• Title/Summary/Keyword: Lymphocyte culture

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Isolation and molecular characterizations of canine distemper virus from a naturally infected Korean dog using Vero cells expressing dog signaling lymphocyte activation molecule

  • Yang, Dong-Kun;Kim, Ha-Hyun;Lee, Siu;Yoon, Yoon-Seek;Park, Jungwon;Oh, Dongryul;Yoo, Jae Young;Ji, Miryeon;Han, Bokhee;Oh, Subin;Hyun, Bang-Hun
    • Journal of Veterinary Science
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    • v.21 no.5
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    • pp.64.1-64.14
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    • 2020
  • Background: Canine distemper virus (CDV) infection results in high morbidity and mortality in dogs. There has been no report about Isolation of Korean CDV since 1980 in Korea. Objectives: To investigate the biological properties and the genetic characterization of Korean CDV. Methods: Vero cells expressing dog signaling lymphocyte activation molecule (dSLAM) gene named as Vero/dSLAM were used to isolate CDV using 17 samples. Diagnostic methods such as cytopathic effects, immunofluorescence assay, peroxidase linked assay, electron microscopy, rapid immunodiagnostic assay, and reverse transcription polymerase chain reaction were used to confirm the Korean CDV isolate as a CDV. The genetic analysis was performed through cloning and sequencing of hemagglutinin gene of CDV isolate. Results: A virus propagated in Vero/dSLAM cell was confirmed as CDV (CD1901 strain) based on the above methods. The CD1901 strain showed the highest viral titer (105.5 50% tissue culture infectious dose [TCID50]/mL) in the Vero/dSLAM cells at 4 days post inoculation, but did not form a fork on chorioallantoic membrane of 7-day-old egg. Ribavirin, a nucleotide analogue anti-viral agent, inhibits moderately the Korean CDV propagation in the Vero/dSLAM cells. The nucleotide and amino acid sequences of the H gene of CD1901 strain were compared with those of other CDV strains. The CD1901 strain belonged to Asia 1 group and had the highest similarity (99.9%) with the BA134 strain, which was isolated in China in 2008. Conclusions: We constructed successfully Vero/dSLAM and isolated one Korean CDV isolate (CD1901 strain) from a naturally infected dog. The CD1901 strain belonged to Asia 1 genotype.

Induction of Rice Allergen-Specific IgE Synthesis by KU8l2 Cells (KU812세포에 의한 쌀 알레르겐 특이적 IgE항체 합성의 유도)

  • Shim, Sun-Yup;Katakura, Yoshinori;Shirahata, Sanetaka
    • Journal of Life Science
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    • v.17 no.11
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    • pp.1492-1496
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    • 2007
  • In vitro IgE class switching could be induced through co-culture of CD40L-expressing KU812 cells and CD40-expressing B cells in the presence of IL-4 or IL-13. It has been generated several B cell lines, which produce rice allergen (RA)-specific IgM antibody by in witγo immunization (IVI) using peripheral blood lymphocyte (PBL). In this study, induction of RA-specific IgE antibody by KU812 cells was attempted. Before co-culture, we determined the CD40 expression in RA-specific B cell lines, RA9G11 and the CD40 ligand (CD40L) expression in activated KU812 cells by treatments with phorbol myristate acetate (PMA) and ionomycin for 6 hrs. Flow cytometric analysis shown that RA9G11 and activated KU812 cells expressed high level of CD40 and CD40L, respectively. RA9G11 cells were cultured with activated KU812 cells for 12 days in the presence of IL-4 for IgE class switching. Mature $C{\varepsilon}$ mRNA level and RA-specific IgE spot forming cells (SFC) were observed in all culture condition, and especially, high level of RA-specific IgE synthesis was determined the same ratio of RA9G11 and activated KU812 cells in the presence of 50U IL-4. Therefore, induction of RA-specific IgE synthesis by activated KU812 cells can be contributed in the application for allergic therapy and prevention.

Prospective Study of the Immunologic Factors Affecting the Prognosis of Severe Community-Acquired Pneumonia (중증 지역사회획득 폐렴환자의 예후에 영향을 미치는 면역지표에 대한 연구)

  • Hwang, Jae-Kyung;Lee, Ho-Moeng;Song, Kwang-Sik;Park, Gye-Young;Park, Jeong-Woong;Park, Jae-Kyung;Jeong, Seong-Hwan;Ahn, Jeong-Yeal;Seo, Yiel-Hea;Nam, Gui-Hyun
    • Tuberculosis and Respiratory Diseases
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    • v.50 no.4
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    • pp.437-449
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    • 2001
  • Background : In the severe community-acquired pneumonia, it has been known that the immune status is occasionally suppressed. This study was performed to identify the immunologic markers related with the prognostic factors in severe community-acquired pneumonia. Methods : 23 patients with severe community-acquired pneumonia were involved in this study, and divided into survivor (16) and nonsurvivor (7) groups. In this study, the medical history, laboratory tests(complete blood counts, routine chemistry profile, immunoglobulins, complements, lymphocyte subsets, cytokines, sputum and blood culture, urine analysis), and chest radiographs were scrutinized. Results : 1) Both groups had lymphopenia(total lymphocyte count $995.6{\pm}505.7/mm^3$ in the survivor and $624.0{\pm}287.6/mm^3$ in the nonsurvivor group). 2) The T-lymphocyte count of the nonsurvivor group($295.9{\pm}203.0/mm^3$) was lower than the survivor group($723.6{\pm}406.5/mm^3$) (p<0.05). 3) The total serum protein(albumin) was $6.0{\pm}1.0(2.7{\pm}0.7)\;g/d{\ell}$ in the survivor and $5.2{\pm}1.5(2.3{\pm}0.8)g/d{\ell}$ in the nonsurvivor group. The BUN of the nonsurvivor group($41.7{\pm}30.0mg/d{\ell}$) was higher than that of the survivor group($18.9{\pm}9.8mg/d{\ell}$)(p<0.05). The creatinine concentration was higher in the nonsurvivor group($1.8{\pm}1.0mg/d{\ell}$) than that in the survivor group($1.0{\pm}0.3mg/d{\ell}$)(p<0.05). 4) The immunoglobulin G level was higher in the survivor group($1433.0{\pm}729.5mg/d{\ell}$) than in the nonsurvivor group($849.1{\pm}373.1mg/d{\ell}$) (p<0.05). 5) The complement $C_3$ level was $108.0{\pm}37.9mg/d{\ell}$ in the survivor group and $88.0{\pm}32.1mg/d{\ell}$ in the nonsurvivor group. 6) A cytokine study showed an insignificant difference in both groups. 7) Chronic liver disease, DM, and COPD were major underlying diseases in both groups. Conclusion : These results suggest that decreased a T-lymphocyte count and immunoglobulin G level, and an increased BUN and creatinine level may be associated with the poor prognosis of severe community-acquired pneumonia.

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Anti-stress Effect by the Treatment of Vitamin C (Vitamin C 투여에 의한 항스트레스 효과)

  • 오찬호;최동성
    • KSBB Journal
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    • v.8 no.5
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    • pp.424-430
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    • 1993
  • The anti-stress effect by the treatment of vitamin C was investigated in this study. The treatment of ascorbic acid in the presence of $Cu^{2+}$ion induced strong time- and dose-dependent degradation of hitamine, and also the addition of histamine accelerated time-dependent decomposition of ascorbic acid in vitro. The treatment of ascorbic acid in $ODS^{od}/_{od}$rats, which cannot synthesize ascorbic acid, significantly decreased the urinary histamine. The protreatment of ascorbic acid, dexamethasone and promethazine inhibited the lethal effect induced by immobilization stress, but that of dimethylsulfoxide did not. The addition of ascorbic and to a culture of spleen cells of $ODS^{od}/_{od}$rats significantly increased the Con A-dependent T lymphocyte proliferation.

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The Immunoadjuvant Activity of The Water-Extract of Korean mistletoe (Viscum album var. coloratum) Fruit (한국산 겨우살이 열매 추출물의 Immunoadjuvant 효과)

  • Lee, Jung-Lim;Ahn, Jae-Hyung;Hwang, Seong-Gu;Jung, Yeon-Hwa;Yang, Hyo-Seon;Kang, Tae-Bong;Kim, Jong-Bae;Yoo, Yung-Choon
    • Korean Journal of Pharmacognosy
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    • v.41 no.4
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    • pp.275-281
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    • 2010
  • To evaluate the immunomodulatory activity of a water extract (KMF-WE) of Korean mistletoe (Viscum album var. coloratum) fruit, we examined its ability to induce humoral and cellular immune response against keyhole limpet hemocyanine (KLH). Immunized mice with KLH admixed with KMF-WE (KLH/KMF-WE) showed significant induction of KLH-specific antibodies compared to mice immunized with KLH alone. The assay for determining isotypes of antibodies revealed that KMFWE augmented KLH-specific-IgG1 and -IgG2a production. In vitro T lymphocyte proliferation analysis against KLH revealed that the splenocytes of mice immunized with KLH/KMF-WE showed a significantly higher proliferative ability than those from mice immunized with KLH alone. The culture supernatants of splenocytes, which were harvested from mice immunized with KLH/KMF-WE, showed higher levels of both Th-1 type (IL-2, IFN-${\gamma}$) and Th-2 type (IL-4) cytokines in response to KLH stimulation compared to those from mice immunized with KLH alone. Also, in delayed-type hypersensitivity (DTH) assay, mice immunized with KLH/KMF-WE showed a significantly higher reaction to KLH than mice treated with KLH alone. These results suggest that KMF-WE possess immunoadjuvant activity to enhance both antigen-specific humoral and cellular immune responses against protein antigens (KLH).

Differentiation and Labeling of Mouse Preadipocytes for Allogenic Transplantation Study (동종이식 연구를 위한 마우스 지방전구세포의 표지 및 분화 방법의 확립)

  • Kim, In Ok;Kim, Taek Seung;Kim, Mi Hyung;Hyon, Won Sok;Mun, Goo Hyun;Oh, Kap Sung;Bang, Sa Ik
    • Archives of Plastic Surgery
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    • v.32 no.4
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    • pp.533-538
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    • 2005
  • Due to its safety and softness, autologous fat transplantation has been commonly performed for soft tissue correction. However, the injected fat is absorbed resulting in the reduction of volume of the graft by 40-60% within a few months. Thus, there was an attempt to use adipocytes differentiated from preadipocytes in vitro for transplantation. Differentiated adipocytes were biocompatible and matured with gradual volume increase at transplantation site in clinical study(unpublished data). In addition, they did not induce immune rejection in response to nonself lymphocytes in a mixed lymphocyte reaction(MLR)(unpublished data). The purpose of this study is to differentiate mouse preadipocytes following labeling into adipocytes to establish an animal model for allogenic transplantation. Preadipocytes isolated from inguinal and retroperitoneal fat pad of C57BL/6 mice were proliferated with growth medium by passage 3 and differentiated into adipocytes with different culture conditions after labeled with BrdU. At most suitable conditions, above 90% of preadipocytes were differentiated and BrdU labeling did not affect differentiation rate and function of differentiated adipocytes. These results demonstrate that BrdU-labeled adipocytes resulting from this in vitro differentiation protocol are useful for allogenic transplantation study.

Expression of the Functional Recombinant Interleukin-16 in E. coli and Mammalian Cell Lines

  • Kim, Seon-Young;Lee, Chang-Hun;Kim, Kyung-Joo;Kim, Yeon-Soo
    • Journal of Microbiology and Biotechnology
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    • v.11 no.2
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    • pp.234-241
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    • 2001
  • The C-terminal 393 bp region of the human interleukin-16 (IL-16) gene was cloned and expressed in E. coli along with mammalian cell lines. Recombinant IL-16 expressed from E. coli was 22 kDa on SDS-PAGE and showed 260% of chemoattractant activity at a concentration of $0.1\;{\mu}g/ml$. HeLa, COS, and Neuro-2a cells were transduced by recombinant retrovirus vector pLNC/IL-16/IRES/TK and the intracellular and secreted amounts of IL-16 produced by HeLa/IL-16/TK, COS/IL-16/TK, and Neuro-2a/IL-16/TK cells were determined by enzyme-linked immunosorbent assay (ELISA). HeLa/IL-16/TK $(1{\times}10^5)$ and COS/IL-16/TK $(1{\times}10^5)$ cells secreted 36.1 and 13.3 ng of IL-16 for 48 h, respectively. Forty-nine ng and 86.4 ng of IL-16 remained in the cell lysates of HeLa/IL-16/TK and COS/IL-16/TK. Intracellular and secreted amounts of IL-16 from Neuro-2a/IL-16/TK $(5{\times}10^5)$ cells during 24 h cultivation were 50 ng and 3.3 ng, respectively. Also, HeLa and COS cells wee stably transfected with mammalian expression vector pCRIII/IL-16. Both culture media and cell lysates prepared from HeLa/IL-16 cells and COS/IL-16 cells showed chemoattractant activity ranging from 190% to 460% as compared to the control experiment. Expression of the herpes simplex virus thymidine kinase (HSV0tk) gene in pLNC/IL-16/ IRES/TK bicistronic retroviral expression vector was verified by performing a genciclovir (GCV) sensitivity assay. Finally, IL-16 repressed Tat-transactivated human immunodeficiency virus type 1 long terminal repeat (HIV-1 LTR) promoter activity.

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Effects of Nano-sized Carbon Black on the Lungs of High Fat-diet Induced Overweight Rats

  • Lim, Cheol-Hong;Kang, Mingu;Han, Jeong-Hee;Yun, Hyo-In
    • Environmental Analysis Health and Toxicology
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    • v.28
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    • pp.14.1-14.9
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    • 2013
  • Objectives This study was conducted to determine whether nano-sized carbon black exposure results in greater damage in high fat diet-induced overweight rats than normal weight ones and to identify the possible causes of any differences. Methods Two groups of Sprague-Dawley rats allocated by body weight (normal and overweight) were exposed to aerosolized nano-sized carbon black for 6 hours a day, 5 days per week over a 4-week period. Differential cell counts, lactate dehydrogenase (LDH) activities and albumin concentrations were measured in bronchoalveolar lavage (BAL) fluid, and histopathological findings in the lungs were evaluated. Tumor necrosis factor-alpha (TNF-${\alpha}$) and interleukin (IL)-6 were measured in BAL fluid and supernatants of lipopolysaccharide(LPS)-stimulated lymphocyte culture. Results Rats exposed to high concentrations of nano-sized carbon black showed significantly increased (p <0.05) polymorphonuclear leukocyte number and LDH activity in the BAL fluid from both overweight and normal rats. Mild histopathological changes were observed in normal rats irrespective of carbon black concentrations. However, severe histological scores were found in overweight rats ($1.75{\pm}0.46$, $2.25{\pm}0.46$, and $2.88{\pm}0.35$ after low, medium, and high concentration exposures). Proinflammatory cytokine levels of TNF-${\alpha}$ and IL-6 were significantly higher in the supernatant of LPS-stimulated lymphocytes of overweight rats, whereas there was no significant difference in the BAL fluid between normal and overweight rats. Conclusions Inflammation and damage to lungs exposed to nano-sized carbon black was more severe in high fat diet-induced overweight rats compared to normal rats.

The Effects of Juglandis Semen Extract on the Experimental Asthma induced by Ovalbumin (호도(胡桃) 물추출물이 Ovalbumin으로 유도된 천식동물모델에 미치는 영향)

  • Kim, Sang-Chan;Byun, Sung-Hui
    • Herbal Formula Science
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    • v.13 no.1
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    • pp.145-159
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    • 2005
  • Objective : Juglandis Semen has a function that to invigorate the lung and kidney. And It is commonly used as a supporting agent in the treatment of coughing and bronchitis. This study was performed to investigate the effect of oral administration of Juglandis Semen Extract (JSE) against the experimental asthma induced by ovalbumin. Methods : Asthma was induced to Balb/c mouse by i.p. injection and aerosol immunization with ovalbumin. It was observed the change of the cell number in the BAL fluid. Concentrations of IL-4, IL-5 in splenoc yte were assessed by ELISA, IgG and IgE from serum were calculated by same method. Results : 1. Number of macrophage in BAL fluid was significantly decreased in JSE group compared with control group, but not eosinophil and lymphocyte. 2. Levels of IgG and IgE in serum were significantly decreased in JSE group compared with control group, respectively. 3. Concentration of IL-4 in culture supernatant of splenocyte was significantly decreased in JSE group compared with control group, but there was no significant in IL-5. Conclusion : We found that the effect of JSE extract in asthma was implicated in reduction of IL4released from Th2 cell, and decreases of IgG and IgE from plasma cell. These findings suggest that JSE can produce anti-asthmatic effect, which may play a role in allergen-induced asthma therapy.

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The effects of properties and interactions of surface molecules in antigen presenting cells on T cell activation (인공 항원제시세포의 표면 분자의 특성 및 상호작용이 T 세포 활성화에 미치는 영향)

  • Min, Youngsil;Kang, Yoon Joong
    • Journal of Convergence for Information Technology
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    • v.10 no.6
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    • pp.164-176
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    • 2020
  • Efficient production of antigen specific cytotoxic T cells is critical for appropriate adoptive immune response. In vitro culture and expansion of human T lymphocyte clones are very sophisticated and subtle procedure in immune cell therapy and hard to control. Therefore, many groups devoted their efforts to manipulate artificial antigen presenting cells (aAPCs) that can induce T cell activation and clonal expansion. To mimicking of natural antigen-presenting cells, aAPCs encompass basic signal molecules required for T cell activation: MHC:antigen complexes, co-stimulatory molecules and soluble immune modulating molecules. Orchestrated organization of these molecules is important for efficient T cell activation. Here, we discuss how those molecules have been incorporated in several aAPC models, but also how physical properties od aAPC are important for interaction with T cells.