감초 (Glycyrrhiza uralensis Fisch, GLU) 가천식모델생쥐의 BALF 내면역세포및 Cytokine 에미치는효과
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1 감초 (Glycyrrhiza uralensis Fisch, GLU) 가천식모델생쥐의 BALF 내면역세포및 Cytokine 에미치는효과 Effects of Glycyrrhiza uralensis Fisch on Immunocyte and Cytokine Production in Asthma Model Mouse Young-Joo Han, Yang-Chun Park Department of Internal Medicine College of Oriental Medicine, Daejeon University, Daejeon, Korea Objective : The aim is to identify the effects of Glycyrrhiza uralensis Fisch on immunocyte and cytokine production in asthmatic laboratory mice. This experiment was designed to investigate the antiallergic and antiinflamatary the effects of Glycyrrhiza uralensis Fisch on asthma. Materials and Methods : We measured the eosinophil, IL-4, IL-5, IL-13, IFN-, CD4, CD8, CD69, CCR3, CD11b, Gr-1 in bronchoalveolar lavage fluid of ovalbumin induced asthmatic mice. Results : Glycyrrhiza uralensis Fisch increased the proliferation of eosinophils, IL-4, IL-5, IL-13, IgE, granulocyte, CCR3, CD4, IgE, CD69. Glycyrrhiza uralensis Fisch increased the proliferation of IFN-. Conclusion : Results suggest that Glycyrrhiza uralensis Fisch extract is useful in treatment and prevention allergic asthma. Key Words: Glycyrrhiza uralensis Fisch, cytokine, antiallergy, :, (Tel , Fax : , omdpyc@dju.ac.kr), 1-3.,,,,,,,
2 ,,,,,,,,, 3.,,, 6.,,,,,,, 7. 8, Wang 9., IL-4, IL-5, IL-13, IgE, IFN-, CD4, CD8, CD69, CCR3. 1) 18~25g C57BL/6( ), ( 22.1%, 8.0%, 5.0%, 8.0%, 0.6%, 0.4% Co.), 22 2, 50 10%, 12 (07:00~19:00), 150~300Lux 2. 2) (Glycyrrhiza uralensis: GLU). 1) 200g 2,000 3 (Rotary evaporator, BUCHI B-480, Switzerland). (Freeze dryer, EYELA FDU-540, Japan) (-84 ). 2) 500 g/ (OVA, chicken egg ovalbumin; Grade ) 10% (w/v) aluminum potassium sulfate (Alum; Sigma) PBS. 10N NaOH ph g 5. OVA/Alum (pellet) 100 g OVA (500 / ) / , PBS Alum,. 3) OVA/Alum 3 (150 /kg) 5.. 4) (broncho-alveolar lavage fluid: BALF) 6. 10% FBS/DMEM 1 (trachea), 409
3 3. ACK 37oC % trypan blue. 5) BALF cytospin( ) 1,000 rpm 5 slide. Diff-Quick (Baxter Healthcare, U.S.A.). type, (Nikon, japan) 400 infiltration. 6) ELISA analysis IL-4, IL-5, IgE, IFN- enzyme-linked immuno-sorbent assay(elisa, Endogen, USA), IL-13 ELISA kit(r&d system). 7) (flow cytometry analysis) 2~5 105, (1% ( ), 0.01% NaN3, ph 7.4) 1. FITC(flu-ores-ceinat-ediso thiocyanate) PE(phycoerythrin) CD3, CD4, CD8, CD69, CCR3, B (FACScan, BD Biosciences). (No. of 400 photo) , , GLU GLU (p<0.001) (Fig. 1). Fig. 1. Effect of GLU extract drug on eosinophil in BALF of OVA-induced asthm a murine. group data by T test ( p < 0.01, p < 0.01) Fig. 2. Effect of GLU extract drug on IL-4level in murine OVA - indued asthma of BAL fluid. group data by T test ( p < 0.05, p < 0.01, p < 0.001) 410
4 Fig. 3. Effect of GLU extract drug on IFN- level in murine OVA - indued asthma of BAL fluid. group data by T test ( p < 0.05, p < 0.01, p < 0.001) Fig. 4. Effect of GLU extract drug on IL-5 level in murine OVA - indued asthma of BAL fluid. group data by T test ( p < 0.05, p < 0.01, p < 0.001) Fig. 5. Effect of GLU extract drug on IL-13level in murine OVA - indued asthma of BAL fluid. group data by T test ( p < 0.05, p < 0.01, p < 0.001) Fig. 6. Effect of GLU extract drug on IgE level in murine OVA - indued asthma of BAL fluid. group data by T test ( p < 0.05, p < 0.01, p < 0.001) IL-4 (pg/ ), , , GLU GLU (p<0.001) (Fig. 2). IFN- (pg/ ), , , GLU GLU (p<0.05) (Fig. 3). IL-5 (pg/ ), 0 0, , GLU GLU 411
5 Granulocytes / lymphocytes population SSC Granulocytes (6.7%) Lymphocytes (55.7%) % 30.1% % 31.9% FSC Fig. 7. Effect of GLU extract drug on Graanulocytes / lym phoocytes population (%) in murine OVAA-indued asthma lung cells. WT : Nomal C57 BL/6 mice CD3 / CCR3 Wiid Type OVA-Control OVA-GLU CD3e CCR Fig. 8. Effect of GLU extract drug on CD 3e/CCR3( ) murine OVA- indued asthma lung cells. CD4 / CD8 Wiid Type OVA-Control OVA-GLU CD CD Fig. 9. Effect of GLU extract drug on CD4/CD8( ) murine OVA- indued asthma lung cells. 412
6 CD3e / CD69 CD69 Wiid Type OVA-Control OVA-GLU CD3e Fig. 10. Effect of GLU extract drug on CD3e/CD69 murine OVA- indued asthma lung cells. lge / B220 Wiid Type OVA-Control OVA-GLU lge B220 Fig. 11. Effect of GLU extractdrug on IgE/B220( ) murine OVA- indued asthmalung cells. (p<0.05) (Fig. 4). IL-13 (pg/ ), , , GLU GLU (p<0.05) (Fig. 5). IgE (ng/ ), 0 0, , GLU GLU (p<0.05) (Fig. 6). FACS, / (%) 6.7/55.7, 17.8/30.1, GLU 10.7/31.9 (Fig. 7). CD3e/CCR3(%) 17.8/19.9, 13.9/34.8, GLU 18.5/21.9 CCR3 (Fig. 8). 413
7 CD4/CD8(%) 19.9/8.8, 29.6/12.5, GLU 22.9/11.5 CD4 (Fig. 9). CD69/CD3e(%) 35.7/4.4, 16.3/28.3, GLU 11.1/31.5 CD69 (Fig. 10). IgE/B220(%) 1.2/36.4, 15.7/16.9, GLU 8.9/35.0 IgE (Fig. 11)., IgE cytokine, 4.,, leukotriens,,,,, 10. T ( Th) cytokine Th1, Th2. Th1 IL-2, IL-12, IFN-,,, Th2 IL-4, IL-5, IL-6, IL-10,, 11. Th1 Th2 Th2 12.,,, 6.,,,,,, 7. 8, Wang 9., IL-4, IL-5, IL-13, IgE, IFN-, CD4, CD8, CD69, CCR3. arylsulphatase, histaminase, phospholipase D,, 13. GLU.(Fig. 1) IL-4 Th2 B, T thymocyte, monocyte, monocyte. B IgE, IgE Fc R. IL IL-4 GLU 414
8 (Fig. 2). IFN- Th1 B Th1 Th2. IL- 4 IL-4 IgE 15. IFN- GLU (Fig. 3). IL-5 IL-4 B CD4+ T. IL-5 IL-2 IL-4 B B IgA. IL IL-5 GLU (Fig. 4). IL-13 Th2 IgE 17. IL-13 GLU (Fig. 5). IgE. IgE IgE 13. IgE GLU (Fig. 6). BALF GLU, IL-4, IL-5, IL-13, IFN- Th2 cytokine Th1 cytokine. 30.1%, GLU 31.9% 17.8%, GLU 10.7% (Fig. 7). T T (T cell recepter : TCR) TCR (major histocompatibility complex : MHC). TCR MHC TCR T CD4, CD8, CD3 CD3 T CD3 Th1 Th2 18. Chemokine cytokine cytokine CCR3 chemokine,, Th2 18. CCR3 34.8% GLU 21.9% (Fig. 8). CD4+ T T Th2-type cytokine CD3+, CE4+, CD4+/CD8+. CD4 CD8 1.5~2:1 19. CD4/CD8(%) 29.6/12.5, GLU 22.9/11.5 (Fig. 9) CD4. CD3 T CD69 B T, 415
9 T, 20. CD69 (%) 16.3 GLU 11.1 (Fig. 10) (Fig. 1). B220 B IgE B 18. IgE/B220(%) 15.7/16.9 GLU 8.9/35.0 (Fig. 11) (Fig. 6) IgE. BALF GLU, IL-4, IL-5, IL-13 IFN-,, CCR3, CD4, IgE, CD69., IL-4, IFN-, IL-5, IL-13, IgE, CCR3, CD4, CD8, CD69, IgE. 1., IL-4, IL-5, IL-13, IgE. 2. IFN-. 3., CCR3, CD4, IgE, CD69. allergy : ; 1990,p.208, ; 42(5): : ; 2002,p , : ; 2002p Lee SI. Prevalences of Symptoms of Asthma and Other Allergic Diseases in Korean Children: A Nationwide Questionnaire Survey. J Korea Med Sci. 2001;16(2): : ; 1991,p : ; 1999,p selectin ;18(1): Wang H, Chang B, Wang B. The effect of herbal medicine including astragalus membranaceus (fisch) bge, codonpsis pilosula and glycyrrhiza uralensis fisch on airway responsiveness. Zhonghua Jie He He Hu Xi Za Zhi. 1998;21(5): ;46 (1): ,,,,,. cytokines ;42(1): Kasakura S. A role for T helper type 1 and type 2 cytokine in the pathogenesis of various human diseases. Rinsho Byori, Symp. 1998;46(9): ,,,,.... : ; 2002,p
10 14.. IMMUNOLOGY. : ; 2000,p.1,65, 154-6, ,,,,,,,,,. Interferon- Gamma ;44(4): Greenfeder S, Umland SP, Cuss FM, Chapman RW, Egan RW. Th2 cytokines and asthma. The role of interleukin-5 in allergic eosinophilic disease. Respir Res. 2001;2(2): ,,. IL-10 IL-13 mrna ;9(1): Allison JP, Lanier LL. Structure, function and serology of the T-cell antigen receptor complex. Annu Rev Immunol. 1987;5: Krug N, Erpenbeck VJ, Balke K, Petschallies J, Tschernig T, Hohlfeld JM, Fabel H. Cytokine profile of bronchoalveolar lavage-derived CD4(+), CD8(+), and gammadelta T cells in people with asthma after segmental allergen challenge. Am J Respir Cell Mol Biol. 2001;25(1): Hartnell A, Robinson DS, Kay AB, Wardlaw AJ. CD69 is expressed by human eosinophils activated in vivo in asthma and in vitro by cytokines. Immunology. 1993;80(2):
Abstract. IgE. IgE Th2. x x IL-4 IL-5 IgE CD4 +
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