• IMMUNE NETWORK
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• 제3권4호
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• pp.302-309
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• 2003

Background: CTLA4 (CD152), which is expressed on the surface of T cells following activation, has a much higher affinity for B7 molecules comparing to CD28, and is a negative regulator of T cell activation. In contrast to stimulating and agonistic capabilities of monoclonal antibodies specific to CTLA-4, CTLA4Ig fusion protein appears to act as CD28 antagonist and inhibits in vitro and in vivo T cell priming in variety of immunological conditions. We've set out to confirm whether inhibition of the CD28-B7 costimulatory response using a soluble form of human CTLA4Ig fusion protein would lead to persistent inhibition of alloreactive T cell activation. Methods: We have used CHO-$dhfr^-$ cell-line to produce CTLA4Ig fusion protein. After serum free culture of transfected cell line we purified this recombinant molecule by using protein A column. To confirm characterization of fusion protein, we carried out a series of Western blot, SDS-PAGE and silver staining analyses. We have also investigated the efficacy of CTLA4Ig in vitro such as mixed lymphocyte reaction (MLR) & cytotoxic T lymphocyte (CTL) response and in vivo such as experimental autoimmune encephalomyelitis (EAE), graft versus host disease (GVHD) and skin-graft whether this fusion protein could inhibit alloreactive T cell activation and lead to immunosuppression of activated T cell. Results: In vitro assay, CTLA4Ig fusion protein inhibited immune response in T cell-specific manner: 1) Human CTLA4Ig inhibited allogeneic stimulation in murine MLR; 2) CTLA4Ig prevented the specific killing activity of CTL. In vivo assay, human CTLA4Ig revealed the capacities to induce alloantigen-specific hyporesponsiveness in mouse model: 1) GVHD was efficiently blocked by dose-dependent manner; 2) Clinical score of EAE was significantly decreased compared to nomal control; 3) The time of skin-graft rejection was not different between CTLA4Ig treated and control group. Conclusion: Human CTLA4Ig suppress the T cell-mediated immune response and efficiently inhibit the EAE, GVHD in mouse model. The mechanism of T cell suppression by human CTLA4Ig fusion protein may be originated from the suppression of activity of cytotoxic T cell. Human CTLA4Ig could not suppress the rejection in mouse skin-graft, this finding suggests that other mechanism except the suppression of cytotoxic T cell may exist on the suppression of graft rejection.

• IMMUNE NETWORK
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• 제1권2호
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• pp.116-125
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• 2001

Background: Nitric oxide (NO) production has been described as a double-edged sword eliciting both pro- and anti-inflammatory effects in different immune reactions. This work was undertaken to investigate the immunoregulatory role of NO in experimental allergic encephalomyelitis (EAE) and experimental allergic uveitis (EAU). Method: We examined whether molsidomine (MSDM), a NO donor, administration to the myelin basic protein (MBP)- or interphotoreceptor retinoid binding protein (IRBP)-immunized rats could suppress EAE development by shifting toward the Th2 cytokine response. In the EAE experiments, the rats were treated orally with MSDM (10 mg/kg/day) at the early stage (-1~4 days) or throughout the experimental period (-1~15 days). Results: This resulted in significant amelioration of the disease and mild clinical symptoms, while MBP-immunization without MSDM administration showed severe EAE development. A marked reduction in inflammation was also observed in the spinal cord, indicating the crucial role of NO in the pathogenesis of EAE in in vivo. In the EAU experiments, a 24 h pre-treatment with MSDM prior to IRBP immunization resulted in significant inhibition of the disease. Furthermore, MSDM administration for 2 1 days completely reduced the incidence and severity of EAU. To investigate whether MSDM could modulate cytokine switching from Th 1 to Th2, culture supernatants of MBP- or IRBP-stimulated inguinal lymphocytes were analyzed. MSDM treatment enhanced IL-10 secretion but decreased IFN-${\gamma}$. IL-4 was undetectable in all groups. In contrast, the MBP-or IRBP-immunized rats without MSDM secreted high concentrations of IFN-${\gamma}$, but low concentrations of IL-10. Conclusion: In conclusion, NO administation suppresses EAE and EAU by modulating the Th1/Th2 balance during inflammatory immune responses. This work further suggests that NO may be useful in the therapeutic control of autoimmune disease.

• 대한수의학회지
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• 제38권1호
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• pp.173-181
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• 1998

Esherichia coli O157 : H7의 slt I, slt II, uid A 및 eaeA 4종 유전자를 동시에 검출하기 위한 multiplex PCR 기법을 확립하고 쇠고기중 직접 E coli O157 : H7 검출시험을 실시하였다. 4 set의 primers를 이용한 multiplex PCR 기법으로 31종의 장내세균에 대한 특이성을 조사한 결과 E coli O157 : H7 에서 1,087bp (eae A), 584bp (slt II), 348bp (slt I) 또는 252bp (uid A)크기의 DNA를 동시에 특이적으로 검출할 수 있었다. E coli O157 : H7 15주는 모두 uid A 및 eae A 유전자가 동시에 검출되었고, 다른 장내세균에서는 검출되지 않았다. slt I 또는 slt II 유전자를 가지고 있는 E coli 표준균주 24종을 이용하여 multiplex PCR 기법과 Vero cell cytotoxicity assay을 비교검사한 결과 베로톡신 산생능과 PCR법의 결과는 일치하였다. mutiplex PCR 기법의 쇠고기중 검출한계는 modified EC(mEC)에서 증균없이는 E coli O157 : H7균 $10^4cells/g$ 이상에서 검출이 가능하였으나 mEC에 1차 증균후 modified TSB 증균하였을 경우에는 10cells/g이하까지도 검출이 가능하였다. 개발된 multiplex PCR 기법을 쇠고기 40종에 직접 적용한 결과 E coli O157 : H7은 검출되지 않았으나 slt I 및 slt II유전자를 가지고 있는 E coli 4종이 검출되었으며, 이들의 혈청형은 O6, O112, O115 및 O139 였다. 이 연구에서 개발된 multiplex PCR은 쇠고기중 E coli O157 : H7을 신속하고 특이적으로 검출하는데 사용할 수 있을 것으로 사료된다.

• 대한수의학회지
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• 제54권4호
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• pp.209-218
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• 2014

Experimental autoimmune encephalomyelitis (EAE), an animal model of human multiple sclerosis (MS), reflects pathophysiologic steps in MS such as the influence of T cells and antibodies reactive to the myelin sheath, and the cytotoxic effect of cytokines. Galectin-9 (Gal-9) is a member of animal lectins that plays an essential role in various biological functions. The expression of Gal-9 is significantly enhanced in MS lesions; however, its role in autoimmune disease has not been fully elucidated. To identify the role of Gal-9 in EAE, we measured changes in mRNA and protein expression of Gal-9 as EAE progressed. Expression increased with disease progression, with a sharp rise occurring at its peak. Gal-9 immunoreactivity was mainly expressed in astrocytes and microglia of the central nervous system (CNS) and macrophages of spleen. Flow cytometric analysis revealed that $Gal-9^+CD11b^+$ cells were dramatically increased in the spleen at the peak of disease. Increased expression of tumor necrosis factor (TNF)-R1 and p-Jun N-terminal kinase (JNK) was observed in the CNS of EAE mice, suggesting that TNF-R1 and p-JNK might be key regulators contributing to the expression of Gal-9 during EAE. These results suggest that identification of the relationship between Gal-9 and EAE progression is critical for better understanding Gal-9 biology in autoimmune disease.

• 한국융합학회논문지
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• 제6권5호
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• pp.77-84
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• 2015

본 연구는 항균제에 내성을 보이는 대장균의 특성을 알아보기 위해 설사환자에서 분리된 대장균에 대한 항균제 감수성 및 병원성 인자의 상관성을 분자융합적 기술을 통해 조사하였다. 분리한 대장균의 항균제 내성은 60주에서 ESBL(extendede spectrum ${\beta}$-lactamase) positive균주가 8주이고, negative균주는 52주였다. ESBL 양성 8주 중 2주는 병원성 유전자가 검출되지 않았으며, stb(3주), flich7(1주), flich7-eae(2주)로 나타났다. ESBL 음성 52주 중 26주는 병원성 유전자가 검출되지 않았고, stx1(3주), stb(10주), flich7 및 eae(각 2주), stx1-flich7(2주), stx1-stb(4주), flich7-stb(2주), flich7-stb-eae(1주)이었다. 결론적으로 항균제 내성이 증가하는 시대에 분자 융합적 관점에서 독력 유전자의 분포와 항균제 내성과의 관계는 적게 나타났으나, 향후 다양한 독력 유전자의 분석을 통한 융합기술연구가 이루어진다면 보다 정확한 병원성 인자를 추정할 수 있을 것으로 사료된다.

• 대한수의학회지
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• 제43권4호
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• pp.525-529
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• 2003

The phosphorylation of extracellular signal-regulated kinases (p-ERK) in the spinal cord of rats with acute monophasic experimental autoimmune encephalomyelitis (EAE) was studied using immunohistochemistry and treatment with inhibitor. P-ERK is constitutively expressed in glial cells in the normal spinal cord. In EAE, some inflammatory cells in the subarachnoid space were positive for p-ERK at the early stage, and its immunoreactivity declined when those cells infiltrated the parenchyma at the peak stage. In a blocking experiment using its inhibitor, the intravenous administration of PD98059 from day 7 to 13 post-immunization did not modulate EAE paralysis. Considering the results, we postulate that intravenous administration of PD98059 is not effective in ameliorating EAE paralysis, although many inflammatory cells express ERK in the subarachnoid space.

• Journal of Microbiology
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• 제38권2호
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• pp.93-98
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• 2000

Nine hundred patients diagnosed with diarrhea or hemorrhagic uremic syndrome in the Kyungpook Province, Korea, were examined from November 1998 to February 2000. One patient in Kumi appeared to possess the Escherichia coli O157:H7 strain, which is very important in clinical decision making and public health action. The isolated strain, an E. coli O157:H7 KM, contained a 60 MDa plasmid and typical virulence genes including the verotoxin 2 gene, ehxA gene (encoding enterohemorrhagic hemolysin), and eae (encoding attaching and effacing protein-intimin) gene. This strain produced only verotoxin 2. Pulsed field gel electrophoretic analysis showed that the genomic organization of the E. coli O157:H7 KM strain may differ greatly from those of representative strains previously reported in the United States and Japan.

• 대한약학회:학술대회논문집
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• 대한약학회 2001년도 Proceedings of the Pharmaceutical Society of Korea
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• pp.134.2-135
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• 2001

• 한국축산식품학회지
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• 제30권1호
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• pp.148-153
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• 2010

시중에서 판매되는 계육과 돈육 가검물 1085개를 구입하고, 총 217개의 대장균 분리주를 확보하였다. 현재 알려진 대장균 병원성 인자 11개(astA, elt, estI, estII, cdt, cnf, eae, afa, stx, inve, aggR)의 보유여부를 PCR 기법으로 조사하였다. 돈육유래 대장균에서 존재하는 병원성 인자는 astA(6.9%), eae와 elt(4.6%), estII(4.0%), estI(2.3%), afa (1.1%), cnf(0.6%)의 순으로 검출되었으나, cdt, stx, aggR, inve는 검출되지 않았다. 계육에서 분리된 대장균 42개 중에서 astA와 inve를 보유하고 있는 병원성 대장균이 각각 1개씩 확인되었다. 특히 astA는 돈육과 계육에서 분리된 대장균 중에서 높은 빈도로 존재하고 있음을 확인하였으며, cnf와 afa같이 기존에 알려지지 않은 병원성 인자들의 존재도 확인되었다.

• ALGAE
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• 제26권4호
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• pp.343-350
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• 2011

Fermentation and enzyme-assisted extraction (EAE) improve nutritional and functional properties of foods by increasing the extraction of active compounds, ingestion rates, and body absorption. In this study, we investigated whether applying the EAE process improves the extraction and isolation efficiency of a polysaccharide from fermented Ecklonia cava (FE), which inhibited NO production in lipopolysaccharide (LPS)-activated RAW 264.7 cells. The results showed that the FE using the fungi Candida utilis and two different bacteria, namely Lactobacillus brevis and Saccharomyces cerevisiae increased protein and carbohydrate contents in comparison with those in non-fermented E. cava (NE). Aqueous extracts of fermented E. cava increased extraction yields and carbohydrate content, compared with the aqueous extract of NE. In addition, treating LPS-stimulated RAW 264.7 cells with aqueous extracts resulted in reduced NO production compared to that in LPS-treated cells. Ten EAEs of L. brevis-fermented E. cava (LFE) improved NO inhibitory effects in LPS-activated RAW 264.7 cells and the Viscozyme extract (VLFE) from the resulting extracts showed the highest NO inhibitory effect. We found that the >30 kDa fraction of VLFE led to markedly high inhibition of LPS-induced NO production as compared to that in the <30 kDa fraction. The crude polysaccharide isolated from >30 kDa fraction (VLFEP) consisted of fucose and markedly decreased NO production induced by LPS stimulation. VLFEP could be useful as an anti-inflammatory agent to suppress macrophage activation.