• Title/Summary/Keyword: Flow Identification

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Identification of Histone Deacetylase 1 Protein Complexes in Liver Cancer Cells

  • Farooq, Muhammad;Hozzein, Wael N.;Elsayed, Elsayed A.;Taha, Nael A.;Wadaan, Mohammad A.M.
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.2
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    • pp.915-921
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    • 2013
  • Background: Hepatocellular carcinoma is one of the leading causes of mortalities worldwide. The search for new therapeutic targets is of utmost importance for improved treatment. Altered expression of HDAC1 in hepatocellular carcinoma (HCC) and its requirement for liver formation in zebrafish, suggest that it may regulate key events in liver carcinogenesis and organogenesis. However, molecular mechanisms of HDAC1 action in liver carcinogenesis are largely unknown. The present study was conducted to identify HDAC1 interacting proteins in HepG2 cells using modified SH-double-affinity purification coupled with liquid mass spectrophotemetery. Materials and Methods: HepG2 cells were transfected with a construct containing HDAC1 with a C-terminal strepIII-HA tag as bait. Bait proteins were confirmed to be expressed in HepG2 cells by western blotting and purified by double affinity columns and protein complexes for analysis on a Thermo LTQ Orbitrap XL using a C18 nano flow ESI liquid chromatography system. Results: There were 27 proteins which showed novel interactions with HDAC1 identified only in this study, while 14 were among the established interactors. Various subunits of T complex proteins (TCP1) and prefoldin proteins (PFDN) were identified as interacting partners that showed high affinity with HDAC1 in HepG2 cells. Conclusions: The double affinity purification method adopted in this study was very successful in terms of specificity and reproducibility. The novel HDAC1 complex identified in this study could be better therapeutic target for treatment of hepatocellular carcinoma.

Generation of Anti-HLA-DR4 Specific Antibodies by Immunization of the Recombinantly Expressed Allelic Subtype-Specific Region of the $HLA-DRB1^*0405$ Molecules

  • Park, Jung-Hyun;Cho, Eun-Wie;Lee, Yun-Jung;Chung, Jin;Hahm, Kyung-Soo;Kim, Kil-Lyong
    • BMB Reports
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    • v.31 no.2
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    • pp.111-116
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    • 1998
  • HLA-DR4 is the dominant allele of MHC class II genes in Koreans. In particular, the $DRB1^*0405$ subtype has been reported to be almost exclusively expressed in Far East Asians, and has also been observed to be strongly associated with rheumatoid arthritis in Koreans and the Japanese. Identification of this specific allele has been mainly performed by PCR-based methods, which is often time consuming, costly, and involves tedious procedures such as the isolation of genomic DNA, PCR, and gel electrophoresis. To develop a more convenient tool for screening vast amounts of samples as well as to generate reagents which might also be used in other applications, in this study, antibodies were produced against this specific HLA subtype. By PCR, an allelespecific region covering the ${\beta}1$ domain of $DRB1^*0405$ was amplified and recombinantly expressed in E.coli. Immunization of Lewis rats with the purified protein yielded an allele specific antiserum. Western blot analysis showed the selective detection of the HLA-DR ${\beta}-chain$. Using this antiserum, established cell lines and peripheral blood lymphocytes were analyzed on their HLA haplotype by fluorescence activated flow cytometry. These novel antibodies will provide a powerful tool in the detection and investigation of DR4 alleles.

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Extraction and Identification of Volatile Isothiocyanates from Wasabi using Supercritical Carbon Dioxide (초임계 이산화탄소를 이용한 고추냉이로부터 휘발성 Isothiocyanates류 추출 및 동정)

  • Kim, Sung-Jin;Lee, Min-Kyung;Back, Sung-Sin;Chun, Byung-Soo
    • KSBB Journal
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    • v.22 no.3
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    • pp.174-178
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    • 2007
  • The aim of this study was to identifyisothiocyanates (ITCs) from wasabi (Wasabi japonica Matsum) using supercritical carbon dioxide ($SCO_2$) and to compare the composition in the extracts between $SCO_2$ and organic solvents extraction. A semi-continuous high pressure apparatus was used to extract wasabi (roots, stems and leaves) at following conditions pressure 80$\sim$120 bar, temperature $40\sim50^{\circ}C$. Ether, ethanol, chloroform and dichloromethane were used as organic solvents. The ITCs extracted by means of both separation technologies were analyzed by a gas chromatography system. As the results of study, AITC and ITCs were highly extracted at 40$^{\circ}C$ and 80 bar. To extract AITC from wasabi, $SCO_2$ extraction is more effective than organic solvents extraction, resulted in thermal degeneration and remaining of organic solvents.

Control Oriented Storage and Reduction Modeling of the Lean NOx Trap Catalyst (제어를 위한 Lean NOx Trap의 흡장 및 환원 모델링)

  • Lee, Byoungsoo;Han, Manbae
    • Transactions of the Korean Society of Automotive Engineers
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    • v.22 no.2
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    • pp.60-66
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    • 2014
  • A control oriented model of the Lean $NO_x$ trap (LNT) was developed to determine the timing of $NO_x$ regeneration. The LNT model consists of $NO_x$ storage and reduction model. Once $NO_x$ is stored ($NO_x$ storage model), at the right timing $NO_x$ should be released and then reduced ($NO_x$ reduction model) with reductants on the catalyst active sites, called regeneration. The $NO_x$ storage model simulates the degree of stored $NO_x$ in the LNT. It is structured by an instantaneous $NO_x$ storage efficiency and the $NO_x$ storage capacity model. The $NO_x$ storge capacity model was modeled to have a Gaussian distribution with a function of exhaust gas temperature. $NO_x$ release and reduction reactions for the $NO_x$ reduction model were modeled as Arrhenius equations. The parameter identification was optimally performed by the data of the bench flow reactor test results at space velocity 50,000/hr, 80,000/hr, and temperature of $250-500^{\circ}C$. The LNT model state, storage fraction indicates the degree of stored $NO_x$ in the LNT and thus, the timing of the regeneration can be determined based on it. For practical purpose, this model will be verified more completely by engine test data which simulate the NEDC transient mode.

Influence of rutin on the effects of neonatal cigarette smoke exposure-induced exacerbated MMP-9 expression, Th17 cytokines and NF-κB/iNOS-mediated inflammatory responses in asthmatic mice model

  • Liu, Li-Li;Zhang, Yan;Zhang, Xiao-Fang;Li, Fu-Hai
    • The Korean Journal of Physiology and Pharmacology
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    • v.22 no.5
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    • pp.481-491
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    • 2018
  • Allergic asthma is one of the most enduring diseases of the airway. The T-helper cells and regulatory T-cells are critically involved in inflammatory responses, mucus hypersecretion, airway remodelling and in airway hyper-responsiveness. Cigarette smoke (CS) has been found to aggravate inflammatory responses in asthma. Though currently employed drugs are effective, associated side effects demand identification and development of novel drugs with negligible or no adverse effects. Rutin, plant-derived flavonoid has been found to possess antioxidant and anti-inflammatory effects. We investigated the ability of rutin to modulate T-cells and inhibit inflammation in experimentally-induced asthma in cigarette smoke exposed mice. Separate groups of neonatal mice were exposed to CS for 10 days from post-natal days 2 to 11. After 2 weeks, the mice were sensitized and challenged with ovalbumin (OVA). Treatment group were given rutin (37.5 or 75 mg/kg body weight) during OVA sensitization and challenge. Rutin treatment was found to significantly inhibit cellular infiltration in the airways and Th2 and Th17 cytokine levels as well. Flow cytometry revealed effectively raised $CD4^+CD25^+Fox3^+$ Treg cells and supressed Th17 cell population on rutin treatment. Airway hyper-responsiveness observed following CS and OVA challenge were inhibited by rutin. $NF-{\kappa}B$ and iNOS, chief regulators of inflammatory responses robustly activated by CS and OVA were down-regulated by rutin. Rutin also inhibited the expression of matrix metalloproteinase 9, thereby aiding in prevention of airway remodelling in asthma thereby revealing to be a potent candidate in asthma therapy.

Simultaneous Determination of Six Components in the Traditional Herbal Medicine 'Oryeongsan' by HPLC-DAD and LC-MS/MS

  • Lee, Jiwoo;Weon, Jin Bae;Lee, Bohyoung;Yun, Bo-Ra;Eom, Min Rye;Ma, Choong Je
    • Natural Product Sciences
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    • v.19 no.1
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    • pp.20-27
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    • 2013
  • A simple high performance liquid chromatography - diode array detector (HPLC-DAD) method has been developed and validated for simultaneous determination of the six components (cinnamic acid, cinnamaldehyde, eugenol, atractylenolide I, atractylenolide III, and ergosterol) in Oryeongsan. In addition, identification of six marker compounds was conducted by a LC-MS/MS analysis. The six compounds in Oryeongsan were separated on Shishedo $C_{18}$ column (5 ${\mu}m$, $4.6{\times}250$ mm) at a column temperature of $30^{\circ}C$. The mobile phase was a mixture of 0.1% trifluoroacetic acid (TFA) water and acetonitrile employing gradient elution at a flow rate of 1.0 mL/min. The detection wavelength was set at 205, 250, 280, and 330 nm. The developed method had good linearity ($R^2$ > 0.9997) and the limit of detection (LOD) and limit of quantification (LOQ) were observed within the ranges 0.01~0.15 and 0.05~0.45 ${\mu}g/mL$, respectively. The relative standard deviation (RSD) values of intra- and inter-day testing were indicated that less than 3% and 90.31~103.31% of accuracy. The results of recovery test were 90.56~106.72% and RSD range was measured from 0.84 to 2.95%. In conclusion, this HPLC-DAD method has been successfully applied to the simultaneous determination of six compounds in Oryeongsan samples.

Applications and Prospects of Stable Isotope in Aquatic Ecology and Environmental Study (수생태 환경 연구에 있어 안정동위원소의 활용과 전망)

  • Choi, Bohyung;Shin, Kyung-Hoon
    • Korean Journal of Ecology and Environment
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    • v.51 no.1
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    • pp.96-104
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    • 2018
  • Stable isotope approach for aquatic ecology and environmental sciences has been introduced as very useful technique since 1980s and also has been applied to investigate various issues in aquatic ecology and environmental study last 10 years in Korea. Especially carbon and nitrogen isotope ratios have been mainly used to understand food web energy flow and ecosystem structure. In addition, nitrogen isotope ratio has been applied for nitrogen cycle and source identification as well as biomagnification studies. However, large temporal or spatial variations of nitrogen isotope ratio of primary producer have been found in many aquatic environments, and it is regarded as the critical problems to determine trophic level of aquatic animals. Recently, the compound specific isotope analysis of nitrogen within individual amino acids has been developed as an alternative method for trophic ecology. This article introduces the progress history of stable isotope application in aquatic ecology and environmental sciences, and also suggests new direction based on future prospects in stable isotope ecology and environmental study.

Morphological and Cytogenetic Analysis of Colchicine-induced Tetraploids of Fallopia multiflolra Haraldson (Colchicine 처리에 의해 유기된 4배체 하수오의 형태 및 세포유전학적 특성)

  • Kim, Ki Hyun;Youn, Cheol Ku;Kim, In Jae;Lee, Kyung Ja;Kim, Young Ho;Hong, Seong Tack;Woo, Sun Hee
    • Korean Journal of Medicinal Crop Science
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    • v.26 no.5
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    • pp.362-369
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    • 2018
  • Background: For stable induction of tetraploidy in Fallopia multiflora Haraldson, colchicine was treated to establish the condition of induction and investigated the morphological and cytogenetic traits of the tetraploid plants obtained compared to those of diploid ones. Methods and Results: For the induction of tetraploidy, F. multiflora plants were soaked in aqueous solutions of colchicine at various concentration (0.1, 0.5, and 1.0%). After this, 2% dimethyl sulfoxide (DMSO) was added at room temperature on a shaker set at 150 rpm for periods of 12, 24, and 48 h. The induction rate of tetraploids appeared to be the highest in plants treated with 0.5% colchicine for 24 h. As the colchicine concentration and soaking time increased above these levels, the growing tip of the roots did not develop and they began to rot. When compared to diploid plants, tetraploids differed greatly in various characteristics, including the sizes and shapes of the leaves, fruits, flowers and roots. The induced tetraploid F. multiflora had larger guard cells, and chloroplasts, increased number of chloroplast in the guard cells and decreased stomatal densities. Conclusions: When colchicine induced plants for tetraploid, it can be distinguished from diploids, in various characteristics such as morphological changes as stomatal size, number of chloroplasts per guard cell, number of chromosomes and flow cytometry. Therefore, it proved that these methods are suitable, quick and easy methods for the identification of the ploidy level of F. multiflora.

Estimation of shelf-life by long-term storage test of Pyungwi-san (평위산 전탕팩의 장기보존 시험에 따른 유통기한 설정)

  • Seo, Chang-Seob;Kim, Jung-Hoon;Lim, Soon-Hee;Shin, Hyeun-Kyoo
    • Herbal Formula Science
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    • v.19 no.1
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    • pp.183-194
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    • 2011
  • Objectives : To estimate the shelf-life by long-term storage test of Pyungwi-san. Methods : Experiments were conducted to evaluate the stability such as the selected physicochemical, heavy metal, microbilogical experiment under an acceleration test and long-term storage test of Pyungwi-san in different storage under room temperature, refrigeration and freezing. Futhermore, HPLC analysis was performed for the determinations of glycyrrhizin in the Pyungwi-san on an Inertsil ODS-3 column(250 mm ${\times}$ 4.6 mm, 5 um) using solvent 35% acetonitrile include 0.05% phosphoric acid at 254 nm. The flow rate was 1.0 mL/min. Results : The significant change was not showed in pH, heavy metal, microbiological, identification test and quantitative analysis based on acceleration test and long-term storage test. Retention time of glycyrrhizin in HPLC chromatogram was about 16.065 min and calibration curve showed good linearity($R^2$ = 0.9999). The contents of glycyrrhizin in acceleration test and long-term storage test were 0.068~0.076 mg/mL and 0.066~0.077 mg/mL, respectively. Shelf-lifes of room temperature, refrigeration and freezing by long-term storage test were predicted 41, 24 and 34 months, respectively. Conclusions : The suggested shelf-life would be helpful on the storage and distribution of herbal medicine.

fabrication of DMMP Thick Film Gas Sensor Based on SnO2 (산화주석을 기반으로 한 DMMP 후막가스센서 제작)

  • 최낙진;반태현;곽준혁;백원우;김재창;허증수;이덕동
    • Journal of the Korean Institute of Electrical and Electronic Material Engineers
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    • v.16 no.12S
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    • pp.1217-1223
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    • 2003
  • Nerve gas sensor based on tin oxide was fabricated and its characteristics were examined. Target gas is dimethyl methyl phosphonate(C$_3$ $H_{9}$ $O_3$P, DMMP) that is simulant gas of nerve gas. Sensing materials were Sn $O_2$ added a-Al$_2$ $O_3$ with 0∼20wt.% and were physically mixed each material. They were deposited by screen printing method on alumina substrate. The sensor device was consisted of sensing electrode with interdigit(IDT) type in front and a heater in back side. Total size of device was 7${\times}$10${\times}$0.6㎣. Crystallite size & phase identification and morphology of fabricated Sn $O_2$ powders were analyzed by X-ray diffraction and by a scanning electron microscope, respectively. Fabricated sensor was measured as flow type and resistance change of sensing material was monitored as real time using LabVIEW program. The best sensitivity was 75% at adding 4wt.% $\alpha$-Al$_2$ $O_3$, operating temperature 30$0^{\circ}C$ to DMMP 0.5ppm. Response and recovery time were about 1 and 3min., respectively. Repetition measurement was very good with $\pm$3% in full scale.TEX>$\pm$3% in full scale.