• Title/Summary/Keyword: Transient analysis

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Effects of Mutagenesis for Glycosylation Sites of Recombinant Human EPO During Production from Cultured CHO Cell

  • Lee, Hyun-Gi;Seong, Hwan-Hoo;Im, Seok-Ki;Chung, Hee-Kyoung;Lee, Poongyeon;Lee, Yeun-Kun;Min, Kwan-Sik;Chang, Won-Kyoung;Lee, Hoon-Taek
    • Proceedings of the Korean Society of Embryo Transfer Conference
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    • 2002.11a
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    • pp.97-97
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    • 2002
  • Human eryhropoietin (EPO) is acidic glycoprotein hormone that plays key role in hematopoiesis by facilitating differentiation of erythrocyte and formation of hemoglobin (Hb) and is used for the treatment of anemia. Human EPO is consist of 166 amino acids which is modified by three N-glycosylations (24, 38, 83) and single O-glycosylation (126). N-glycosylation is reported to be related to the cellular secretion and activity of EPO. In this study, we examined effects of mutagenesis in glycosylation site of recombinat hEPO for the cellular secretion during production from cultured CHO cell. We produced rhEpo which was cloned by PCR from human liver cDNA (TaKaRa) in cultured CHO cell. Using supernatant of the culture, ELISA assay and western analysis were performed. To estimate biological activity, 20IU of rhuEpo was subcutaneously injected into four ICR mice. After 8 days, HCT level was increased average 13 per cent, RBC was increased ca. 2${\times}$10$\^$6//${\mu}\ell$. In disease model Rat (anemia c-kit, WSRC-WS/WS), HCT was increased ca. 12%, RBC was increased ca. 1.6${\times}$10$\^$6//${\mu}\ell$. These results suggests that rhEpo we produced has biological activity. To remove glycosylation site by substituting 24, 38, 83, and 126th asparagine (or serine) with glutamic acid, overlapping -extension site-directed mutagenesis was performed. To add novel glycosylation sites, 69, 105th leucine was mutated to asparagine. Mutant EPO construct was transfected into CHO cell. Supernatant of the cell culture was analyzed using ELISA assay with monoclonal anti-EPO antibody (Medac, Germany). Since, several reports for mutagenesis of glycosylation sites showed case-by-case results, we examined both transient expression and stable expression. Addition of novel glycosylation sites resulted no secretion while deletion mutants had little effect except some double deletion mutants (24/83 and 38/83) and triple mutant. We suggest that not single but combination of glycosyl group affect secretion of EPO.

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Isolation and Molecular Characterization of a New CRT Binding Factor Gene from Capsella bursa-pastoris

  • Wang, Xinglong;Liu, Li;Liu, Sixiu;Sun, Xiaoqing;Deng, Zhongxiang;Pi, Yan;Sun, Xiaofen;Tang, Kexuan
    • BMB Reports
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    • v.37 no.5
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    • pp.538-545
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    • 2004
  • A new CRT binding factor (CBF) gene designated Cbcbf25 was cloned from Capsella bursa-pastoris, a wild grass, by the rapid amplification of cDNA ends (RACE). The full-length cDNA of Cbcbf25 was 898 bp with a 669 bp open reading frame (ORF) encoding a putative DRE/CRT (LTRE)-binding protein of 223 amino acids. The predicted CbCBF25 protein contained a potential nuclear localization signal (NLS) in its N-terminal region followed by an AP2 DNA-binding motif and a possible acidic activation domain in the C-terminal region. Bioinformatic analysis revealed that Cbcbf25 has a high level of similarity with other CBF genes like cbf1, cbf2, and cbf3 from Arabidopsis thaliana, and Bncbf5, Bncbf7, Bncbf16, and Bncbf17 from Brassica napus. A cold acclimation assay showed that Cbcbf25 was expressed immediately after cold triggering, but this expression was transient, suggesting that it concerns cold acclimation. Our study implies that Cbcbf25 is an analogue of other CBF genes and may participate in cold-response, by for example, controlling the expression of cold-regulated genes or increasing the freezing tolerance of plants.

Stability Analysis for a Dyke Subjected to Tidal Fluctuations (조위변동(潮位變動)을 받는 호안제(護岸堤)의 사면안정해석(斜面安定解析))

  • Kim, Sang Kyu
    • KSCE Journal of Civil and Environmental Engineering Research
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    • v.8 no.3
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    • pp.91-100
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    • 1988
  • Assuming that tidal level is constantly changed with an amplitude of 10 meters and a cycle of 12 hours, the slope stability for a typical dyke is analysed. The variation of pore water pressure within the dyke during the tidal change is obtained using a computer program, FLUMP, which is incorporated with saturated-unsaturated and transient flow. The results show that the variation of free water surface and distribution of pore water pressure within the dyke during the tidal fluctuations can be clearly predicted with the computer program. When a tide is lowered to the minimum level, the predicted pressure head is higher than the level of the free water surface in some parts of the dyke; that is, excess pore water pressure is generated in a zone affected by the tidal change. Also an unsaturated zone which shows negative pore water pressure is temporally created when a tide is lowered. The shear strength of the zone can be predicted based on the proposal suggested by Fredlund et al. It is emphasized that the excess pore water pressure generated during tidal fluctuations and strength parameters for the unsaturated zone should be considered in analyzing the slope stability of dykes. When those are considered, the critical slip surface seems to be located below the free water surface obtained when a tide is at the lowest.

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Analysis of Hydraulic Characteristics of Flood Plain Using Two-Dimensional Unsteady Model (2차원 부정류 모형을 이용한 둔치의 수리특성 분석)

  • Ku, Young Hun;Song, Chang Geun;Kim, Young Do;Seo, Il Wo
    • KSCE Journal of Civil and Environmental Engineering Research
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    • v.33 no.3
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    • pp.997-1005
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    • 2013
  • Since the cross-sectional shape of the Nakdong river is compound type, the water stage rises up to the top of the flood plane, as the flow discharge increases during the extreme rain storm in summer. The recent increase of rainfall intensity and flood frequency results in the immersions of parks and hydrophilic facilities located in the flood plain. Therefore it is necessary to analyze the hydraulic characteristics evolved by the extreme rain storm in the flood plain. The study reach ranging from the Gangjeong Goryeong Weir and the Dalseong Weir, where several hydraulic facilities are located along the channel, was selected and numerical simulations were conducted for 42 hours including the peak flood of the typhoon Sanba. The 2-D transient model, FaSTMECH was employed and the accuracy of the model was assessed by comparing the water level between the simulation results and the measured ones at a gauging station. It showed a high correlation with $R^2$ of 0.990, AME of 0.195, and RMSE of 0.252. In addition, the inundation time, the inundation depth, the inundation velocity, and the shear stress variation in the flood plain facilities were analyzed.

Optimal arrangement of multiple wind turbines on an offshore wind-wave floating platform for reducing wake effects and maximizing annual energy production (다수 풍력터빈의 후류영향 최소화 및 연간발전량 극대화를 위한 부유식 파력-해상풍력 플랫폼 최적배치)

  • Kim, Jong-Hwa;Jung, Ji-Hyun;Kim, Bum-Suk
    • Journal of Advanced Marine Engineering and Technology
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    • v.41 no.3
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    • pp.209-215
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    • 2017
  • A large floating offshore wind-wave hybrid power generation system with an area of 150 m2 and four 3 MW class wind turbine generators was installed at each column top. In accordance with the wind turbine arrangement, the wake generated from upstream turbines can adversely affect the power performance and load characteristics of downstream turbines. Therefore, an optimal arrangement design, obtained through a detailed flow analysis focusing on wake interference, is necessary. In this study, to determine the power characteristics and annual energy production (AEP) of individual wind turbines, transient computational fluid dynamics, considering wind velocity variation (8 m/s, 11.7 m/s, 19 m/s, and 25 m/s), was conducted under different platform conditions ($0^{\circ}$, $22.5^{\circ}$, and $45^{\circ}$). The AEP was calculated using a Rayleigh distribution, depending on the wind turbine arrangement. In addition, we suggested an optimal arrangement design to minimize wake losses, based on the AEP.

Effect of Gd-DTPA on Diffusion in Canine Brain with Hyperacute Stroke (초급성 뇌경색을 일으킨 개에서 Gd-조영제의 주입이 뇌의 확산에 미치는 영향)

  • 김범수;정소령;신경섭
    • Investigative Magnetic Resonance Imaging
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    • v.6 no.2
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    • pp.158-165
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    • 2002
  • Purpose : To evaluate the effect of Gd-DTPA on signal intensity of diffusion-weighted magnetic resonance(MR) image and apparent diffuse coefficient (ADC) in dog brain with hype racute stroke. Materials and methods : Experimental canine model of hyperacute cerebral infarction was made by selective intraarterial embolization with particulate embolic material. Diffusion-weighted MR imaging was performed in five dogs at 1 hour after the embolization of internal carotid artery. After intravenous bolus injection of Gd- DTPA, additional 11 diffusion-weighted MR images were serially obtained from 2 minutes to 90 minutes after injection in each dog. The author evaluated findings of hyperacute cerebral infarction on diffusion-weighted MR imaging, and calculated mean signal intensity and mean ADC in infarcted region and contralateral normal region. Statistical analysis of mean signal intensity, mean ADC and contrast-noise ratio before and after Gd-DTPA injection was performed. Results : Hyperacute cerebral infarction developed in all five dogs on diffusion-weighted MR images obtained 1 hour after embolization. The area of hyperacute infarction had steady increase in signal intensity on diffusion-weighted MR image and decrease in ADC. In normal perfusion area, decrease in signal intensity was observed at 2 minutes the Gd-DTPA injection, whereas ADC did not changed. Conclusion : Intravenous injection of Gd-DTPA had no influence on ADC in both hyperacute infarction and normally perfused are a, but caused initial transient signal reduction in normally perfused area on diffusion-weighted MR image due to susceptibility effect of Gd-DTPA. It is important to calculate ADC in evaluating the effect of diffusion after injection of Gd-DTPA.

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Biochemical changes and drug residues in ascidian Halocynthia roretzi after formalin-hydrogen peroxide treatment regimen designed against soft tunic syndrome

  • Lee, Ji-Hoon;Kim, Ju-Wan;Shin, Yun-Kyung;Park, Kyung-Il;Park, Kwan Ha
    • Fisheries and Aquatic Sciences
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    • v.20 no.7
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    • pp.12.1-12.7
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    • 2017
  • Soft tunic syndrome (STS) is a protozoal disease caused by Azumiobodo hoyamushi in the edible ascidian Halocynthia roretzi. Previous studies have proven that combined formalin-hydrogen peroxide ($H_2O_2$) bath is effective in reducing STS progress and mortality. To secure target animal safety for field applications, toxicity of the treatment needs to be evaluated. Healthy ascidians were bathed for 1 week, 1 h a day at various bathing concentrations. Bathing with 5- and 10-fold optimum concentration caused 100% mortality of ascidians, whereas mortality by 0.5- to 2.0-fold solutions was not different from that of control. Of the oxidative damage parameters, MDA levels did not change after 0.5- and 1.0-fold bathing. However, free radical scavenging ability and reducing power were significantly decreased even with the lower-than-optimal 0.5-fold concentration. Glycogen content tended to increase with 1-fold bathing without statistical significance. All changes induced by the 2-fold bathing were completely or partially restored to control levels 48 h post-bathing. Free amino acid analysis revealed a concentration-dependent decline in aspartic acid and cysteine levels. In contrast, alanine and valine levels increased after the 2-fold bath treatment. These data indicate that the currently established effective disinfectant regimen against the parasitic pathogen is generally safe, and the biochemical changes observed are transient, lasting approximately 48 h at most. Low levels of formalin and $H_2O_2$ were detectable 1 h post-bathing; however, the compounds were completely undetectable after 48 h of bathing. Formalin-$H_2O_2$ bathing is effective against STS; however, reasonable care is required in the treatment to avoid unwanted toxicity. Drug residues do not present a concern for consumer safety.

Temperature-Induced Stresses and Deformation in Composite Box Girder Bridges (합성 박스형 교량의 온도에 의한 응력 및 변형)

  • Chang, Sung Pil;Im, Chang Kyun
    • Journal of Korean Society of Steel Construction
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    • v.9 no.4 s.33
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    • pp.659-672
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    • 1997
  • Thermal response induced from nonlinear temperature distribution in composite box gilder bridges depends on several variables(environmental conditions, physical and material properties, location and orientation of bridge, and cross-section geometry). In this paper, parametric study are conducted in order to find the effects of variations of seasons, location and orientation of bridge, sectional geometry and some material properties on the axial deformation, curvature and stresses in composite box girder bridge. A two-dimensional transient finite element model to conduct this parametric studies is briefly presented. Firstly, the effects of the parameters on the diurnal variation of curvature are considered, and for the time of maximum curvature, on the distribution of temperature and stresses of composite box girder sectional are considered. Finally, some considerations about the influence of the parameters on the daily maximum values of axial deformation, curvature and stresses are carried out. The influence of thermal effect on structures is important as much as the influence of live or dead load in some cases. In the design of steel composite bridges, the thermal stresses calculated on the supposition that the temperature difference between the concrete slab and steel girder is $10^{\circ}C$ and the temperature distributions are uniform in concrete slab and steel girder can be underestimated.

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A Study on the Characteristics Linked to Obesity with Acute Stroke Patients (급성기 뇌중풍환자의 비만 및 복부비만과의 임상적 제특징 분석)

  • Hsia, Yu-Chun;Jung, Ki-Yong;Go, Ho-Yeon;Jung, Hee;Baik, Jong-Woo;Yeo, Hyun-Soo;Ko, Mi-Mi;Choi, You-Kyung;Kim, Dong-Woo;Han, Chang-Ho;Cho, Ki-Ho;Bang, Ok-Sun;Park, Jong-Hyung;Ko, Seung-Gyu;Jun, Chan-Yong
    • The Journal of Internal Korean Medicine
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    • v.29 no.1
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    • pp.149-159
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    • 2008
  • Background and Purpose : This study was to assess characteristics in acute stroke patients according to obesity and abdominal obesity. Methods : From1 Oct. 2005 to 31 Jul. 2007, 629 patients with a stroke were included. Patients were hospitalized within 30 days after the onset of stroke at Kyungwon University Incheon Oriental Hospital, DongGuk University International Hospital and Department of Cardiovascular and Neurologic Disease (Stroke Center), or Kyung Hee University Oriental Hospital. We investigated general characteristics preferences according to general obesity and abdominal obesity. Results : 629 acute stroke patients were included in the final analysis. Male, young age, hypertension, heart disease in the family history, hypertension in the past history, care of invalids or grandchildren and Taeumin in the Sasang constitution were higher among obese patients. Male, transient ischemic attack in the past history, smoker and Soyangin in the Sasangconstitution were higher among patients with abdominal obesity. The incidence of abdominal obesity corresponded to lower education level. Conclusions : In our study, we observed the general disposition of various characteristic distributions according to obesity and abdominal obesity in acute stroke patients.

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Mapping of Human Cytomegalovirus IE1 Responsive Elements in the c-jun Promoter (사람세포거대바이러스 (Human Cytomegalovirus)의 극초기항원-1 (Immediate Early-1, IE-1)에 반응하는 c-jun Promoter의 유전자 지도 분석)

  • Park, Chung-Gyu;Han, Tae-Hee;Kim, Dae-Joong;Kim, Jin-Hee;Hwang, Eung-Soo;Choi, Sung-Bae;Cha, Chang-Yong
    • The Journal of Korean Society of Virology
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    • v.28 no.3
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    • pp.267-274
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    • 1998
  • Human cytomegalovirus (HCMV) has the ability to activate the expression of many viral and cellular genes. Among various viral proteins, the immediate early proteins (IE1-72kDa, IE2-86kDa) have been known to be potent transactivators. The product of c-jun proto-oncogene is important in cell activation and differentiation. Here, we tried to find out if the IE could activate the c-jun promoter and also tried to identify the responsible sequence elements in the c-jun activation by IE1-72kDa. We found HCMV IE expression transactivated the c-jun promoter in human embryonal lung fibroblasts (HEL). The activation fold by IE1-72kDa, IE2-86kDa and IE2-55kDa was 23, 35, and 5, respectively. When the expression of each IE was combined, it showed synergism. Expression of (IE1-72kDa + IE2-86kDa) and (IE1-72kDa + IE2-86kDa + IE2-55kDa) resulted in 131 and 162 fold increase, respectively. The c-jun promoter region between -117 and -59 contains binding sites for the transcription factors Spl, CAAT, AP-l like (ATF/CREB), and MEF2. Transient expression assays were performed using various reporter plasmids containing the c-jun promoter-regulatory region linked to the luciferase gene and a plasmid expressing HCMV IE1 gene. Deletional and point mutational analysis showed that the sequence between -225 to -160 and the CTF binding site were involved in the up-regulation of c-jun promoter.

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