling pathway, as well as c-FLIP, play essential roles in regulating cells in the escape from TRAIL-induced apoptosis. Based on these results, this study provides a potential application of quercetin in combination with TRAIL in the treatment of human colon cancer.

• Animal cells and systems
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• v.6 no.2
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• pp.171-180
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• 2002

Nitric oxide has high affinity for iron, and thus it can cause intracellular iron loss. We tested the idea that intracellular iron can be the primary target of NO toxicity by comparing the signaling mechanisms involved in cell death caused by iron depletion and that caused by NO. Treatment of HL-60 cells with a NO donor, S-nitroso-N-acetyl-DL-penicillamine (SNAP), decreased the intracellular iron level rapidly as that observed with the iron chelator deferoxamine (DFO). Iron chelators such as DFO and mimosine could induce death of human leukemic HL-60 cells by a mechanism requiring activation of p38 kinase, c-Jun N-terminal kinase, caspase-3 and caspase-8. DFO and SNAP also caused release of cytochrome c from mitochondria. Inhibition of p38 kinase by a selective inhibitor, SB203580, abolished the NO and DFO-induced cell death, release of cytochrome c, and activation of caspase-3 and caspase-8, thus indicating that p38 kinase lies upstream in the cell death processes. In a parallel situation, the cells that are sensitive to NO showed similar sensitivity to DFO. Moreover, simultaneous addition of ferric citrate, an iron-containing compound, inhibited the SNAP and DFO-induced activation of caspases and also blocked the NO-mediated cell cycle arrest at $G_1$ phase. Collectively, our data implicate that the NO-induced cell death of tumor cells including HL-60 cells is mediated by depletion of iron and further suggest that activation of p38 kinase lies upstream of cytochrome c release and caspase activation involved in this apoptotic process.

• Journal of Life Science
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• v.19 no.8
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• pp.1047-1054
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• 2009

Oral squamous carcinoma (OSC) cells present resistance to chemotherapeutic agents-mediated apoptosis in the late stages of malignancy. Advances in the understanding of bacterial toxins have produced new strategies for the treatment of cancers. It was demonstrated here that Pseudomonas aeruginosa exotoxin A (PEA) significantly decreased the viability of chemoresistant YD-9 cells in the apoptosis mechanism. Apoptotic manifestations were evident through changes in nuclear morphology and generation of DNA fragmentation. PEA treatment induced caspase-3, -6 and -9 cleavage, and activation. These events preceded proteolysis of the caspase substrates poly (ADP-ribose) polymerase (PARP), DNA fragmentation factor 45 (DFF45), and lamin A in YD-9 cells. The reduction of mitochondrial membrane potential, release of cytochrome c and SmacjDlABLO from mitochondria to cytosol, andtranslocation of AlF into nucleus were shown. While p53, p21 and $14-3-3{\gamma}$ were upregulated, cyclin Band cdc2 were downregulated by PEA treatment. Taken together, PEA induces apoptosis in chemoresistant YD-9 cells via activation of caspases, mitochondrial events and regulation of cell cycle genes.

• Journal of Life Science
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• v.19 no.8
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• pp.1023-1032
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• 2009

Despite the fact that many cancer cells are sensitive to TNF-related apoptosis-inducing ligand (TRAIL)-induced apoptosis, some cancer cells show either partial or complete resistance to TRAIL. Human leukemic K562 and CEM cells also show resistance to TRAIL-induced apoptosis. Novel molecular target and treatment strategies are required to overcome TRAIL resistance of human leukemia cells. Therefore, the purpose of this study was to target key anti-apoptotic molecules deciding TRAIL resistance for sensitization of TRAIL-resistant K562 and CEM cells, and to evaluate the effect of quercetin as a TRAIL sensitizer on these TRAIL-resistant cells. We found that quercetin acted in synergy with TRAIL to enhance TRAIL-induced apoptosis in K562 cells by inhibition of the DNA-PK/Akt signaling pathway, which leads to enhancement of TRAIL-mediated activation of caspases and concurrent cleavage of PARP and up-regulation of Bax. The findings suggest that the DNA-PK/Akt signaling pathway plays an essential role in regulating cells to escape from TRAIL-induced apoptosis, and quercetin could act in synergy with TRAIL to increase apoptosis by inhibition of the DNA-PK/Akt signaling pathway, which overcomes TRAIL-resistance of K562 and CEM cells. This study suggests that DNA-PK might interfere with TRAIL-induced apoptosis in human leukemic cells through activation of the Akt signaling pathway.

• Journal of the Korean Society of Food Science and Nutrition
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• v.38 no.4
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• pp.442-450
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• 2009

Piperine is an alkaloid-amine found in pepper and has been reported to have anticarcinogenic properties. To explore the possibility that piperine has cancer chemopreventive and chemotherapeutic effects in colon cancer, we examined whether piperine inhibits the growth of HT-29 human colon cancer cells and investigated the mechanisms for this effect. Cells were cultured with various concentrations ($0{\sim}40{\mu}M$) of piperine. Piperine decreased the cell viability and induced apoptosis of HT-29 cells. Western blot analysis of total cell lysates revealed that piperine decreases the protein levels of Bcl-2, Mcl-1, and intact Bid but increases Bik levels. Piperine increased the percentage of cells with depolarized mitochondrial membrane, and the release of cytochrome c into cytoplasm. Piperine induced the cleavage of poly (ADP-ribose) polymerase and caspases 8, 9, 7, and 3 and increased the Fas levels. In addition, piperine significantly decreased the protein levels of survivin. The present results indicate that piperine inhibits the growth of HT-29 colon cancer cells by the induction of apoptosis, which may be mediated by its ability to change the Bcl-2 family proteins, increase the activation of caspases, and decrease survivin levels. Overall, our findings suggest that piperine has cancer chemotherapeutic effects in colon cancer.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2019.04a
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• pp.112-112
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• 2019

Baicalein is one of the main flavonoids derived from roots of Scutellaria baicalensis Georgi, a traditional Oriental medicine. Although baicalein has high antitumor effect on several human carcinomas, the mechanism responsible for this property is not unclear. In this study, the data revealed that baicale-ininduced growth inhibition was associated with the induction of apoptosis connecting with cytochrome c release, down-regulation of anti-apoptotic Bcl-xl and increased the percentage of cells with a loss of mitochondria membrane permeabilization. Baicalein also induced the proteolytic activation of caspases and cleavage of PARP; however, blockage of caspases activation by z-VAD-fmk inhibited baicalein-induced apoptosis. In addition, baicalein enhanced the formation of autophagosomes and up-regulated LC3-II/LC3-I ratio. Interestingly, the pretreatment of bafilomycin A1 recovered baicalein-induced cell death suggesting that autophagy by baicalein roles as protective autophagy. Taken together, our results indicated that this flavonoid induces apoptosis and cell protective autophagy. These data means combination treatment with baicalein and autophagy inhibitor might be a promising anticancer drug.