• Journal of Life Science
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• v.21 no.7
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• pp.923-931
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• 2011

In the present study, we investigated the effect of indole-3-carbinol (I3C), a natural compound present in vegetables, on the cell migration and invasion of OVCAR-3 ovarian cancer cells. Our results indicated that I3C inhibited the proliferation of OVCAR-3 cells, a process which was associated with inhibition of cell motility as determined by wound healing experiments and cell invasion studies. I3C treatment increased the tightness of the tight junctions (TJs), which was demonstrated by an increase in transepithelial electrical resistance and a decrease in paracellular permeability. The RT-PCR and immunoblotting results indicated that I3C repressed the levels of claudin-3 as well as claudin-4, proteins that comprise a major part of TJs and play a key role in the control and selectivity of paracellular transport. Furthermore, the activities of matrix metalloproteinase (MMP)-2 and MMP-9 were also decreased by treatment with I3C, which was connected with the down-regulation of their mRNAs and protein expression. The results suggest that I3C may be expected to inhibit cancer cell metastasis and invasion by restoring TJs and decreasing MMP activity in ovarian cancer cell line OVCAR-3.

• Clinical and Experimental Reproductive Medicine
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• v.33 no.2
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• pp.115-123
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• 2006

Objective: To investigate new signal transduction cascade through integrin, FAK and ERK in the suppressed cell proliferation by GnRH-I and -II. Method: Human endometrial cancer cells (HEC1A) were cultured under the following condition: DMEM/F12 (10% FBS). GnRH-I and -II were treated time (0, 5, 10, 15, 20, 30 min; 100 nM) and dose (10 nM or 100 nM; 20 min) dependent manner according to experimental purposes. Cell proliferation was measured using [$^3H$] thymidine incorporation assay. Immunoblotting was utilized to detect proteins. Results: GnRH-I and -II inhibited proliferation of HEC1A cells and induced expression of integrin ${\beta}3$. Phosphorylation of FAK and ERK were induced by GnRH-I and -II. Conclusion: GnRH inhibited cell proliferation via the expression of integrin and FAK, ERK phosphorylation.

• Journal of Ginseng Research
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• v.41 no.1
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• pp.43-51
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• 2017

Background: BIOGF1K, a compound K-rich fraction prepared from the root of Panax ginseng, is widely used for cosmetic purposes in Korea. We investigated the functional mechanisms of the anti-inflammatory and antioxidative activities of BIOGF1K by discovering target enzymes through various molecular studies. Methods: We explored the inhibitory mechanisms of BIOGF1K using lipopolysaccharide-mediated inflammatory responses, reporter gene assays involving overexpression of toll-like receptor adaptor molecules, and immunoblotting analysis. We used the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay to measure the antioxidative activity. We cotransfected adaptor molecules, including the myeloid differentiation primary response gene 88 (MyD88) and Toll/interleukin-receptor domain containing adaptor molecule-inducing interferon-${\beta}$ (TRIF), to measure the activation of nuclear factor (NF)-${\kappa}B$ and interferon regulatory factor 3 (IRF3). Results: BIOGF1K suppressed lipopolysaccharide-triggered NO release in macrophages as well as DPPH-induced electron-donating activity. It also blocked lipopolysaccharide-induced mRNA levels of interferon-${\beta}$ and inducible nitric oxide synthase. Moreover, BIOGF1K diminished the translocation and activation of IRF3 and NF-${\kappa}B$ (p50 and p65). This extract inhibited the upregulation of NF-${\kappa}B$-linked luciferase activity provoked by phorbal-12-myristate-13 acetate as well as MyD88, TRIF, and inhibitor of ${\kappa}B$ ($I{\kappa}B{\alpha}$) kinase ($IKK{\beta}$), and IRF3-mediated luciferase activity induced by TRIF and TANK-binding kinase 1 (TBK1). Finally, BIOGF1K downregulated the NF-${\kappa}B$ pathway by blocking $IKK{\beta}$ and the IRF3 pathway by inhibiting TBK1, according to reporter gene assays, immunoblotting analysis, and an AKT/$IKK{\beta}$/TBK1 overexpression strategy. Conclusion: Overall, our data suggest that the suppression of $IKK{\beta}$ and TBK1, which mediate transcriptional regulation of NF-${\kappa}B$ and IRF3, respectively, may contribute to the broad-spectrum inhibitory activity of BIOGF1K.

• Development and Reproduction
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• v.8 no.2
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• pp.99-111
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• 2004

The present study aimed to investigate whether the expression of ADAM-8, 9, 10, 12, 15, 17 and ADAMTS-1 genes is controlled by ovarian steroid hormones. Ovariectomized mice were injected with 17 ${\beta}$-estradiol ($E_2$), progesterone ($P_4$, or $E_2+P_4$. Uterine tissues were processed for RT-PCR and immunoblotting. The results of RT-PCR showed that administration of $E_2$ increases the level of ADAM-8, 12 and ADAM17 expression compared to $P_4$ or control group. In contrast, administration of $P_4$ markedly stimulated the expression of ADAM-9, 10, 15 and ADAMTS-1, whereas $E_2$ did not. Immunoblotting analysis using anti-mouse ADAM polyclonal antibodies demonstrated that $E_2$ alone or $E_2+P_4$ treatment results in the strong expression of ADAM-8, 12 and ADAM17 proteins but $P_4$ alone or control group gave weak expression. In contrast, $P_4$ alone or $E_2$ plus $P_4$ treatment increased the expression level of ADAM-9, 10, 15 and DAMTS-1 proteins. $E_2$ alone or control group did not increase the expression. These results indicate that expression of ADAM-8, 12 and ADAM17 genes is upregulated by $E_2$ and that of ADAM-9, 10, 15 and ADAMTS-1 gene is upregulated by $P_4$.

• Korean Journal of Medicinal Crop Science
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• v.15 no.1
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• pp.38-45
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• 2007

Korean mistletoe (Viscum album L) extract has been found to posses immunoregulating activity. In this study, Korean mistletoe extract, M11C (non-lectin components), was used to know whether this extract activates splenocytes to secret interleukin $1\beta(IL-1\beta)$. The splenocytes were treated with M11C, and then collected the supernatant and cell lysate that were prepared to analyze the level of $IL-1\beta$, using ELISA, immunoblotting, and RT-PCR. Maximum effective dose and time of M11C on $IL-1\beta$ secretion from splenocytes were $200{\mu}g/m\ell$ and 8 hours, respectively. Treatment dose and time for the maximum expression of $IL-1\beta$ mRNA were $200{\mu}g/m\ell$ and 4 hours, respectively. Saccharide degradation enzyme Viscozyme L completely blocked the effect of M11C on $IL-1\beta$ secretion from splenocytes. As the result, among non-lectin components saccharide could be regarded as a main component for $IL-1\beta$ expression from splenocytes.

• Korean Journal of Pharmacognosy
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• v.34 no.3 s.134
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• pp.210-217
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• 2003

Korean mistletoe (Viscum album) extract has been found to posses immunostimulatory activity. In this study, Korean mistletoe extract, M11C (non-lectin components), was used to know whether this extract might activate mouse splenic macrophages to produce tumor necrosis $factor-{\alpha}\;(TNF-{\alpha}$) and might play a role in anticancer. To know the effect of M11C on the production of $TNF-{\alpha}$, the splenic macrophages were treated by the M11C, and then collected the supernatant (M11C stimulated splenic macrophage-conditioned media; MSCM). MSCM was analyzed for the $TNF-{\alpha}$ secretion by means of ELISA and immunoblotting, and mRNA expression was analyzed by RT-PCR. The S-180 murine sarcoma model was established to know the effect of M11C on the inhibition of tumor growth. M11C had the effect of $TNF-{\alpha}$ production from splenic macrophages performed by ELISA technique. This ELISA data was reconfirmed by immunoblotting assay. The effects of M11C on the expression of $TNF-{\alpha}$ mRNA from the macrophages was also shown. M11C also had the inhibitory effect of S-180 tumor growth. These data suggest that Korean mistletoe extract M11C may be used for an immunomodulator.

• Journal of Life Science
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• v.17 no.3 s.83
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• pp.350-355
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• 2007

The rat RBL-2H3 mast cells contain various Src-family kinases. Previous reports with this cell line indicated that Lyn activation is an important initial signaling for the activation of the cells. However, the role and location of other Src-family kinase isoforms which are expressed in the cells are not clear. In this study, we now show that isoforms of Src-family kinases, Lyn, fyn, Fgr, c-Src, and Yes are differentially expressed and located differently in the cells as indicated by RT-PCR, immunoblotting analysis, and confocal microscopy. Lyn and Fgr were located on plasma membrane but on the other hand c-Src and Yes were located on intracellular organelle. All of Src-family kinases were cloned and overexpressed for investigating the roles of the isoforms. Overexpression of Fyn and Fgr, not Lyn and c-Src, stimulated Ag-induced degranulation in the cells. Our findings strongly suggest for the first time that each of Src-family kinase isoform can regulate differentially $Fc{\varepsilon}RI$-mediated signaling in RBL-2H3 mast cells.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.13 no.8
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• pp.3785-3791
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• 2012

Dissolved oxygen is one of the most important factors controlling growth in aquatic organisms. Hypoxia is generally defined as dissolved oxygen less than 2.8 mg $O_2/L$ (equivalent to 2 mL $O_2/L$ or 91.4 mM). Therefore, hypoxia zone can cause a serious problem in marine ecosystem. In this study, to investigate embryotoxic (fertilization and embryo development rates) effects of hypoxia on sea urchin Strongylocentrotus nudus were exposed to dissolved oxygen levels of 7.6 mg $O_2/L$ (normoxia) and 1.8 mg $O_2/L$ (hypoxia) for 2 days at $15^{\circ}C$ and 33 ‰. Also, Expression levels of stress related gene (HSP70) and antioxidant related gene (glutathione reductase) in the sea urchins exposed to hypoxia were confirmed by Immunoblotting and RT-PCR analysis. In results, we showed that developmental rates were dramatically reduced in hypoxia condition. Molecular analysis demonstrated that higher HSP70 (5.5 fold) and glutathione reductase gene (2.79 fold) were present in the sea urchin exposed to hypoxia. Our results suggested that hypoxia can cause the abnormal development and elicits a stress and antioxidant response on sea urchin.

• Journal of Ginseng Research
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• v.47 no.6
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• pp.726-734
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• 2023

Background: Skeletal muscles play a key role in physical activity and energy metabolism. The loss of skeletal muscle mass can cause problems related to metabolism and physical activity. Studies are being conducted to prevent such diseases by increasing the mass and regeneration capacity of muscles. Ginsenoside Rg5 has been reported to exhibit a broad range of pharmacological activities. However, studies on the effects of Rg5 on muscle differentiation and growth are scarce. Methods: To investigate the effects of Rg5 on myogenesis, C2C12 myoblasts were induced to differentiate with Rg5, followed by immunoblotting, immunostaining, and qRT-PCR for myogenic markers and promyogenic signaling (p38MAPK). Immunoprecipitation confirmed that Rg5 increased the interaction between MyoD and E2A via p38MAPK. To investigate the effects of Rg5 on prevention of muscle mass loss, C2C12 myotubes were treated with dexamethasone to induce muscle atrophy. Immunoblotting, immunostaining, and qRT-PCR were performed for myogenic markers, Akt/mTOR signaling for protein synthesis, and atrophy-related genes (Atrogin-1 and MuRF1). Results: Rg5 promoted C2C12 myoblast differentiation through phosphorylation of p38MAPK and MyoD/E2A heterodimerization. Furthermore, Rg5 stimulated C2C12 myotube hypertrophy via phosphorylation of Akt/mTOR. Phosphorylation of Akt induces FoxO3a phosphorylation, which reduces the expression of Atrogin-1 and MuRF1. Conclusion: This study provides an understanding of how Rg5 promotes myogenesis and hypertrophy and prevents dexamethasone-induced muscle atrophy. The study is the first, to the best of our knowledge, to show that Rg5 promotes muscle regeneration and to suggest that Rg5 can be used for therapeutic intervention of muscle weakness and atrophy, including cancer cachexia.

• Molecules and Cells
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• v.26 no.2
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• pp.200-205
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• 2008

The mitogen-activated protein kinase (MAPK) signaling pathway is activated in response to extracellular stimuli and regulates various activities in eukaryotic cells. Following exposure to stimuli, MAPK is known to be activated via dual phosphorylation at a conserved TxY motif in the activation loop; both threonine and tyrosine residues are phosphorylated by an upstream kinase. However, the mechanism underlying dual phosphorylation is not clearly understood. In the budding yeast Saccharomyces cerevisiae, the Hog1 MAPK mediates the high-osmolarity glycerol (HOG) signaling pathway. Tandem mass spectrometry and phosphospecific immunoblotting were performed to quantitatively monitor the dynamic changes occurring in the phosphorylation status of the TxY motif of Hog1 on exposure to osmotic stress. The results of our study suggest that the tyrosine residue is preferentially and dynamically phosphorylated following stimulation, and this in turn leads to the dual phosphorylation. The tyrosine residue was hyperphosphorylated in the absence of a threonine residue; this result suggests that the threonine residue is critical for the control of signaling noise and adaptation to osmotic stress.