• Title/Summary/Keyword: light-chain

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Intergeneric Transfer of Isolated Nuclei from Lentinus edodes into Protoplasts of Pleurotus florida (사철느타리버섯 원형질체내에 표고 핵의 전이)

  • Yoo, Young-Bok;Shin, Pyung-Gyun
    • The Korean Journal of Mycology
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    • v.24 no.2 s.77
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    • pp.111-126
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    • 1996
  • Transfer of the isolated nuclei from Lentinus edodes into protoplasts of Pleurotus florida was induced with polyethlene glycol (PEG) and $CaCl_2$. The intergeneric transfer products were classified into nuclear hybrid, heterokaryon or synkaryon, and reconstituted cell. These progenies except nuclear hybrids formed mature fruiting bodies on sawdust rice bran medium. Formation of fruit bodies was influenced by several factors such as light, temperature, nutrition and physic state of the culture media. Most of fruiting body characters were similar to those of P. florida in synkaryon and L. edodes in reconstituted cell, respectively. All these basidiocarps had clamp connections though initial heterokaryon colonies were lacking. Isozyme patterns of intergeneric progenies were quite different from those of parents. DNA polymorphisms of transfer products were also compared by random amplified polymorphic DNAs (RAPD) analysis based on polymerase chain reaction. The RAPD patterns were different from those of donor and recipient. DNA fingerprints ranged in size from 0.25 to 4.0 Kb. On the basis of RAPD, the transfer products were classified into five groups. Two synkaryon were analysed with distribution of progenies and segregation of genetic markers by random spore analyses. The genetic markers were segregated into wild type and riboflavine requiring auxotrophs.

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[6]-Gingerol Attenuates Autophagy and Increases Activities of Antioxidative Defense Enzymes in Mice with Cerulein-induced Acute Pancreatitis (Cerulein 유도 급성췌장염 마우스모델에서 자가분해 조절과 항산화 활성에 미치는 [6]-gingerol의 영향)

  • Kim, Sung Ok;Choi, Yung Hyun
    • Journal of Life Science
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    • v.23 no.10
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    • pp.1280-1287
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    • 2013
  • The current study investigated the effects of [6]-gingerol, a ginger phytochemical, on the expression of autophagy-related genes and the activation of antioxidative enzymes in the pancreas of mice with cerulein-induced acute pancreatitis. The following were studied: pancreatic edema, ${\alpha}$-amylase activity in serum, expression of autophagy genes, activities of antioxidative defense enzymes, such as superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), and the production of lipid peroxidation (LPO). The results revealed that cerulein-induced edema in the pancreas and ${\alpha}$-amylase activity in the cerulein group significantly increased compared with that of the control. However, that of the [6]-gingerol pretreated group was significantly decreased compared with that of the cerulein-alone injected group (positive control). There was no significant difference compared with that of control. The expression of autophagy-related proteins, including Beclin-1 and cleaved microtubule-associated protein 1 light chain 3, were significantly increased in the positive control but significantly decreased in the [6]-gingerol-pretreated group. Furthermore, the activities of SOD and GSH-Px in the positive control were decreased compared with those of the control. However, those of the [6]-gingerol pretreated group were significantly increased compared with those of the cerulein-alone group. The mRNA levels and antioxidant enzyme activities were similar. The production of LPO in the cerulein with and without [6]-gingerol groups was increased by 133.1% and 26.3%, respectively, compared with that of the control, whereas that of the [6]-gingerol-pretreated group was significantly decreased by 48.5% compared with that of the positive control. Therefore, [6]-gingerol may be a strong candidate in reducing autophagy and LPO production and in enhancing antioxidative enzyme activities to help prevent acute and chronic pancreatitis.

Role of NFAT5 in Osteogenic Differentiation of Human Adipose Tissue-Derived Mesenchymal Stem Cells (인체 지방 유래 중간엽 줄기세포의 골분화 조절 기전에서 NFAT5의 역할)

  • Lee, Sun Young;Yang, Ji won;Jung, Jin Sup
    • Journal of Life Science
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    • v.23 no.4
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    • pp.471-478
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    • 2013
  • Human adipose tissue-derived mesenchymal stem cells (hADSCs) have therapeutic potential, including the ability to self-renew and differentiate into multiple lineages. Understanding of molecular mechanisms of stem cell differentiation is important for improving the therapeutic efficacies of stem cell transplantation. In this study, we determined the role of nuclear factor of activated T cells (NFAT5) in the osteogenic differentiation of hADSCs. The down-regulation of NFAT5 expression by the transfection of a specific siRNA significantly inhibited osteogenic differentiation of hADSCs and decreased the activity of the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-${\kappa}B$) promoter without affecting their proliferation and adipogenic differentiation. The inhibition of NFAT5 expression inhibited the basal and Tumor Necrosis Factor ${\alpha}$ (TNF-${\alpha}$) induced activation of NF-${\kappa}B$, but it did not affect TNF-${\alpha}$-induced degradation of the $I{\kappa}B$ protein. These findings indicate that NFAT5 plays an important role in the osteogenic differentiation of hADSCs through the modulation of the NF-${\kappa}B$ pathway.

Substitution of Heavy Complementarity Determining Region 3 (CDR-H3) Residues Can Synergistically Enhance Functional Activity of Antibody and Its Binding Affinity to HER2 Antigen

  • Moon, Seung Kee;Park, So Ra;Park, Ami;Oh, Hyun Mi;Shin, Hyun Jung;Jeon, Eun Ju;Kim, Seiwhan;Park, Hyun June;Yeon, Young Joo;Yoo, Young Je
    • Molecules and Cells
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    • v.39 no.3
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    • pp.217-228
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    • 2016
  • To generate a biobetter that has improved therapeutic activity, we constructed scFv libraries via random mutagenesis of several residues of CDR-H3 and -L3 of hu4D5. The scFv clones were isolated from the phage display libraries by stringent panning, and their antiproliferative activity against HER2-positive cancer cells was evaluated as a primary selection criterion. Consequently, we selected AH06 as a biobetter antibody that had a 7.2-fold increase in anti-proliferative activity ($IC_{50}$: 0.81 nM) against the gastric cancer cell line NCI-N87 and a 7.4-fold increase in binding affinity ($K_D$: 60 pM) to HER2 compared to hu4D5. The binding energy calculation and molecular modeling suggest that the substitution of residues of CDR-H3 to W98, F100c, A101 and L102 could stabilize binding of the antibody to HER2 and there could be direct hydrophobic interactions between the aromatic ring of W98 and the aliphatic group of I613 within HER2 domain IV as well as the heavy and light chain hydrophobic interactions by residues F100c, A101 and L102 of CDR-H3. Therefore, we speculate that two such interactions were exerted by the residues W98 and F100c. A101 and L102 may have a synergistic effect on the increase in the binding affinity to HER2. AH06 specifically binds to domain IV of HER2, and it decreased the phosphorylation level of HER2 and AKT. Above all, it highly increased the overall level of p27 compared to hu4D5 in the gastric cancer cell line NCIN82, suggesting that AH06 could potentially be a more efficient therapeutic agent than hu4D5.

Ginseng improves cognitive deficit via the RAGE/NF-κB pathway in advanced glycation end product-induced rats

  • Tan, Xiaobin;Gu, Junfei;Zhao, Bingjie;Wang, Shuyuan;Yuan, Jiarui;Wang, Chunfei;Chen, Juan;Liu, Jiping;Feng, Liang;Jia, Xiaobin
    • Journal of Ginseng Research
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    • v.39 no.2
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    • pp.116-124
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    • 2015
  • Background: Ginseng, the root of Panax ginseng (PG), is used widely as a herbal medicine to prevent and treat various diseases. Panax ginseng has pharmacological effects on neurodegenerative diseases such as Alzheimer's disease (AD). The present study evaluated the neuroprotective effects of PG and its possible neuroprotective mechanisms in advanced glycation end product (AGE)-induced AD in a rat model. Methods: Advanced glycation end products were injected bilaterally into the CA3 region of the rats' brains. The Morris water maze test and step-down type passive avoidance test were performed to evaluate their memory and cognitive abilities. The oxidation indexes in the hippocampus were detected. Immunohistochemistry was conducted to visualize the receptors for advanced glycation end products (RAGEs) and nuclear factor-kappa-light-chain-enhancer of activated B cell (NF-${\kappa}B$). Results: Behavioral results showed that PG (1 g/kg, 0.5 g/kg, and 0.25 g/kg) significantly shortened the escape latency, remarkably increased the number of crossing times, significantly decreased the number of errors, and prolonged the latency in rats with AGE-induced AD. Panax ginseng also significantly reduced the malondialdehyde level, increased the glutathione content, and increased superoxide dismutase activity in the hippocampus. Panax ginseng significantly decreased the expression of RAGE and NF-${\kappa}B$. The blockade of anti-RAGE antibody could significantly reduce AGE-induced impairments and regulate these expressions. Conclusion: Our results demonstrated that PG significantly inhibits AGE-induced memory impairment and attenuates Alzheimer-like pathophysiological changes. These neuroprotective effects of PG may be associated with the RAGE/NF-${\kappa}B$ pathway. Our results provided the experimental basis for applying PG in preventing and treating AD.

A Comparative Study on the Characteristics of Accelerated aging at Low and High Temperatures of the Fluorocarbon Rubber Composites (불소 고무복합체의 저온과 고온촉진노화 특성에 대한 비교 연구)

  • Park, JeongBae;Lee, BeomCheol;Jeong, YoonSeok;Park, SungHan
    • Proceedings of the Korean Society of Propulsion Engineers Conference
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    • 2017.05a
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    • pp.915-922
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    • 2017
  • The study on the thermal and oil resistance rubber composite, 2016. [6] predicted the lifetime of Fluorocarbon Rubber by accelerating aging at high temperature ($150^{\circ}C$, $175^{\circ}C$, $200^{\circ}C$). general rubber products are likely to exhibit different properties depending on the degradation factors such as temperature, humidity, ozone, light, emulsion, mechanical and electrical stress. To solve these problems, We compared the rate of change about tensile strength, elongation rate, volume change rate, weight change rate, thickness change rate, thermal conductivity in low temperature promoting aging on the basis of predictive lifetime of high temperature promoting aging. As a result of the review, the required life expectancy was satisfied, but there was a slight difference in the rate of change between the high-temperature promoted aging life result and the low temperature promoted aging life result. The cause was a reduction in "tensile strength / elongation" and an increase in "volume / weight / thickness" caused by the main chain decomposition of fluorine rubber due to aging at high temperature promoting aging. However, the low temperature promoting aging was caused by the curing reaction of fluorine rubber at $80^{\circ}C$. The tensile strength / elongation and volume / weight / thickness changes were small.

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Mode of Action of the New Diphenyl Ether Herbicide KC6361 (새로운 백화형 디페닐에테르계 화합물 KC-6361의 제초작용기작)

  • Kim, J.S.;Kim, T.J.;Kim, Y.S.;Cho, K.Y.
    • Korean Journal of Weed Science
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    • v.14 no.2
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    • pp.81-93
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    • 1994
  • This study was conducted to characterize herbicidal activity of the new type diphenylether compounds inducing bleaching(whitening) in relation to their chemical structures and to find out the herbicidal action mechanism. Bleaching was highly appeared in the compounds which have short chain alkyl or allyl group of cabamoyl type in meta position of A ring and nitro group in para position of B ring, and KC6361 showed the highest efficacy among these derivatives. Rice was tolerant to KC6361 at 2.0kg/ha. The applications of this compound at 0.25~1.0kg/ha provided acceptable control of the various weeds including large crabgrass, barnyardgrass, pigweed and so on. In the dark condition, KC6361 decreased carotenoid contents but increased chlorophyll in etiolated cucumber cotyledon. By the increase of light intensity, the carotenoid biosynthesis was more inhibited than the chlorophyll biosynthesis. The incorporation of $C^{14}$-acetate into lipid was slightly inhibited. Both KC6361 and norflurazon stimulated growth of dwarf rice(cv. Daneunbanju). These results suggest that both KC6361 and norflurazon may have the same action sites. To confirm this hypothesis, the changes of carotenoid intermediates in cucumber cotyledon and barnyardgrass leaf treated with KC6361 were investigated. Phytoene and phytofluene were increased but ${\beta}$-carotene was decreased, indicating that KC6361 inhibited phytoene and/or phytofluene dehydrogenase like norflurazon.

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Immunomodulating Effect of Extract of Cheonggukjang Fermented with Bacillus amyloliquefaciens (SRCM100730) on RAW 264.7 Macrophages (Bacillus amyloliquefaciens(SRCM 100730)로 발효된 청국장 추출물의 RAW 264.7 대식세포 면역증강 활성)

  • Choo, Seung Bin;Yang, Hui;Jeong, Do-Yuon;Jeong, Seong-Yeop;Ryu, Myeong Seon;Oh, Kwang-Hoon;Yoo, Yung Choon
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.46 no.11
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    • pp.1300-1307
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    • 2017
  • Cheonggukjang is well known as a traditional fermented food in Korea and has various biological activity. In this study, immune-enhancing activity of extract of cheonggukjang fermented with Bacillus amyloliquefaciens (SRCM100730) was examined in RAW 264.7 murine macrophages. Treatment with extract augmented production of nitric oxide (NO) and tumor necrosis $factor-{\alpha}$ ($TNF-{\alpha}$) from RAW 264.7 macrophages in a dose-dependent manner. Similarly, increased mRNA expression of inducible nitric oxide synthase (iNOS) and $TNF-{\alpha}$ was observed. In addition, the extract synergistically enhanced production of NO and $TNF-{\alpha}$ from lipopolysaccharide (LPS)-stimulated macrophages. Analysis of intracellular pathways revealed that the immune-enhancing activity of cheonggukjang extract was related to activation of mitogen-activated protein kinases (MAPK) and nuclear factor kappa-light-chain-enhancer of activated B cells ($NF-{\kappa}B$). These results suggest that cheonggukjang fermented with B. amyloliquefaciens (SRCM100730) is a beneficial food effective for activation of immune responses.

Remifentanil Protects Human Keratinocyte Through Autophagic Expression

  • Kim, Eok Nyun;Park, Chang Hoon;Woo, Mi Na;Yoon, Ji Young;Park, Bong Soo;Kim, Yong Ho;Kim, Cheul Hong
    • Journal of The Korean Dental Society of Anesthesiology
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    • v.14 no.2
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    • pp.101-106
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    • 2014
  • Background: Remifentanil, an ultra-short-acting mu-opioid receptor agonist, is unique from other opioids because of its esterase-based metabolism, minimal accumulation, and very rapid onset and offset of clinical action. Remifentanil can prevent the inflammatory response and can suppress inducible nitric oxide synthase expression in a septic mouse model. However, the effects of remifentanil on human keratinocyte and autophagy have yet to be fully elucidated during hypoxia-reoxygenation. Here we investigated whether remifentanil confers protective effect against hypoxia-reoxygenation in human keratinocyte and, if so, whether autophagy mediates this effect. Methods: The human keratinocytes were cultured under 1% oxygen tension. The cells were gassed with 94% $N_2$, and 5% $CO_2$ and incubated for 24 h at $37^{\circ}C$. To determine whether the administration of affects human keratinocytes hypoxia-reoxygenation injury, cells were then exposed to various concentrations of remifentanil (0.01, 0.1, 0.5 and 1 ng/ml) for 2 h. After remifentanil treatment, to simulate reoxygenation and recovery, the cells were reoxygenated for 12 h at $37^{\circ}C$. Control group did not receive remifentanil treatment. Normoxia group did not receive hypoxia and remifentanil treatment for 36 h. 3-MA group was treated 3-methyladenine (3-MA) for 1h before remifentanil treatment. Cell viability was measured using a quantitative colorimetric assay with MTT, showing the mitochondrial activity of living cells. Cells were stained with fluorescence and analyzed with Western blot analysis to find out any relations with activation of autophagy. Results: Prominent accumulation of autophagic specific staining MDC was observed around the nuclei in RPT group HaCaT cells. Similarly, AO staining, red fluorescent spots appeared in RPT group HaCaT cells, while the Normoxia, control and 3-MA groups showed mainly green cytoplasmic fluorescence. We here examined activation of autophagy related protein under H/R-induced cells by Western blotting analysis. Atg5, Beclin-1, LC3-II (microtubule-associated protein 1 light chain 3 form II) and p62 was elevated in RPT group cells. But they were decreased when autophagy was suppressed by 3-MA (Fig. 5). Conclusions: Although the findings of this study are limited to an in vitro interpretation, we suggest that remifentanil may have a beneficial effect in the recovery of wound from hypoxia-reoxygenation injury.

Melatonin protects endothelial progenitor cells against AGE-induced apoptosis via autophagy flux stimulation and promotes wound healing in diabetic mice

  • Jin, Haiming;Zhang, Zengjie;Wang, Chengui;Tang, Qian;Wang, Jianle;Bai, Xueqin;Wang, Qingqing;Nisar, Majid;Tian, Naifeng;Wang, Quan;Mao, Cong;Zhang, Xiaolei;Wang, Xiangyang
    • Experimental and Molecular Medicine
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    • v.50 no.11
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    • pp.13.1-13.15
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    • 2018
  • Wound healing is delayed in diabetic patients. Increased apoptosis and endothelial progenitor cell (EPC) dysfunction are implicated in delayed diabetic wound healing. Melatonin, a major secretory product of the pineal gland, promotes diabetic wound healing; however, its mechanism of action remains unclear. Here, EPCs were isolated from the bone marrow of mice. Treatment of EPCs with melatonin alleviated advanced glycation end product (AGE)-induced apoptosis and cellular dysfunction. We further examined autophagy flux after melatonin treatment and found increased light chain 3 (LC3) and p62 protein levels in AGE-treated EPCs. However, lysosome-associated membrane protein 2 expression was decreased, indicating that autophagy flux was impaired in EPCs treated with AGEs. We then evaluated autophagy flux after melatonin treatment and found that melatonin increased the LC3 levels, but attenuated the accumulation of p62, suggesting a stimulatory effect of melatonin on autophagy flux. Blockage of autophagy flux by chloroquine partially abolished the protective effects of melatonin, indicating that autophagy flux is involved in the protective effects of melatonin. Furthermore, we found that the AMPK/mTOR signaling pathway is involved in autophagy flux stimulation by melatonin. An in vivo study also illustrated that melatonin treatment ameliorated impaired wound healing in a streptozotocin-induced diabetic wound healing model. Thus, our study shows that melatonin protects EPCs against apoptosis and dysfunction via autophagy flux stimulation and ameliorates impaired wound healing in vivo, providing insight into its mechanism of action in diabetic wound healing.