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The optimal conditions to improve retrovirus-mediated transduction efficiency to NIH 3T3 cells (레트로바이러스(retrovirus)의 NIH 3T3 세포로의 유전자 전달효율을 증가시키기 위한 적절한 조건들)

  • Lee, Jun Ah;Lee, Kang-Min;Lee, Hyun Jae;Lee, Yun Jeong;Kim, Dong Ho;Lim, Jung Sub;Park, Kyung-Duk
    • Clinical and Experimental Pediatrics
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    • v.50 no.10
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    • pp.1011-1017
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    • 2007
  • Purpose : We tried to assess the optimal conditions to improve low transduction efficiency and their effect on target cells. Methods : Cultured NIH 3T3 cells were incubated with retroviral vectors bearing an enhanced green fluorescent protein (eGFP) gene. We varied the ratio of viral vectors to target cells (1:1-1:8) and the number of transfections (${\times}1$, ${\times}2$), and compared transduction efficiencies. Also, the effects of polybrene on transduction efficiency and viability of target cells were assessed. Transduction of the eGFP gene was evaluated by observing NIH 3T3 cells under a fluorescence microscope and efficiencies were measured by the percentage of eGFP positive cells using FACscan. Results : As the ratio of retroviral vectors to target cells increased, transduction efficiency was greatly improved, from 7% (1:1) to 38% (1:4). However, transduction efficiency did not increase any more when the ratio increased from 1:4 to 1:8. Cells transfected twice showed higher transduction efficiencies than cells transfected once, at a ratio of 1:8. The eGFP gene transduced to NIH 3T3 cells sustained its expression during repeated passages. However, after the third passage (day 9), the percentage of eGFP positive cells began to decline. The degree of this decline in eGFP expression was lower in cells transfected twice than in cells transfected once (P<0.05). The addition of polybrene did not have any toxic effect on NIH 3T3 cells and greatly increased transduction efficiency (P=0.007). In addition to vector component, transduction efficiency was very sensitive to culture confluence. Cells cultured and transfected in 24-well plate showed higher transduction efficiency, although cells cultured in 6- well plate proliferated more (P=0.024). Conclusion : Our data could be used as a basis for retrovirus-based gene therapy. Further study will follow using human cells as target cells.

Estimation of Long-term Water Demand by Principal Component and Cluster Analysis and Practical Application (주성분분석과 군집분석을 이용한 장기 물수요예측과 활용)

  • Koo, Ja-Yong;Yu, Myung-Jin;Kim, Shin-Geol;Shim, Mi-Hee;Akira, Koizumi
    • Journal of Korean Society of Environmental Engineers
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    • v.27 no.8
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    • pp.870-876
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    • 2005
  • The multiple regression models which have two factors(population and commercial area) have been used to forecast the water demand in the future. But, the coefficient of population had a negative value because proper regional classification wasn't performed, and it is not reasonable because the population must be a positive factor. So, the regional classification was performed by principal component and cluster analysis to solve the problem. 6 regional characters were transformed into 4 principal components, and the areas were divided into two groups according to cluster analysis which had 4 principal components. The new regression models were made by each group, and the problem was solved. And, the future water demands were estimated by three scenarios(Active, moderate, and passive one). The increase of water demand ore $89.034\;m^3/day$ in active plat $49,077\;m^3/day$ in moderate plan, and $19,996\;m^3/day$ in passive plan. The water supply ability as scenarios is enough in water treatment plant, however, 2 reservoirs among 4 reservoirs don't have enough retention time in all scenarios.

Distribution of CD4+CD25+ T cells and graft-versus-host disease in human hematopoietic stem cell transplantation (사람의 동종 조혈모세포이식에서 CD4+CD25+ T세포의 분포와 이식편대숙주병)

  • Lee, Dae Hyoung;Chung, Nak Gyun;Jeong, Dae Chul;Cho, Bin;Kim, Hack Ki
    • Clinical and Experimental Pediatrics
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    • v.51 no.12
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    • pp.1336-1341
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    • 2008
  • Purpose : This study aimed to determine the frequencies of $CD4^+CD25^+$ T cells in donor graft and peripheral blood $CD4^+CD25^+$ T cells in recipients after hematopoietic stem cell transplantation (HSCT) and their association with graft-versus-host disease (GVHD). Methods : Seventeen children who underwent HSCT were investigated. $CD4^+CD25^+$ T cells in samples from donor grafts and recipient peripheral blood were assessed by flow cytometry at 1 and 3 months after transplantation. Results : $CD4^+CD25^+$ T cell frequencies in the grafts showed no significant difference between patients with and without acute GVHD (0.90% vs. 1.06%, P=0.62). Absolute $CD4^+CD25^+$ T cell number in grafts were lower in patients with acute GVHD than in those without acute GVHD ($6.18{\times}10^5/kg$ vs. $25.85{\times}10^5/kg$, P=0.09). Patients without acute GVHD showed a significant decrease in peripheral blood $CD4^+CD25^+$ T cell percentage at 3 months compared to those at 1 month after HSCT (2.11% vs. 1.43%, P=0.028). However, in patients with acute GVHD, $CD4^+CD25^+$ T cell percentage at 3 months was not different from the corresponding percentage at 1 month after HSCT (2.47% vs. 2.30%, P=0.5). Conclusion : The effect of frequencies of $CD4^+CD25^+$ T cells in donor grafts on acute GVHD after HSCT could not be identified, and the majority of peripheral blood $CD4^+CD25^+$ T cells in patients who underwent HSCT may be activated T cells related to acute GVHD rather than regulatory T cells. Further studies with additional markers for regulatory T cells are needed to validate our results.

Clinical application of D-dimer in Kawasaki Disease (가와사끼병에서 D-dimer의 임상적 응용)

  • Han, Jae Joon;Ko, Hong Ki;Yoo, Young;Lee, JungHwa;Lee, Kwang Chul;Son, Chang Sung;Lee, Joo Won
    • Clinical and Experimental Pediatrics
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    • v.50 no.2
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    • pp.205-208
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    • 2007
  • Purpose : Vascular endothelial cell damage and alteration of a fibrinolytic system was suggested to play a role in the development of coronary artery abnormalities in Kawasaki disease (KD). D-dimer is one of the markers of endothelial damage and fibrinolysis. We evaluated the clinical usefulness of D-dimer to differentiate KD from other febrile diseases and predict coronary artery abnormalities in KD. Methods : Sixty eight patients diagnosed as KD and twenty eight patients presented with acute febrile illnesses other than KD from September 2005 to July 2006 were included. Blood levels of D-dimer and various inflammatory markers were measured before treatment and the clinical course of KD was followed. Serial echocardiography was performed at the onset of disease and thereafter at a monthly interval for at least 2 months. Results : KD patients showed a higher D-dimer level than febrile controls, but the difference was not significant ($1.21{\pm}0.77{\mu}g/mL$ vs $0.92{\pm}0.71{\mu}g/mL$, P=0.083). Neither was the difference between KD patients who had coronary artery abnormalities and those who had not ($1.49{\pm}0.98{\mu}g/mL$ vs $1.15{\pm}0.71{\mu}g/mL$, P=0.169). D-dimer was significantly correlated with other inflammatory markers, such as C-reactive protein and erythrocyte sedimentation rate in both KD patients and febrile controls. Conclusion : D-dimer was not specific for KD. But it may be useful as an inflammatory marker to assess the severity of KD.

Distribution and Mobility of Herbicide $^{14}C$-Molinate in a Rice-Paddy-Soil Lysimeter (벼 재배 Lysimeter 환경에서 제초제 $^{14}C$-molinate의 분포 및 이동성 평가)

  • Park, Byung-Jun;Kim, Chan-Sub;Park, Kyung-Hun;Park, Hyeon-Ju;Im, Geon-Jae;Choi, Ju-Hyeon;Shim, Jae-Han;Ryu, Gab-Hee
    • The Korean Journal of Pesticide Science
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    • v.10 no.3
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    • pp.172-182
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    • 2006
  • This study was designed to assess molinate fate in the lysimeter by measuring the total radioactivity in the leachate, evolved $^{14}CO_2$, and $^{14}C$-residues in soil and rice plant. The amounts of applied $^{14}C$ in the leachate from the lysimeter for 20 weeks were 1.05% in 2.31 pore volume (217,465 mL) at the first and 0.34% in the second year, respectively. The amount of $^{14}CO_2$ evolved from the lysimeter accounted for 6.47% and 0.03% of applied $^{14}C$ in the first and second year. The $^{14}C$-activities in the soil layer of the lysimeter were distributed 18.0% (1st) and 13.3%(2nd) in the depth of 0 to 10 cm, 4.3 (1st) and 1.1% (2nd) in the depth of 10 to 20 cm. Most of the applied $^{14}C$ was detected in the top 20 cm soil layer. Total $^{14}C$ in rice plants grown at lysimeter were detected 11.46% of applied $^{14}C$. 11.11% in straw, 0.24% in brown rice grain, 0.08% in chaff and 0.03% in ears were distributed in the first year. Consequently, environmental fate of molinate using lysimeter simulating a paddy rice field were investigated 25.24% in soil, 11.64% in rice plant, 1.05% in leachate, 6.74% in evolved $^{14}CO_2$ and 0.02% in volatilized organic chemicals in the first year.

Study of the Optimize Radiotherapy Treatment Planning (RTP) Techniques in Patients with Early Breast Cancer; Inter-comparison of 2D and 3D (3DCRT, IMRT) Delivery Techniques (유방암 방사선치료 시 최적의 방사선치료계획기법에 대한 고찰)

  • Kim, Young-Bum;Lee, Sang-Rok;Chung, Se-Young;Kwon, Young-Ho
    • The Journal of Korean Society for Radiation Therapy
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    • v.18 no.1
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    • pp.35-41
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    • 2006
  • Purpose: A various find of radiotherapy treatment plans have been made to determine appropriate doses for breasts, chest walls and loco-regional lymphatics in the radiotherapy of breast cancers. The aim of this study was to evaluate the optimum radiotherapy plan technique method by analyzing dose distributions qualitatively and quantitatively. Materials and Methods: To evaluate the optimum breast cancer radiotherapy plan technique, the traditional method(two dimensional method) and computed tomography image are adopted to get breast volume, and they are compared with the three-dimensional conformal radiography (3DCRT) and the intensity modulated radiotherapy (IMRT). For this, the regions of interest (ROI) such as breasts, chest walls, loco-regional lymphatics and lungs were marked on the humanoid phantom, and the computed tomography(Volume, Siemens, USA) was conducted. Using the computed tomography image obtained, radiotherapy treatment plans (XiO 5.2.1, FOCUS, USA) were made and compared with the traditional methods by applying 3DCRT and IMRT. The comparison and analysis were made by analyzing and conducting radiation dose distribution and dose-volume histogram (DVH) based upon radiotherapy techniques (2D, 3DCRT, IMRT) and point doses for the regions of interest. Again, treatment efficiency was evaluated based upon time-labor. Results: It was found that the case of using 3DCRT plan techniques by getting breast volume is more useful than the traditional methods in terms of tumor delineation, beam direction and confirmation of field boundary. Conclusion: It was possible to present the optimum radiotherapy plan techniques through qualitative and quantitative analyses based upon radiotherapy plan techniques in case of breast cancer radiotherapy. However, further studies are required for the problems with patient setup reproducibility arising from the difficulties of planning target volume (PVT) and breast immobilization in case of three-dimensional radiotherapy planning.

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Potential Allelic Association of Microsatellite Markers on Chromosome 1 with Economic Traits in Korean Native Chicken (한국재래닭 1번 염색체내 초위성체 유전표지를 이용한 경제형질 연관 지역 탐색)

  • Kim, H.K.;Oh, J.D.;Kang, B.S.;Park, M.N.;Chae, E.J.;Jung, H.M.;Seo, O.S.;Choe, H.S.;Jeon, G.J.;Lee, H.K.;Kong, H.S.
    • Korean Journal of Poultry Science
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    • v.35 no.2
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    • pp.163-169
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    • 2008
  • A total of 17 polymorphic microsatellite markers on chromosome 1 were used for allelic association tests with phenotypic traits in Korean native chicken. Chi-square tests were performed to compare the frequencies of individual alleles between the high and the low trait groups. The frequency of allele 123 of MCW0160 showed a significant difference between the high and the low groups in the trait of egg weight (EW). Three markers, namely ADL0234, UMA1.125 and ADL0101, were found to show significant differences in allelic distribution for the trait of the first lay day (FLD). UMA1.117, ADL0020, UMA 1.019, LMA1 and ADL0238 were found to show significant differences in allelic distribution for the trait of body weight (BW). ADL0101 and ADL0238 were found to show significant differences in allelic distribution for the trait of number of egg production(EP). In this study, we identified the QTL for economic traits at around 94 (MCW0160), 151 (ADL0234), 170 (UMA1.125), 225 (UMA1.117), 285 (ADL0020), 387 (UMA1.019), 418 (LMA1), 500 (ADL0101) and 520 (ADL0238) cM on chromosome 1 in Korean native chicken. The results provided a useful guideline for identification of positional candidate gene and marker-assisted selection for economic traits in Korean native chicken.

Irradiation Alone in Stage IB, IIA, and IIB Cervix Cancer : I Analysis of Survival and Failure Patterns (자궁경부암 병기 IB, IIA, IIB에서 방사선 단독치료성적 : I 생존기간 및 재발양상에 관한 분석)

  • Ahn Sung Ja;Chung Woong Ki;Nah Byung Sik;Nam Taek Keun;Choi Ho Sun;Byun Ji Soo
    • Radiation Oncology Journal
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    • v.15 no.2
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    • pp.129-136
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    • 1997
  • Purpose : We analyzed the survival and failure patterns of cervix cancer patients treated with irradiation alone to evaluate our treatment method and to compare with the others Methods and Materials : Two hundred and twenty cervical cancer patients, Stage IB, II A, and II B who completed the planned treatment between Mar 1987 and December 1991 were analyzed retrospectively. The Stage IB patients were restaged to the Stage IB1 and IB2 by the recently revised FIGO classification, Patients were treated with a combination of external irradiation and the intracavitary brachytherapy Determination of the tumor control was done at the time of 6 months Postirradiation. The follow-up time was ranged from 3 to 115 months and the mean was 62 months and the follow-up rate was $93.6\%$(206/220) Results : The overall 5-year survival rate of Stage IB1 (N=50), IB2(N: 15). II A(N=58), and II B(N=97) was $94\%,\;87\%,\;69\%,\;and\;56\%$. respectively. In the univariate analysis of prognostic factors, stage(0.00), initial Hg level (P=0.00), initial TA-4(tumor-associated) antigen level(p= 0.02), initial CEA level(p=0.02), barrel-shaped tumor(p=0.02), whole cervical involvement (0.00), pelvic tyrnphadenopathy(LAP) in CT(p=0.04), and Post-irradiation adiuvant chemotherapy(P=0.00) were statistically significant in survival analysis. In a while multivariate analysis showed that the stage was the most powerful Prognostic indicator and the Post-irradiation chemotherapy factor also showed the statistical significance. The overall local control rate was $81\%$ and by the stage, $100\%$ in Stage IBI, $86.7\%$ in Stage IBS, $84.5\%$ in Stage IIA, and $68.1\%$ in Stage IIB, respectively The overall tumor recurrence rate was $15.5\%$(27/174) and by the stage, $8\%$(4/50) in Stage IB1, $0\%$(0/l3) in Stage IB2, $22.4\%$(l1/49) in Stage II A, and $19.4\%$(12/62) in Stage II B, respectively. Conclusions : We obtained the similar treatment results to the other's ones in early stage cervical cancer patients. But in Stage II B, the local control rate was lower than that of the other institutes and also the survival was poorer. So it seems to be necessary to reevaluate the treatment method in advanced cervical cancer patients.

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Is Nitrogen Uptake Rate by Phytoplankton below the Euphotic Zone in the Yellow Sea Considerable? (황해의 무광대에서 식물플랑크톤에 의한 질소 섭취율은 상당한가?)

  • Yang, Sung-Ryull;Shim, Jae-Hyung;Chung, Chang-Soo;Hong, Gi-Hoon;Pae, Se-Jin;Yang, Dong-Beom;Park, Myung-Gil
    • The Sea:JOURNAL OF THE KOREAN SOCIETY OF OCEANOGRAPHY
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    • v.7 no.2
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    • pp.60-67
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    • 2002
  • To determine whether nitrogen (N) uptake by phytoplankton below the euphotic zone in the Yellow Sea is considerable, we measured the uptake rates of nitrate and ammonium using $^{15}N$-labeled stable isotope $K^{15}NO_{3}$ and $^{15}NH_{4}Cl$, in May and November 1997 at total 10 stations. Depth-integrated uptake rates of nitrate and ammonium over the euphotic zone during this study ranged from 1.8 to 15.3 mg N $m^{-2}$ $d^{-1}$ and from 5.0 to 132.2 mg N $m^{-2}$ $d^{-1}$, respectively, and ammonium uptake predominated at 9 of 10 stations (1.9-19.4 fold). Depth-integrated uptake rates of nitrate and ammonium over the whole water column ranged from 2.9 to 22.0 mg N $m^{-2}$ $d^{-1}$ and from 15.7 to 175.5 mg N $m^{-2}$ $d^{-1}$, respectively. The significant proportion of whole water column N uptake was attributed to uptake by phytoplankton below the euphotic zone, ranging from 13.0 to 86.2% for nitrate and from 13.8 to 67.8% for ammonium, indicating that phytoplankton N uptake below the euphotic zone is at times considerable in the study area. The results suggest that when phytoplankton below the euphotic zone in the Yellow Sea are again entrained into the euphotic zone by a certain physical forcing such as turbulent mixing and the vertical movement of thermocline, these episodic events may significantly affect the material fluxes within the euphotic zone. Furthermore, the results suggest that a portion of regenerated production estimated from $^{15}N$-ammonium uptake should be included in new production estimates, which otherwise could be underestimated in the Yellow Sea.

Expression of Hypoxia-inducible Factor-1 $\alpha$ in Esophageal Squamous Cell Carcinoma: Relationship to Prognosis and Tumor Biomarkers (식도 편평세포암에시 Hypoxia-inducible Factor-1 $\alpha$의 발현: 예후와 종양표지자와의 상관성)

  • 양일종;김종인;이해영;천봉권;조성래
    • Journal of Chest Surgery
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    • v.37 no.8
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    • pp.691-701
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    • 2004
  • Background: Tissue hypoxia is a characteristic of many human malignant neoplasms, and hypoxia inducible factor-1 (HIF-1) plays a pivotal role in essential adaptive response to hypoxia, and activates a signal pathway for the expression of the hypoxia-regulated genes, resulting in increased oxygen delivery or facilitating metabolic adaptation to hypoxia. Increased level of HIF-1 a has been reported in many human malignancies, but in esophageal squamous cell carcinoma, the influence of HIF-1 a on tumor biology, including neovascularization, is not still defined. Material and Method: The influence of HIF-1 a expression on angiogenic factors, correlation between the tumor proliferation and HIF-1 a expression, interaction of HIF-1 a expression and p53, and correlation between HIF-1 a expression and clinicopathological prognostic parameters were investigated, using immunohistochemical stains for HIF-1 a, VEGF, CD34, p53, and Ki-67 on 77 cases of resected esophageal squamous cell carcinoma. Result: HIF-1 a expression in cancer cells was found in 33 of 77 esophageal squamous cell carcinoma cases. The 33 cases (42.9%) showed positive stain for HIF-1 a. High HIF-1 a expression was significantly associated with several pathological parameters, such as histologic grade (p=0.032), pathological TMN stage (p=0.002), the depth of tumor invasion (p=0.022), regional lymph node metastasis (p=0.002), distant metastasis (p=0.049), and lymphatic invasion (p=0.004). High HIF-1 a expression had significant VEGF immunoreactivity (p=0.008) and Ki-67 labeling index (p<0.001), but was not correlated with microvascular density within tumors (p=0.088). The high HIF-1 a expression was correlated with aberrant p53 accumulation with a marginal significance (p=0.056). The overall 5-year survival rate was 34.9%. The survival rate of patients with a high HIF-1 a expression was worse than that of patients with low-expression tumors (log-rank test, p=0.0001). High HIF-1 a expression was independent unfavorable factors although statistical significance is marginal in multivariate analysis. Conclusion: It is suggested that (1) high HIF-1 a expression in esophageal squamous cell carcinoma is associated with tumor hypoxia, or with genetic alteration in early carcinogenesis and progressive stages, (2) high HIF-1 a expression may be associated with intratumoral neovascularization through HIF-VEGF pathway, and (3) high HIF-1 a expression is associated with poor prognosis in patients with esophageal squamous cell carcinoma and may playa role as biomarker for regional lymph node metastasis.