• Development and Reproduction
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• v.12 no.1
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• pp.97-105
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• 2008

Vinclozolin (VCZ) is a systemic fungicide commonly used in fruits, vegetables and the wine industry. VCZ and its metabolites, butenoic acid (M1) and enanilide (M2) derivatives, act as anti-androgens through actions on the androgen receptor. Although there is growing body of evidence that VCZ's action as an endocrine disrupting chemical (EDC) in male reproductive physiology and pathphysiology, no evidence on the VCZ's EDC action in female is available yet. Previously we found that the prepubertal VCZ exposures could effectively delay the onset of puberty in female rats, suggesting the postponed or weakened activities of hypothalamus-pituitary-ovary (H-P-O) reproductive hormonal axis. The present study was performed to examine whether the VCZ administration affects the transcriptional activities of reproductive hormone-related genes in the same animal model. VCZ (10 mg/kg/day) was administered daily from postnatal day 21 (PND 21) through the day when the first vaginal opening (V.O.) was observed. To determine the transcriptional changes of reproductive hormone-related genes in hypothalamus and pituitary, total RNAs were extracted and applied to the semiquantitative reverse transcription polymerase chain reaction (RT-PCR). As a result, treatment with VCZ significantly lowered the transcriptional activity of nitric oxide synthase-2 (NOS-2) which is known to adjust gonadotropin-releasing hormone (GnRH) secretion in the hypothalamus (p<0.01). Similarly, the mRNA levels of KiSS-1, G protein-coupled receptor 54 (GPR54) and GnRH were significantly decreased in hypothalamus (p<0.01) from VCZ-treated group. As expected, the transcriptional activities of luteinizing hormone-${\beta}$ (LH-${\beta}$) and follicle stimulating hormone-${\beta}$ (FSH-${\beta}$) in the anterior pituitary from VCZ-treated group were also significantly lower than those from the control group. The present study indicates that(i) the inhibitory effect of VCZ exposure on the onset of puberty in immature female rats could be derived from the reduced transcriptional activities of gonadotropin subunits and their upstream modulators such as GnRH and KiSS-1 in hypothalamus-pituitary neuroendocrine axis, and (ii) these inhibitory effects could be mediated by NO signaling pathway.

• Journal of Applied Biological Chemistry
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• v.61 no.2
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• pp.205-211
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• 2018

Rebaudioside A is a natural sweetener isolated from Stevia rebaudiana Bertoni, one of the glycosides based on steviol. Recent studies have shown that rebaudioside A inhibits the inflammatory response by inhibiting cytokines secretion such as interleukin-$1{\alpha}/1{\beta}$ in activated RAW264.7 mouse macrophage cells by LPS. However, the inhibitory mechanism of inflammation by rebaudioside A in the presence of LPS has not been fully elucidated. Therefore, in this study, we tried to investigate the anti-inflammatory activity of rebaudioside A at the protein level when RAW264.7 cells were stimulated by LPS. The inducible nitric oxide synthase protein expression level was reduced in the group treated with $250{\mu}M$ rebaudioside A compared to the LPS-treated group. In addition, the mRNA expression level of $NF-{\kappa}B$, which is a representative nuclear transcription factor by inflammatory signal, was also decreased as compared with that of LPS-treated group. In addition, $NF-{\kappa}B$ and inhibitor-${\kappa}B$ ($I-{\kappa}B$) complexes that are known to be dissociated by $I-{\kappa}B$ phosphorylation and ubiquitination were less phosphorylated than LPS treated group in the presence rebaudioside A. Finally, we could find that rebaudioside A was involved in the $NF-{\kappa}B$ pathway through reducing extracellular signal-regulated kinase1/2 phosphorylation in a concentration-dependent manner. These results suggest that rebaudioside A might suppress inflammatory reaction through MAPK and $NF-{\kappa}B$ regulation in LPS-stimulated RAW264.7.

• Clinical and Experimental Pediatrics
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• v.49 no.4
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• pp.431-438
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• 2006

Purpose : Insulin-like growth factor binding protein(IGFBP)-3 has been known as a tumor suppressor gene, and its anti-tumor function was divided into insulin-like growth factor(IGF)-dependent and IGF-independent mechanism. In IGF-independent mechanism, IGFBP-3 directly interacts with a cell without binding of IGFs, becoming an interesting object in oncology. Several studies demonstrate that one of the well-known tumor suppressor genes, p53, induces directly IGFBP-3 transcription, and the increment of IGFBP-3 expression induces apoptosis of many cancer cells. Recently, the anti-tumor mechanisms of IGFBP-3 have been reported, but post-translational modification of IGFBP-3 and its anti-tumor mechanism are not well known. In this study, we examined whether p53 regulated the glycosylation of IGFBP-3, and analysed the meaning of IGFBP-3 glycosylation related to the apoptosis of cancer cell. Methods : The p53-mutated status of MDA-MB-231 human breast cancer cells was used in this experiment. The expression and glycosylation of IGFBP-3 were tested by Western blot analysis after infection of adenovirus mediated Ad/p53 and/or Ad/IGFBP-3. Results : Ad/p53 infected cells resulted in growth retardation and the induced apoptosis. p53 induced direct expression and glycosylation of IGFBP-3. The increase of glcosylated IGFBP-3 was able to promote cellular apoptosis, and the glycosylation of IGFBP-3 was more activated by the double treatment of Ad/p53 and Ad/IGFBP-3. Conclusion : From this study, the anti-tumor activity of IGFBP-3 was shown to improve the stabilization of IGFBP-3 through the increment of glycosylation of IGFBP-3 by p53. This result suggests that the combined gene therapy of p53 and IGFBP-3 may appropriate treatment of cancer.

• Journal of Life Science
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• v.28 no.4
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• pp.454-464
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• 2018

Black soybeans are used as food sources as well as for traditional medicines because they contain an abundance of natural phenolic compounds. In this study, total phenolic contents (TPCs) of Korean black seed coat soybean varieties Socheongja (SCJ), Socheong 2 (SC2) and Cheongja 2 (CJ2) as well as their antioxidant capacities were investigated. Among them, TPCs were abundantly present in the order of CJ2$H_2O_2$-stimulated HaCaT human keratinocytes. Our results revealed that treatment with SCJ and SC2 prior to $H_2O_2$ exposure significantly increases the viability of HaCaT cells, indicating that the exposure of HaCaT cells to SCJ and SC2 conferred a protective effect against oxidative stress. SCJ and SC2 also effectively inhibited $H_2O_2$-induced apoptotic cell death through the blocking of mitochondrial dysfunction. SCJ and SC2 also attenuated the phosphorylation of Histone H2AX. Furthermore, they effectively induced the levels of thioredoxin reductase (TrxR) 1, a potent antioxidant enzyme, which is associated with the induction of nuclear transcription factor erythroid-2-like factor 2 (Nrf2); however, the protective effects of SCJ and SC2 were significantly reversed by Auranofin, a TrxR inhibitor. These results indicate that they have protective activity through the blocking of cellular damage related to oxidative stress via the Nrf2 signaling pathway. In conclusion, our study indicated that SCJ and SC2 might potentially serve as novel agents for the treatment and prevention of skin disorders caused by oxidative stress.

• Journal of Life Science
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• v.17 no.8 s.88
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• pp.1090-1099
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• 2007

Myristicin, l-allyl-3,4-methylenedioxy-5-methoxybenzene, was one of the major essential oils of nutmeg. However, its anti-allergic effect in the Th1/Th2 immune response was poorly understood. Recently, it was shown that T-bet and GATA-3 was master Th1 and Th2 regulatory transcription factors. In this study, we have attempted to determine whether myristicin regulates Th1/Th2 cytokine production, T-bet and GATA-3 gene expression in ovalbumin (OVA)-induced asthma model mice. Myristicin reduced levels of IL-4, Th2 cytokine production in OVA-sensitized and challenged mice. In the other side, it increased $IFN-{\gamma}$, Th1 cytokine production in myristicin administrated mice. We also examined to ascertain whether myristicin could influence eosinophil peroxidase (EPO) activity. After being sensitized and challenged with ovalbumin (OVA) showed typical asthmatic reactions. These reactions included an increase in the number of eosinophils in bronchoalveolar lavage fluid, an increase in inflammatory cell infiltration into the lung tissue around blood vessels and airways, and the development of airway hyper-responsiveness (AHR). The administration of myristicin before the last airway OVA challenge resulted in a significant inhibition of all asthmatic reactions. Accordingly, these findings provide new insight into the immunopharmacological role of myristicin in terms of its effects in a murine model of asthma.

• Clinical and Experimental Reproductive Medicine
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• v.29 no.1
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• pp.1-12
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• 2002

Objective: In order to acquire the technique for the establishment of human embryonic stem cells (ESe) derived from the human frozen-thawed embryos produced in IVF-ET program, this study was performed to establish mouse ESC derived from the in vitro fertilized embryos. Materials and Methods: After Fl hybrid (C57BL female $\times$ CBA mael) female mice were superovulated with PMSG and hCG treatment, their oocytes were retrieved and inseminated, and the fertilized embryos were cultured for 96-120 hours until the expected stages of blastocysts were obtained. To isolate the inner cell mass (ICM), either the blastocysts were treated with immunosurgery, or the whole embryos were cultured for 4 days. Isolated ICMs were then cultured onto STO feeder cell layer, and the resultant ICM colonies were subcultured with trypsin-EDTA treatment. During the subculture process, ESC-like cell colonies were observed with phase contrast microscopy. To identify ESC in the subcultured ESC-like cell colonies, alkaline phosphatase activity and Oct-4 (octamer-binding transcription factor-4) expression were examined by immunohistochemistry and RT-PCR, respectively. To examine the spontaneous differentiation, ESC-like cell colonies were cultured without STO feeder cell layer and leukemia inhibitory factor (LIF). Results: Seven ESC-like cell lines were established from ICMs isolated from the in vitro fertilized embryos. According to the developmental stage, the growth of ICMs isolated from the expanded blastocysts was significantly better than that of ICMs isolated from the hatched blastocysts (80.3% vs. 58.7%, p<0.05). ESC-like cell colonies were only obtained from ICMs of expanded blastocysts. However, the ICMs isolated from the embryos treated with immunosurgery were poorly grown and frequently differentiated during the culture process. The established ESC-like cell colonies were positively stained with alkaline phosphatase and expressed Oct-4, and their morphology resembled that observed in the previously reported mouse ESC. In addition, following the extended in vitro culture process, they maintained their expression of cell surface markers characteristic of the pluripotent stem cells such as alkaline phosphatase and Oct-4. When cultured without STO feeder cell layer and LIF, they were spontaneously differentiated into the various types of cells. Conclusion: The findings of this study suggest that the establishment of mouse ESC can be successfully derived from the in vitro fertilized embryos. The established ESC-like cells expressed the cell surface markers characteristic of the pluripotent stem cells and spontaneously differentiated into the various types of cells.

• Tuberculosis and Respiratory Diseases
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• v.66 no.3
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• pp.178-185
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• 2009

Background: Epigallocatechin-3-gallate (EGCG) is the major catechin in green tea, and has shown antiproliferative, antiangiogenic, antimetastatic and cell cycle pertubation activity in various tumor models. Hypoxia can be induced because angiogenesis is insufficient for highly proliferating cancer. Hypoxia-inducible factor-1$\alpha$ (HIF-1$\alpha$) and its downstream target, vascular endothelial growth factor (VEGF), are important for angiogenesis, tumor growth and metastasis. The aim of this study was to determine how hypoxia could cause changes in the cellular phenomena and microenvironment in a non-small cell culture system and to examine the effects of EGCG on a HIF-1$\alpha$ and VEGF in A549 cell line. Methods: A549 cells, a non-small cell lung cancer cell line, were cultured with DMEM and 10% fetal bovine serum. A decrease in oxygen tension was induced using a hypoxia microchamber and a $CO_2-N_2$ gas mixture. Gas analysis and a MTT assay were performed. The A549 cells were treated with EGCG (0, 12.5, 25, 50 ${\mu}mol/L$), and then examined by real-time-PCR analysis of HIF-1$\alpha$, VEGF, and $\beta$-actin mRNA. Results: Hypoxia reduced the proliferation of A549 cells from normoxic conditions. EGCG inhibited HIF-1$\alpha$ transcription in A549 cells in a dose-dependent manner. Compared to HIF-1$\alpha$, VEGF was not inhibited by EGCG. Conclusion: HIF-1$\alpha$ can be inhibited by EGCG. This suggests that targeting HIF-1$\alpha$ with a EGCG treatment may have therapeutic potential in non-small cell lung cancers.

• Asian-Australasian Journal of Animal Sciences
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• v.33 no.12
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• pp.2008-2020
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• 2020

Objective: This study was conducted to investigate the effects of dietary corn resistant starch (RS) on the intestinal morphology and barrier functions of broilers. Methods: A total of 320 one-day-old broilers were randomly allocated to 5 dietary treatments: one normal corn-soybean (NC) diet, one corn-soybean-based diet supplementation with 20% corn starch (CS), and 3 corn-soybean-based diets supplementation with 4%, 8%, and 12% corn resistant starch (RS) (identified as 4% RS, 8% RS, and 12% RS, respectively). Each group had eight replicates with eight broilers per replicate. After 21 days feeding, one bird with a body weight (BW) close to the average BW of their replicate was selected and slaughtered. The samples of duodenum, jejunum, ileum, caecum digesta, and blood were collected. Results: Birds fed 4% RS, 8% RS and 12% RS diets showed lower feed intake, BW gain, jejunal villus height (VH), duodenal crypt depth (CD), jejunal VH/CD ratio, duodenal goblet cell density as well as mucin1 mRNA expressions compared to the NC group, but showed higher concentrations of cecal acetic acid and butyric acid, percentage of jejunal proliferating cell nuclear antigen-positive cells and delta like canonical Notch ligand 4 (Dll4), and hes family bHLH transcription factor 1 mRNA expressions. However, there were no differences on the plasma diamine oxidase activity and D-lactic acid concentration among all groups. Conclusion: These findings suggested that RS could suppress intestinal morphology and barrier functions by activating Notch pathway and inhibiting the development of goblet cells, resulting in decreased mucins and tight junction mRNA expression.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 2005.06a
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• pp.154-164
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• 2005

Saccharomyces cerevisiae KNU5377 is a thermotolerant strain, which can ferment ethanol from wasted papers and starch at 40$^{\circ}C$ with the almost same rate as at 30$^{\circ}C$. This strain showed alcohol fermentation ability to convert wasted papers 200 g (w/v) to ethanol 8.4% (v/v) at 40$^{\circ}C$, meaning that 8.4% ethanol is acceptable enough to ferment in the industrial economy. As well, all kinds of starch that are using in the industry were converted into ethanol at 40$^{\circ}C$ with the almost same rate as at 30$^{\circ}C$. Hyperthermic cell killing kinetics and differential scanning calorimetry (DSC) revealed that exponentially growing cells of this yeast strain KNU5377 were more thermotolerant than those of S. cerevisiae ATCC24858 used as a control. This intrinsic thermotolernace did not result from the stability of entire cellular components but possibly from that of a particular target. Heat shock induced similar results in whole cell DSC profiles of both strains and the accumulation of trehalose in the cells of both strains, but the trehalose contents in the strain KNU5377 were 2.6 fold higher than that in the control strain. On the contrary to the trehalose level, the neutral trehalase activity in the KNU5377 cells was not changed after the heat shock. This result made a conclusion that though the trehalose may stabilize cellular components, the surplus of trehalose in KNU5377 strain was not essential for stabilization of whole cellular components. A constitutively thermotolerant yeast, S. cerevisiae KNU5377, was compared with a relatively thermosensitive control, S. cerevisiae ATCC24858, by assaying the fluidity and proton ATPase on the plasma membrane. Anisotropic values (r) of both strains were slightly increased by elevating the incubation temperatures from 25$^{\circ}C$ to 37$^{\circ}C$ when they were aerobically cultured for 12 hours in the YPD media, implying the membrane fluidity was decreased. While the temperature was elevated up to 40$^{\circ}C$, the fluidity was not changed in the KNU5377 cell, but rather increased in the control. This result implies that the plasma membrane of the KNU5377 cell can be characterized into the more stabilized state than control. Besides, heat shock decreased the fluidity in the control strain, but not in the KNU5377 strain. This means also there's a stabilization of the plasma membrane in the KNU5377 cell. Furthermore, the proton ATPase assay indicated the KNU5377 cell kept a relatively more stabilized glucose metabolism at high temperature than the control cell. Therefore, the results were concluded that the stabilization of plasma membrane and growth at high temperature for the KNU5377 cell. Genome wide transcription analysis showed that the heat shock responses were very complex and combinatory in the KNU5377 cell. Induced by the heat shock, a number of genes were related with the ubiquitin mediated proteolysis, metallothionein (prevent ROS production from copper), hsp27 (88-fold induced remarkably, preventing the protein aggregation and denaturation), oxidative stress response (to remove the hydrogen peroxide), and etc.

• Journal of Life Science
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• v.26 no.12
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• pp.1383-1391
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• 2016

Anti-estrogen drugs such as tamoxifen have been used for treating patients with ER-positive, early breast cancer. However, resistance to anti-estrogen treatment is inevitable in most patients. Breast cancer anti-estrogen resistance-3 (BCAR3) has been identified as the protein responsible for the induction of tamoxifen resistance in estrogen-dependent human breast cancer. We have previously reported that BCAR3 regulates the cell cycle progression and the signaling pathway of EGF and insulin leading to DNA synthesis. In this study, we investigated the functional role of BCAR3 in regulating c-Jun transcription in non-tumorigenic human breast epithelial MCF-12A cells. A transient transfection of BCAR3 increased both the mRNA and protein of c-Jun expression, and stable expression of BCAR3 increased c-Jun protein expression. The overexpression of BCAR3 directly activated the promoter of c-jun, AP-1, and SRE but not that of $NF-{\kappa}B$. Furthermore, single-cell microinjection of BCAR3 expression plasmid in the cell cycle-arrested MCF-12A cells induced c-Jun protein expression, and co-injection of dominant negative mutants of Ras, Rac, and Rho suppressed the transcriptional activity of c-Jun in the presence of BCAR3. Furthermore, stable expression of BCAR3 increased the proliferation of MCF-12A cells. The microinjection of inhibitory materials such as anti-BCAR3 antibody and siRNA BCAR3 inhibited EGF-induced c-Jun expression but did not affect IGF-1 induced upregulation of c-Jun. Taken together, we propose that BCAR3 plays a crucial role in c-Jun protein expression and cell proliferation and that small GTPases (e.g., Ras, Rac, and Rho) are required for the BCAR3-mediated activation of c-Jun expression.