• Toxicological Research
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• v.31 no.1
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• pp.11-16
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• 2015

Our body's immune system has defense mechanisms against pathogens such as viruses and bacteria. Immune responses are primarily initiated by the activation of toll-like receptors (TLRs). In particular, TLR4 is well-characterized and is known to be activated by gram-negative bacteria and tissue damage signals. TLR4 requires myeloid differentiation factor 2 (MD2) as a co-receptor to recognize its ligand, lipopolysaccharides (LPS), which is an extracellular membrane component of gram-negative bacteria. Gambogic acid is a xanthonoid isolated from brownish or orange resin extracted from Garcinia hanburyi. Its primary effect is tumor suppression. Since inflammatory responses are related to the development of cancer, we hypothesized that gambogic acid may regulate TLR4 activation. Our results demonstrated that gambogic acid decreased the expression of pro-inflammatory cytokines ($TNF-{\alpha}$, IL-6, IL-12, and $IL-1{\beta}$) in both mRNA and protein levels in bone marrow-derived primary macrophages after stimulation with LPS. Gambogic acid did not inhibit the activation of Interferon regulatory factor 3 (IRF3) induced by TBK1 overexpression in a luciferase reporter gene assay using IFN-${\beta}$-PRD III-I-luc. An in vitro kinase assay using recombinant TBK1 revealed that gambogic acid did not directly inhibit TBK1 kinase activity, and instead suppressed the binding of LPS to MD2, as determined by an in vitro binding assay and confocal microscopy analysis. Together, our results demonstrate that gambogic acid disrupts LPS interaction with the TLR4/MD2 complex, the novel mechanism by which it suppresses TLR4 activation.

• BMB Reports
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• v.55 no.2
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• pp.98-103
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• 2022

Increased mRNA levels of cancer upregulated gene (CUG)2 have been detected in many different tumor tissues using Affymetrix microarray. Oncogenic capability of the CUG2 gene has been further reported. However, the mechanism by which CUG2 overexpression promotes cancer stem cell (CSC)-like phenotypes remains unknown. With recent studies showing that pyruvate kinase muscle 2 (PKM2) is overexpressed in clinical tissues from gastric, lung, and cervical cancer patients, we hypothesized that PKM2 might play an important role in CSC-like phenotypes caused by CUG2 overexpression. The present study revealed that PKM2 protein levels and translocation of PKM2 into the nucleus were enhanced in CUG2-overexpressing lung carcinoma A549 and immortalized bronchial BEAS-2B cells than in control cells. Expression levels of c-Myc, CyclinD1, and PKM2 were increased in CUG2-overexpressing cells than in control cells. Furthermore, EGFR and ERK inhibitors as well as suppression of Yap1 and NEK2 expression reduced PKM2 protein levels. Interestingly, knockdown of β-catenin expression failed to reduce PKM2 protein levels. Furthermore, reduction of PKM2 expression with its siRNA hindered CSC-like phenotypes such as faster wound healing, aggressive transwell migration, and increased size/number of sphere formation. The introduction of mutant S37A PKM2-green fluorescence protein (GFP) into cells without ability to move to the nucleus did not confer CSC-like phenotypes, whereas forced expression of wild-type PKM2 promoted such phenotypes. Overall, CUG2-induced increase in the expression of nuclear PKM2 contributes to CSC-like phenotypes by upregulating c-Myc and CyclinD1 as a co-activator.

• The Journal of Korean Medicine
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• v.32 no.3
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• pp.35-43
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• 2011

Objectives: This study was performed to screen target-specific inhibitors of ${\beta}$-catenin/TCF signaling whose functional activation plays an important role in early events in carcinogenesis. Methods: To investigate the activation or suppression of ${\beta}$-catenin/TCF transcription, we established a transiently transfected cell line with a constitutively active ${\beta}$-catenin mutant gene whose product is not degraded. This cell line was also co-transfected with luciferase reporter gene constructs containing either an optimized (TOPflash) or mutant (FOPflash) TCF-binding element. We investigated cytotoxic effects using a 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium salt (MTS) assay. To find effective inhibitors of ${\beta}$-catenin/TCF signaling from medicinal herbs, the crude extracts of 99 types of medicinal herbs were screened using a luciferase assay system in HEK-293 and SH-SY5y cells. Results: At a concentration of $50{\mu}g$/ml, extracts of Angelica koreanae radix, Cannabis sativa semen, Ephedrae intermedia Schrenk radix, and Vitis rotundifolia fruit showed the following inhibitory effects on ${\beta}$-catenin/TCF signaling: $40{\pm}5.6%$, $23{\pm}6.1%$, $8{\pm}5.1%$, and $22{\pm}9.8%$ in ${\beta}$-catenin-activated HEK-293 cells and $9{\pm}4.7%$, $39{\pm}8.1%$, $39{\pm}6.4%$, and $42{\pm}10.1%$ in ${\beta}$-catenin-activated SH-SY5y cells, respectively. Crude extracts of E. radix were isolated by silica gel column chromatography, and two non-polar fractions of these extracts showed inhibitory effects on ${\beta}$-catenin/TCF signaling. Conclusions: In this study, we established a transiently transfected cell line as a screening system and found that various medicinal herb extracts had inhibitory effects on ${\beta}$signaling.

• Korean Journal of Poultry Science
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• v.43 no.1
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• pp.47-54
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• 2016

The aim of this study was to investigate the expression patterns of key genes involved in lipid metabolism in response to dietary Coenzyme Q10 (CoQ10) in hens. A total of 36 forty week-old Lohmann Brown were randomly allocated into 3 groups consisting of 4 replicates of 3 birds. Laying hens were subjected to one of following treatments: Control (BD, basal diet), T1 (BD+ CoQ10 100 mg/kg diet) and T2 (BD+ micellar of CoQ10 100 mg/kg diet). Birds were fed ad libitum a basal diet or the basal diet supplemented with CoQ10 for 5 weeks. Total RNA was extracted from the liver for quantitative RT-PCR. The mRNA levels of HMG-CoA reductase(HMGCR) and sterol regulatory element-binding proteins(SREBP)2 were decreased more than 30~50% in the liver of birds fed a basal diet supplemented with CoQ10 (p<0.05). These findings suggest that dietary CoQ10 can reduce cholesterol levels by the suppression of the hepatic HMGCR and SREBP2 genes. The gene expressions of liver X receptor (LXR) and SREBP1 were down regulated due to the addition of CoQ10 to the feed (p<0.05). The homeostasis of cholesterol can be regulated by LXR and SREBP1 in cholesterol-low-conditions. The supplement of CoQ10 caused a decreased expression of lipid metabolism-related genes including $PPAR{\gamma}$, XBP1, FASN, and GLUTs in the liver of birds (p<0.05). These data suggest that CoQ10 might be used as a dietary supplement to reduce cholesterol levels and to regulate lipid homeostasis in laying hens.

• Journal of Life Science
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• v.22 no.10
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• pp.1316-1323
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• 2012

The present study was conducted to investigate whether myoblasts can be transdifferentiated into adipocytes by ectopic expression of adipogenic transcription factors, including peroxisome proliferator-activated receptor-${\gamma}$ ($PPAR{\gamma}$), CCAAT/enhancer-binding protein-${\alpha}$ (C/$EBP{\alpha}$), sterol regulatory element binding protein-1c (SREBP1c), and Krueppel-like factor 5 (KLF5), in primary bovine satellite cells. Transcription factors were transiently transfected into primary bovine myoblasts, and the cells were cultured with adipogenic differentiation medium for 2 days and then cultured on growth medium for an additional 8 days. Ectopic expression of $PPAR{\gamma}$ or C/$EBP{\alpha}$ alone was insufficient to induce adipogenesis in myoblasts. However, overexpression of both $PPAR{\gamma}$ and C/$EBP{\alpha}$ in myoblasts was able to induce adipogenic transdifferentiation as indicated by the appearance of mature adipocytes, the induction of adipogenic gene expressions, and the suppression of myogenic gene expressions. In addition, KLF5 and $PPAR{\gamma}$ co-transfected bovine myoblasts were converted to adipocytes but not in cells transfected with only KLF5 expression vector. Overexpression of SREBP1c alone was sufficient to induce transdifferentiation from myoblasts into adipocytes. These results demonstrate that primary bovine satellite cells can be transdifferentiated into adipocytes either by single ectopic expression or combined expression of adipogenic transcription factors in a culture system.

• The Journal of Korean Medicine
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• v.38 no.3
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• pp.43-58
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• 2017

Objectives: We performed this study to evaluate the antioxidative and hypolipidemic effect of Artemisia iwayomogi (韓茵蔯), Curcuma longa L. (鬱金) and Raphanus sativus L. (蘿?子) (ACR). Method: We enriched Artemisiae Capillaris, Curcumae Longae and Raphani Semen compound with alcohol. ACR extract is treated to HepG2 cell. Cell groups are devided into 3 groups: normal, control and ACR treated group. We measured polyphenol, flavonoids, DPPH and ABTS radical scavenging activity, ROS, glutathione, GSH peroxidase, GSH reductase, SOD, catalase, free fatty acid, lipid peroxidation and suppression of ACAT1 and HMG-CoA reductase expression on mRNA level. Results: 1. ACR contained polyphenol and flavonoids and increased GSH significantly in HepG2 cell. 2. ACR increased GPx, GR, and catalase activity significantly in HepG2 cell. 3. ACR increased DPPH and ABTS radical scavenging activity significantly in HepG2 cell and decreased ROS. 4. ACR decreased free fatty acid and MDA significantly in HepG2 cell. 5. ACR suppressed ACAT1 and HMG-CoA reductase expression on mRNA level in HepG2 cell. Conclusion: This study suggests that ACR has antioxidative and hypolipidemic effect and might be effective in prevention and treatment of dyslipidemia.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.15 no.9
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• pp.5628-5636
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• 2014

Currently, many countries have an interest in developing cosmetics materials using native plants. In this aspect, there is increasing need to develop cosmetics materials using native plants in our county. In the present study, calluses were induced from Artemisia annua Linne, which was highlighted because of its useful effects, such as anti-cancer, anti-fungal and anti-inflammation. Water and ethanol extractions were performed from the calluses of Artemisia annua Linne. After the mass production of Artemisia annua Linne's calluses, water and ethanol extraction was performed to examine its functional roles in healing wounds and inflammation. The differences in the effective elements were observed in the ethanol extract. The callus showed anti-inflammation activity through the suppression of the inflammation-related gene, COX-2, and ethanol extracts showed their ability to heal wounds. Overall, these results suggest that the extract of Artemisia annua Linne's calluses is a natural and environment-friendly material, and can be used as medical supplies associated with anti-inflammation and healing wounds.

• Journal of Microbiology and Biotechnology
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• v.14 no.5
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• pp.1075-1080
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• 2004

Dextransucrase (DSRB742) from Leuconostoc mesenteroides NRRL B-742CB is a glucosyltransferase that catalyzes the synthesis of dextran using sucrose, or the synthesis of oligosaccharides when acceptor molecules, like maltose, are present. The DSRB742 gene (dsrB742) was cloned and the properties were characterized. In order to identify critical amino acid residues, the DSRB742 amino acid sequence was aligned with glucosyltransferase sequences, and three amino acid residues reported as sucrose binding amino acids in Streptococcus glucosyltransferases were selected for site-directed mutagenesis experiments. Asp-533, Asp-536, and His-643 were independently replaced with Ala or Asn. D533A and D536A dextransucrases showed reduced dextran synthesis activities, 2.3% and 40.8% of DSRB742 dextransucrase, respectively, and D533N, D536N, H643A, end H643N dextransucrases showed complete suppression of dextran synthesis activities altogether. Additionally, D536N dextransucrase showed complete suppression of oligosaccharide synthesis activities. However, modifications at Asp-533 or at His-643 retained acceptor reaction activities in the range of 8.4% to 21.3% of DSRB742 acceptor reaction activity. Thus at least two carboxyl groups of Asp-533 and Asp-536, and His-643 as a proton donor, are essential for the catalysis process.

• Journal of Acupuncture Research
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• v.23 no.2
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• pp.61-72
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• 2006

Objectives : Human chondrocytes co-treated with Buthus martensi Karsch herbal acupuncture solution(BMK-HAS) extract produced significantly less NO compared with chondrocytes stimulated with $IL-1{\beta}$ alone Methods : Activation and translocation of and NF-kB DNA binding activity were determined by Western blotting and specific enzyme-linked immunosorbent assay. Results : The inhibition of NO production correlated with the suppression of induction and expression of nuclear factor-kB (NF-kB) and activation protein-1 (AP-1)-dependent gene. BMK-HAS inhibited the activation and translocation of NF-kB to the nucleus, indicating that BMK-HAS inhibits the $IL-1{\beta}-induced$ production of NO in human chondrocytes by interfering with the activation of NF-kB through a novel mechanism. In addition, BMK-HAS reduced prostaglandin E2 (PGE2)production in mouse peritoneal macrophages stimulated with lipopolysaccharide, whereas no influence on the activity of inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2) or cyclooxygenase-1 (COX-1) was observed. My data, therefore, suggest that BMK-HAS may be a therapeutically effective inhibitor of $IL-1{\beta}-induced$ inflammatory effects that are dependent on NF-kB activation in human OA chondrocytes. Conclusion : The results indicate that BMK-HAS exerts anti-inflammatory effects related to the inhibition of neutrophil functions and of NO and PGE2 production, which could be due to a decreased expression of iNOS and COX-2 through the transcription factors NF-kB and AP-1.

• Development and Reproduction
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• v.13 no.4
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• pp.297-303
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• 2009

Implantation of blastocyst into the uterine endometrium is established by the existence of histologically and functionally prepared uterine endometrium. Doc-1, an oral cancer suppressor gene, is expressed under the control of steroid hormones and has been suggested as a proliferation regulator of endometrial cells. However, the role is not much clear and in this study we examined the expression modulation of Doc-1 in decidualizing cells in vitro. In vitro decidualization was performed in endometrial stroma cells using progesterone and estrogen. Until 24 hr after decidual induction the proliferation of stroma cell was significantly increased but decreased after then. On the other hand, most of the cells differentiated into decidual cell after 48 hr of induction. The Doc-1 protein was co-localized in a specific deciudal cells and colocalization rate was increased in a parallel manner with the induction time. Based on these results, it is suggested that Doc-1 expression is under the control of both steroid hormones and decidual signals, and Doc-1 protein is involved in suppression of the proliferation of decidualizing cells.