• The Journal of Internal Korean Medicine
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• v.25 no.4
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• pp.18-24
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• 2004

연구배경 : 플라스미노겐 활성 억제인자-1 (plasminogen activator inhibitor-1; PAI-1)은 허혈성 뇌경색의 발생의 원인이 되는 섬유소 용해작용의 저하를 매개하는 인자로서, PAI-1의 작용이 촉진되면 섬유소 용해기능이 저하되어 관상동맥 및 뇌혈관질환의 발생을 증가시키게 된다. PAI-1 유전자의 촉진자(promoter) 영역에는 -675 4G/5G (4G/5G)와A -844G (A/G)의 두 개의 유전자 다형성이 존재하며, 이는 PAI-1의 유전자 전사과정에 영향을 미쳐 혈청 PAI-1의 농도를 증가시키고 결과적으로 허혈성 뇌경색의 발생확률을 높이는 작용을 하게 된다. 연구방법 : 허혈성 뇌경색으로 진단 받은 167명의 환자와 173명의 건강인의 말초혈액에서 DNA를 분리한 후 PAI-1의 4G/5G와 A/G 유전자 다형성에 대한 연쇄중합반응 및 제한효소 절편길이 다형성 (polymerase chain reaction-restriction fragment length polymorphism; PCR-RFLP) 방법을 이용하여 허혈성 뇌경색 발생과 유전자 다형성과의 관계를 비교 분석하였다. 결과 : 허혈성 뇌경색 환자에서의 4G/4G의 유전자형의 빈도는 15.0%으로 정상 대조군의 33.5%에 비해 현저하게 낮게 나타났다 (P < 0.0001). 각각의 유전자형과 허혈성 뇌경색의 발생 위험도 (odd ratio ; OR)와의 관계를 분석했을 때 4G/4G 유전자형을 가질 경우 위험도는 0.35배로 현저하게 낮아졌으며, (P < 0.0001), 5G/5G 유전자형을 가질 경우 위험도는 4.49배 로 현저하게 높아졌다 (P < 0.0001). 그러나, A/G 유전자 다형성과 허혈성 뇌경색의 발생과는 유의한 연관성을 발견하지 못하였다. 결론 : 이상의 결과로 볼 때 PAI-1 유전자의 4G/4G 유전자형은 허혈성 뇌경색의 발생 비율을 감소시키는 작용을 하는 것으로 여겨진다.

• Journal of Life Science
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• v.15 no.3 s.70
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• pp.397-405
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• 2005

Sulforaphane, an isothiocyanate derived from hydrolysis of glucoraphanin in broccoli and other cruciferous vegetables, was shown to induce phase II detoxification enzymes and inhibit chemically induced mammary tumors in rodents. Recently, sulforaphane is known to induce cell cycle arrest and apoptosis in human cancer cells, however its molecular mechanisms are poorly understood. In the present study, we demonstrated that sulforaphane acted to inhibit proliferation and induce morphological changes of human cervical carcinoma HeLa cells. Treatment of HeLa cells with $10{\mu}M\;or\;15{\mu}M$ sulforaphane resulted in significant G2/M cell cycle arrest as determined by flow cytometry. Moreover, $20{\mu}M$ sulforaphane significantly induced the population of sub-G1 cells (9.83 fold of control). This anti-proliferative effect of sulforaphane was accompanied by a marked inhibition of cyclin A and cyclin-dependent kinase (Cdk)4 protein and concomitant induction of Cdc2, Cdk inhibitor p16 and p21. However, sulforaphane did not affect the levels of cyelooxygenases and telomere-regulatory gene products. Although further studies are needed, the present work suggests that sulforaphane may be a potential chemoprevetive/ chemotherapeutic agent for the treatment of human cancer cells.

• Restorative Dentistry and Endodontics
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• v.30 no.5
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• pp.372-384
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• 2005

The purpose of this study was to monitor the secretion of matrix metalloproteinase (MMP) and tissue inhibitor of metalloproteinase (TIMP) by human periodontal ligament (PDL) fibroblasts stimulated with Prevotella nigrescens lipopolysaccharide (LPS), and to examine the effect of calcium hydroxide treatment on P. nigrescens LPS. LPS was extracted and purified from anaerobically cultured P. nigrescens. PDL fibroblasts were stimulated by the LPS (0, 0.1, 1, 10 ${\mu}g/ml$) or LPS (10 ${\mu}g/ml$) pretreated with 12.5 mg/ml of $Ca(OH)_2$ for 3 days, for various periods of time (12, 24, 48 h). Immunoprecipitation were performed for protein level analysis of MMP-1 MMP-2 and TIMP-1. Total RNA was isolated and real-time quantitative polymerase chain reaction (PCR) was performed for quantification of MMP-1 mRNA. According to this study, the results were as follows: 1. The p개duction of MMP-1 by stimulation with P. nigrescens LPS increased in time-dependent manner, and showed maximum value at 48 h in both protein and mRNA level. But there was no dose-dependent increas. 2. MMP-2 production time-dependently increased when stimulated with 1 and 10 ${\mu}g/ml$LPS, but there was no dose-dependent increase. 3. TIMP-1 p개duction increased to 24 h, but decreased at 48 h. It increased when stimulated with 0.1 and 1${\mu}g/ml$, but suppressed at 10 ${\mu}g/ml$ .4. P. nigrescens LPS pretreated with $Ca(OH)_2$ markedly downregulated MMP-1 gene expression.

• Journal of Life Science
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• v.16 no.7 s.80
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• pp.1235-1242
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• 2006

Sulforaphane, an isothiocyanate derived from hydrolysis of glucoraphanin in broccoli and other cruciferous vegetables, was shown to induce phase II detoxification enzymes and inhibit chemically induced mammary tumors in rodents. Recently, sulforaphane is known to induce cell cycle arrest and apoptosis in human canter cells, however its molecular mechanisms are poorly understood. In tile present study, we demonstrated that sulforaphane acted to inhibit proliferation and induce morphological changes of human hepatocarcinoma HepG2 cells. Treatment of HepG2 cells with $10{\mu}M\;or\;15{\mu}M$ sulforaphane resulted in significant G2/M cell cycle arrest as determined by DNA flow cytometry. Moreover, $20{\mu}M$ sulforaphane significantly induced the population of sub-G1 cells suggesting that sulforaphane induced apoptosis. This anti-proliferative effect of sulforaphane was accompanied by a marked inhibition of ryclin A, cyclin 31 and Cdc2 protein. However, the levels of tumor suppressor p53 and Cdk inhibitor p21 mRNA and protein expression were significantly increased by sulforaphane treatment in a concentration-dependent manner. Although further studies are needed, the present work suggests that sulforaphane may be a potential rhemoprevetiveichemotherapeucc agent for the treatment of human cancer cells.

• Reproductive and Developmental Biology
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• v.37 no.4
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• pp.269-279
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• 2013

Low efficiency of somatic cell nuclear transfer (SCNT) is attributed to incomplete reprogramming of transfered nuclei into oocytes. Trichostatin A (TSA), histone deacetylase inhibitor and 5-aza-2'deoxycytidine (5-aza-dC), DNA methylation inhibitor has been used to enhance nuclear reprogramming following SCNT. However, it was not known molecular mechanism by which TSA and 5-aza-dC improve preimplantation embryo and fetal development following SCNT. The present study investigates embryo viability and gene expression of cloned porcine preimplantation embryos in the presence and absence of TSA and 5-aza-dC as compared to embryos produced by parthenogenetic activation. Our results indicated that TSA treatment significantly improved development. However 5-aza-dC did not improve development. Presence of TSA and 5-aza-dC significantly improved total cell number, and also decreased the apoptotic and autophagic index. Three apoptotic-related genes, Bak, Bcl-xL, and Caspase 3 (Casp3), and three autophagic-related genes, ATG6, ATG8, and lysosomal-associated membrane protein 2 (LAMP2), were measured by real time RT-PCR. TSA and 5-aza-dC treatment resulted in high expression of anti-apoptotic gene Bcl-xL and low pro-apoptotic gene Bak expression compared to untreated NT embryos or parthenotes. Furthermore, LC3 protein expression was lower in NT-TSA and NT-5-aza-dC embryos than those of NT and parthenotes. In addition, TSA and 5-aza-dC treated embryos displayed a global acetylated histone H3 at lysine 9 and methylated DNA H3 at lysine 9 profile similar to the parthenogenetic blastocysts. Finally, we determined that several DNA methyltransferase genes Dnmt1, Dnmt3a and Dnmt3b. NT blastocysts showed higher levels Dnmt1 than those of the TSA and 5-aza-dC blastocysts. Dnmt3a is lower in 5-aza-dC than NT, NTTSA and parthenotes. However, Dnmt3b is higher in 5-aza-dC than NT and NTTSA. These results suggest that TSA and 5-aza-dC positively regulates nuclear reprogramming which result in modulation of apoptosis and autophagy related gene expression and then reduce apoptosis and autophagy. In addition, TSA and 5-aza-dC affects the acetylated and methylated status of the H3K9.

• The Korean Journal of Physiology and Pharmacology
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• v.16 no.6
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• pp.393-398
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• 2012

Mast cells are involved in allergic responses, protection against pathogens and autoimmune diseases. Dexamethasone (Dex) and other glucocorticoids suppress $Fc{\varepsilon}RI$-mediated release of inflammatory mediators from mast cells. The inhibition mechanisms were mainly investigated on the downstream signaling of Fc receptor activations. Here, we addressed the effects of Dex on Fc receptor expressions in rat mast cell line RBL-2H3. We measured mRNA levels of Fc receptors by real-time PCR. As expected, Dex decreased the mRNA levels of activating Fc receptor for IgE ($Fc{\varepsilon}R$) I and increased the mRNA levels of the inhibitory Fc receptor for IgG $Fc{\gamma}RIIb$. Interestingly, Dex stimulated transcriptions of other activating receptors such as Fc receptors for IgG ($Fc{\gamma}R$) I and $Fc{\gamma}RIII$. To investigate the mechanisms underlying transcriptional regulation, we employed a transcription inhibitor actinomycin D and a translation inhibitor cycloheximide. The inhibition of protein synthesis without Dex treatment enhanced $Fc{\gamma}RI$ and $Fc{\gamma}RIII$ mRNA levels potently, while $Fc{\varepsilon}RI$ and $Fc{\gamma}RIIb$ were minimally affected. Next, we examined expressions of the Fc receptors on cell surfaces by the flow cytometric method. Only $Fc{\gamma}RIIb$ protein expression was significantly enhanced by Dex treatment, while $Fc{\gamma}RI$, $Fc{\gamma}RIII$ and $Fc{\varepsilon}RI$ expression levels were marginally changed. Our data showed, for the first time, that Dex regulates Fc receptor expressions resulting in augmentation of the inhibitory receptor $Fc{\gamma}RIIb$.

• Korean Journal of Organic Agriculture
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• v.26 no.4
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• pp.775-787
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• 2018

This study was conducted to investigate for the natural methane emission inhibitor as a feed additive no adversely effect on rumen fermentation. Five different Control (Wheat barn (0.05 g), MRA(Methane Reduction Additive)-1 (Allium fistulosum L. (0.05 g)), MRA-2 (Sodium Lauryl Sulfate (0.025 g) + Wheat barn (0.025 g) mixed), MRA-3 (Sodium Dodecyl Sulfate (0.025 g) + Wheat barn (0.025 g) mixed), and MRA-4 (Allium fistulosum L. (0.02 g) + Tannic acid (0.02 g) + Wheat barn (0.01 g) mixed) contents were used to perform 3, 6, 9, 12, 24 and 48 h incubation for in vitro fermentation. Ruminal pH values were ranged within normal ruminal microbial fermentation. Dry matter digestibility was not significantly different across the treatments during the whole fermentation time. Also, the result of microbial growth had no adversely effect on during the whole fermentation time. At 24 h, methane emission was significantly lower (P<0.05) than all treatments except to MRA-1. Especially, MRA-4 carbon dioxide emission was significantly lower (P<0.05) than control at 9, 24 and 48 h incubation. In addition MRA-4 propionate concentration was significantly higher (P<0.05) than control at 24 h incubation. The result of RT-PCR Ciliate-associated methanogens were significantly lower (P<0.05) at MRA-1, MRA-3 and MRA-4 than control at 24 h incubation. Based on the present results, MRA-4 could be suggestible methane emission inhibitor as a natural feed additive.

• Journal of Ginseng Research
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• v.48 no.3
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• pp.298-309
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• 2024

Background: 20(S)-ginsenoside Rh2(GRh2), an effective natural histone deacetylase inhibitor, can inhibit acute myeloid leukemia (AML) cell proliferation. Lactate regulated histone lactylation, which has different temporal dynamics from acetylation. However, whether the high level of lactylation modification that we first detected in acute promyelocytic leukemia (APL) is associated with all-trans retinoic acid (ATRA) resistance has not been reported. Furthermore, Whether GRh2 can regulate lactylation modification in ATRA-resistant APL remains unknown. Methods: Lactylation and METTL3 expression levels in ATRA-sensitive and ATRA-resistant APL cells were detected by Western blot analysis, qRT-PCR and CO-IP. Flow cytometry (FCM) and APL xenograft mouse models were used to determine the effect of METTL3 and GRh2 on ATRA-resistance. Results: Histone lactylation and METTL3 expression levels were considerably upregulated in ATRA-resistant APL cells. METTL3 was regulated by histone lactylation and direct lactylation modification. Overexpression of METTL3 promoted ATRA-resistance. GRh2 ameliorated ATRA-resistance by downregulated lactylation level and directly inhibiting METTL3. Conclusions: This study suggests that lactylation-modified METTL3 could provide a promising strategy for ameliorating ATRA-resistance in APL, and GRh2 could act as a potential lactylation-modified METTL3 inhibitor to ameliorate ATRA-resistance in APL.

• Journal of Life Science
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• v.22 no.4
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• pp.492-498
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• 2012

To investigate whether phytochemicals affect cancer cell viability, human colorectal HCT116 cells were treated with four different phytochemicals. Among these phytochemicals, curcumin is the strongest inhibitor of cell proliferation. In addition, it decreased cell viability in a dose-dependent manner. To unveil the molecular mechanisms involved in the inhibition of cell proliferation by curcumin, we carried out oligo DNA microarray analysis. We found that 137 genes were up-regulated more than 2-fold, and 141 genes were down-regulated more than 2-fold by 25 ${\mu}M$ curcumin treatment. Among the up-regulated genes, we selected 3 genes (ATF-3, GADD45A, and NR4A1) to confirm microarray data. The results of RT-PCR strongly agreed with those of the microarray data. Among the phytochemicals used in this study, curcumin is the strongest inducer of ATF3 expression, and increased ATF3 expression in a dose-dependent manner. Interestingly, FACS analysis showed that the inhibition of cell growth by curcumin was recovered by ATF3-siRNA transfection. Finally, we detected the changes of gene expression by ectopic expression of ATF3. The results indicated that many up-regulated genes were related to apoptosis. Overall, these results suggest that ATF3 may play an important role in the anti-proliferative activity of curcumin in human colorectal cancer cells.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.11
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• pp.4555-4561
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• 2014

Background: Silencing due to methylation of suppressor of cytokine signaling-3 (SOCS-3), a negative regulator gene for the JAK/STAT signaling pathway has been reported to play important roles in leukemogenesis. Imatinib mesylate is a tyrosine kinase inhibitor that specifically targets the BCR-ABL protein and induces hematological remission in patients with chronic myeloid leukemia (CML). Unfortunately, the majority of CML patients treated with imatinib develop resistance under prolonged therapy. We here investigated the methylation profile of SOCS-3 gene and its downstream effects in a BCR-ABL positive CML cells resistant to imatinib. Materials and Methods: BCR-ABL positive CML cells resistant to imatinib (K562-R) were developed by overexposure of K562 cell lines to the drug. Cytotoxicity was determined by MTS assays and $IC_{50}$ values calculated. Apoptosis assays were performed using annexin V-FITC binding assays and analyzed by flow cytometry. Methylation profiles were investigated using methylation specific PCR and sequencing analysis of SOCS-1 and SOCS-3 genes. Gene expression was assessed by quantitative real-time PCR, and protein expression and phosphorylation of STAT1, 2 and 3 were examined by Western blotting. Results: The $IC_{50}$ for imatinib on K562 was 362nM compared to 3,952nM for K562-R (p=0.001). Percentage of apoptotic cells in K562 increased upto 50% by increasing the concentration of imatinib, in contrast to only 20% in K562-R (p<0.001). A change from non-methylation of the SOCS-3 gene in K562 to complete methylation in K562-R was observed. Gene expression revealed down-regulation of both SOCS-1 and SOCS-3 genes in resistant cells. STAT3 was phosphorylated in K562-R but not K562. Conclusions: Development of cells resistant to imatinib is feasible by overexposure of the drug to the cells. Activation of STAT3 protein leads to uncontrolled cell proliferation in imatinib resistant BCR-ABL due to DNA methylation of the SOCS-3 gene. Thus SOCS-3 provides a suitable candidate for mechanisms underlying the development of imatinib resistant in CML patients.