• Journal of the Korean Applied Science and Technology
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• v.35 no.4
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• pp.1003-1012
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• 2018

This study is for checking the possibility of Lentinula edodes as cosmetic materials. For this we carried out biological active evaluation about anti-oxidant and anti-inflammatory effects by Lentinula edodes extracts. We extracted Lentinula edodes with water and 70% ethanol and then in order to evaluate anti-oxidant activity we treated samples by concentrations (100, 500, 1000) ${\mu}g/ml$ and carried out 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging, the activity of 2,2'-azino-bis ( 3-ethylbenzothiazoline-6-sulphonic acid )-diammonium salt (ABTS) cation radical scavenging and superoxide dismutase(SOD) like activity. Also, in order to evaluate effect of anti-inflammatory the samples in macrophages(RAW 264.7 cells), we carried out evaluation of cell viability, nitric oxide inhibitory activity western blot. The results of DPPH, $ABTS^+$ radical scavenging activity and SOD-like activity of the Lentinula edodes extracts increased in dose-dependent manner. The cytotoxic of samples by MTT assay showed no toxicity at the concentrations of 10, 25 and $50{\mu}g/ml$ of Lentinula edodes extract. Nitric oxide inhibition activity results showed that the extracts reduced NO productions in a concentration-dependent manner. Expression of inflammatory cytokines as $TNF-{\alpha}$, $PGE_2$ and $IL-1{\beta}$ decreased in a concentration-dependent manner and iNOS and COX-2 proteins expression rates were decreased significantly in western blot analysis. From the results of the experiment, it was comfirmed that the Lentinula edodes extracts had excellent anti-oxidant and anti-inflammatory effect and could be used as a safe natural cosmetic material in the future.

• Journal of Life Science
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• v.29 no.4
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• pp.492-498
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• 2019

Previously, three kinds of lipases, lipAD1, lipAD2, and lipAD3, and lipase chaperone, lipBD, of Acinetobacter schindleri DYL129 isolated from soil sample were reported. In this report, three lipase and lipase chaperone were cloned into the pET32a(+) or pGEX-6P-1 vectors for protein expression in Escherichia coli, and named as pETLAD1, pETLAD2, pETLAD3 and pETLBD or pGEXLAD1, pGEXLAD 2, pGEXLAD3 and pGEXLBD, respectively. Protein expression rate was higher in pET system than in pGEX system. Although LipAD1 and LipAD2 were produced as inclusion bodies, their expression levels were high. So LipAD1 and LipAD2 could be solubilized in 8 M urea buffer and purified. LipAD3 and LipBD were overexpressed in soluble form and purified. Those proteins were purified by His-tag affinity chromatography connected in AKTA prime system. The activities of the purified lipases were demonstrated with 1% tributyrin agar plate. After purification, molecular mass was determined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis. LipAD1 showed high activity toward ${\rho}$-nitrophenyl acetate and ${\rho}$-nitrophenyl butyrate, LipAD2 showed high activity toward ${\rho}$-nitrophenyl acetate and ${\rho}$-nitrophenyl myristate, and LipAD3 showed high activity toward ${\rho}$-nitrophenyl acetate, ${\rho}$-nitrophenyl butyrate, and ${\rho}$-nitrophenyl miristate, respectively. Three lipases, LipAD1, LipAD2, and LipAD3, showed optimal reaction at $50^{\circ}C$ using ${\rho}$-nitrophenyl butyrate, as substrate.

• Journal of Life Science
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• v.28 no.12
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• pp.1416-1423
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• 2018

With strong biotin binding affinity ($K_D=10^{-14}M$), the tetrameric feature of streptavidin could be used to increase the antigen binding activity of a camel heavy chain (VHH) antibody through their fusion, here stained with biotinylated horseradish peroxidase and subsequent immunoassays ELISA and Western blot analysis. For this application, we cloned the streptavidin gene amplified from the Streptomyces avidinii chromosome by PCR, and this was fused to the gene of the 8B9 VHH antibody which is specific to green fluorescent protein (GFP) antigens. To express a soluble fusion protein in Escherichia coli, we used the pUC119 plasmid-based expression system which uses the lacZ promoter for induction by IPTG, the pelB leader sequence at the N-terminus for secretion into the periplasmic space, and six polyhistidine tags at the C-terminus for purification of the expressed proteins using an $Ni^+$-NTA-agarose column. Although streptavidin is toxic to E. coli because of its strong biotin binding property, this soluble fusion protein was expressed successfully. In SDS-PAGE, the size of the purified fusion protein was 122.4 kDa in its native condition and 30.6 kDa once denatured by boiling, suggesting the tetramerization of the monomeric subunit by non-covalent association through the streptavidin moiety fusing to the 8B9 VHH antibody. In addition, this fusion protein showed biotin binding activity similar to streptavidin as well as GFP antigen binding activity through both ELISA and Western blot analysis. In conclusion, the protein resulting from the fusion of an 8B9 VHH antibody with streptavidin was successfully expressed and purified as a soluble tetramer in E. coli; it showed both biotin and GFP antigen binding activity suggesting the possible production of a tetrameric and bifunctional VHH antibody.

• Korean Journal of Organic Agriculture
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• v.26 no.4
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• pp.661-676
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• 2018

Veronica persica (V. persica) is a perennial plant that is broadly distributed in Europe, Asia and so on. V. persica is used for pain about the lower abdomen and low back. The aim of this study was to investigate the anti-oxidant and anti-inflammatory effects of V. persica ethanol extract in LPS-induced RAW 264.7 cells. To evaluate the anti-oxidant activity, the DPPH and ABTS radical scavenging, total polyphenol and flavonoid contents, and reducing power activity were carried out. The DPPH and ABTS radical scavenging activity were evaluated as 72.0% and 73.0% at the concentrations of 200 and $500{\mu}g/mL$, respectively. Total polyphenol and flavonoid contents of V. persica extracts were measured as 65.22 mg/g and 43.82 mg/g at the concentration of 1 mg/mL. The reducing power activity measurement showed 53.0% activity at 1 mg/mL. The anti-inflammatory effects of the V. persica extract were evaluated in LPS induced RAW 264.7 cells. In the evaluation of cell viability by proliferation ＆ cytotoxicity assay kit, the cytotoxicity of the extract was not confirmed at $0{\sim}800{\mu}g/mL$ concentration. And the V. persica significantly inhibited NO production in a concentration dependent manner. The inhibition effects of NO in cell medium of V. persica was over 80% at $800{\mu}g/mL$. The V. persica also suppressed the expression of iNOS, COX-2, and phosphorylation of $NF-{\kappa}B$ and $IkB-{\alpha}$ proteins. These results indicate that the V. persica has anti-oxidant and anti-inflammatory effects by modulating $NF-{\kappa}B$ signaling pathways and can be used as natural functional materials.

• Korean Journal of Clinical Laboratory Science
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• v.51 no.1
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• pp.71-77
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• 2019

Menadione is known as an anti-tumor factor. Many studies have reported the potential anti-cancer role of menadione against a range of cancer cell lines. In this study, the anti-cancer effects of menadione and the underlying molecular signaling involved in apoptosis was investigated in gastric cancer cell lines. The menadione treatment decreased the cell viability of MKN45 gastric cancer cells. The decreased cell viability was attributed to the induction of apoptosis, which was confirmed by the results indicating the activation of caspase-3 and -7 and the cleavage of PARP in Western blotting. The upstream regulatory molecules involved in apoptosis were investigated further and it was discovered that menadione reduced the expression of survivin, an inhibitor of upstream apoptosis proteins. In addition, a transcription factor ${\beta}$-catenin, which is known to regulate survivin expression, was down-regulated by menadione. A previous report showed that menadione inhibited XIAP expression to induce apoptosis and induced G2/M cell cycle arrest in AGS cells. This study elucidated another inhibitory mechanism of menadione against gastric cancer cells in a different cell line. Although further studies will be needed, the inhibitory mechanism demonstrated in this study will help better understand the anti-cancer effects of menadione.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.63 no.4
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• pp.331-337
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• 2018

The content of nutrients, proteins, and oils of crop seeds is affected by global climate change due to the increase in temperature. Information regarding the effects of increased temperature on soybean seed nutrition is limited despite its vital role in seed quality and food security. The objective of this study was to determine the effect of increasing temperature on seed nutrient, protein, and oil content in two soybean [Glycine max (L.) Merr] cultivars (Daewonkong and Pungsannamulkong during the reproductive period in a temperature-gradient chamber. Four temperature treatments, Ta (near ambient temperature), $Ta+1^{\circ}C$ (ambient temperature+$1^{\circ}C$), $Ta+2^{\circ}C$ (ambient temperature+$2^{\circ}C$), $Ta+3^{\circ}C$ (ambient temperature+$3^{\circ}C$), and $Ta+4^{\circ}C$ (ambient temperature+$4^{\circ}C$), were established by dividing the rows along the temperature gradient. At maturity, increased temperature did not significantly affect the concentration of P, K, Ca, and Mg. The protein and oil content was significantly correlated with temperature. At maturity, the protein content of DWK and PSNK was reduced at $Ta+4^{\circ}C$. The oil content was the highest at $Ta+4^{\circ}C$ in DWK, whereas it decreased in PSNK at $Ta+4^{\circ}C$. Consequently, the biochemical composition of soybean seeds changed with the increase in temperature. These results illustrate the effects of temperature on soybean seed nutrient, protein, and oil content, which can help improve soybean quality at different temperatures. Thus, the biochemical composition of crop seeds can be changed in accordance with nutritional requirements for the benefit of human health in the future.

• Korean Journal of Food Science and Technology
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• v.51 no.4
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• pp.379-392
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• 2019

The current study investigated the effect of Gabjubaekmok (Diospyros kaki) ethanolic extract (GEE) on $H_2O_2$-induced human neuroblastoma MC-IXC cells and amyloid beta $(A{\beta})_{1-42}$-induced ICR (Institute of Cancer Research) mice. GEE showed significant antioxidant activity that was evaluated based on ABTS, DPPH scavenging activity, and inhibition of malondialdehyde (MDA) and acetylcholinesterase activity. Further, GEE inhibited ROS production and increased cell viability in $H_2O_2$-induced MC-IXC cells. Administration of GEE ameliorated the cognitive dysfunction on $A{\beta}$-induced ICR mice as evaluated using Y-maze, passive avoidance, and Morris water maze tests. Results of ex vivo test using brain tissues showed that, GEE protected the cholinergic system and mitochondrial functions by increasing the levels of antioxidants such as ROS, mitochondrial membrane potential (MMP), and adenosine triphosphate (ATP) against $A{\beta}$-induced cognitive dysfunction. Moreover, GEE decreasd the expression levels of apoptosis-related proteins such as $TNF-{\alpha}$, p-JNK, p-tau, BAX and caspase 3. While, expression levels of p-Akt and $p-GSK3{\beta}$ increased than $A{\beta}$ group. Finally, gallic acid was identified as the main compound of GEE using high performance liquid chromatography.

• 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.29 no.10
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• pp.1104-1110
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• 2019

Recently, there has been an increase in the elderly population of the world. Consequently, bone metabolic diseases such as osteoporosis are emerging as a social problem. Osteoclasts play a role in bone resorption, and osteoporosis is induced when bone resorption occurs excessively. Because currently used bone resorption inhibitors may cause side effects when used for a long period of time, it is necessary to develop a new material that effectively inhibits osteoclast differentiation. This study aimed to confirm the inhibitory effect of ethanol extract of Locusta migratoria on RANKL-induced osteoclast differentiation and its mechanism. The toxicity and proliferation effects of LME on RAW264.7 osteoclasts were measured by an MTS assay. There was no cytotoxicity or proliferation when the osteoclasts were treated with up to $2,000{\mu}g/ml$ of LME. In order to confirm the effect of LME on the differentiation of osteoclasts, osteoclasts were treated with RANKL alone or with LME for 3 days. As a result of a TRAP (tartrate-resistant acid phosphatase) assay, the increasing osteoclast differentiation by RANKL decreased in a concentration-dependent manner with the treatment of LME. In addition, LME suppressed the expression of differentiation-related marker genes (TRAP, RANK, NFATc1, and CK) and proteins (NFATc1 and c-Src) that had been increased by RANKL. Also, LME influenced the $NF-{\kappa}B$, ERK and JNK signaling pathways, resulting in the inhibition of osteoclast differentiation. These results suggest that LME may be used as a novel functional material for the prevention and treatment of osteoporosis by playing a role in inhibiting bone absorption.

• Journal of Life Science
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• v.29 no.8
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• pp.904-915
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• 2019

Prostate cancer (PCa) is one of the most metastatic tumor. Although hormone therapy or surgical castration is mostly conducted to treat PCa, it has a lot of side effects. Recently, many researchers have been exploring the tumor microenvironment to remedy these circumstances. Immune cells, especially macrophages, are an important composition of the tumor microenvironment. Under normal conditions, macrophages exhibit mild tumoricidal activity against tumors. However, once activated by interferon gamma or lipopolysaccharides, macrophages can kill cancer cells directly or indirectly by secreting cytokines and chemokines. In this study, murine macrophage RAW 264.7 cells were treated with Phellinus linteus extract. To analyze their pro-inflammatory phenotype, we were used several assays such as a real-time polymerase chain reaction, an enzyme-linked immunosorbent and nitric oxide assay. Prostate cancer cells were treated with the RAW 264.7-conditioned media, which was identified as a pro-inflammatory nature, for 48 h, and the expression of epithelial-mesenchymal transition (EMT)-related genes was determined. Not only N-cadherin, Snail, Twist, Slug, and Cadherin 11, which are mechenchymal-related proteins, were decrease, but epithelial marker of E-cadherin was increased. In addition, the mRNA level of vimentin, ccl2, and vegfa were decreased, as the EMT is closely related to the migration and invasion of cancer cells. In conclusion, the RAW 264.7-conditioned media stimulated with P. linteus extract inhibited migration and invasion and regulated the EMT pathway in human prostate cancer cells.