본문 바로가기
HOME> 논문 > 논문 검색상세

논문 상세정보

NC/Nga 생쥐에서 분리한 T 세포에서 foxp3+ 세포 활성화에 대한 PGA-LM의 효과
Effects of PGA-LM on CD4+CD25+foxp3+ Treg Cell Activation in Isolated CD4+ T Cells in NC/Nga Mice

장순남    (건국대학교 대학원 생물공학과   ); 김금란    (건국대학교 대학원 생물공학과   ); 강상모    (건국대학교 대학원 생물공학과  );
  • 초록

    $\gamma$ -PGA는 우리 전통 콩 발효식품인 청국장의 끈적끈적한 점액성의 성분으로, 매우 다양한 기능을 가지고 있는 천연 소재이다. 이러한 $\gamma$ -PGA가 아토피발진 억제 가능성을 알아보기 위해 NC/Nga 생쥐를 사용하여 in vitro 실험을 실시하였다. $\gamma$ -PGA(PGA-HM, 분자량 300 kDa)를 초음파처리로 저분자화시킨 30 kDa 이하의 저분자 PGA-LM를 만들고, 고분자 PGA-HM과 PGA-LM을 사용하여 실험하였는데 동일한 결과를 얻어 PGA-LM 실험결과 중심으로 보고한 것이다. 아토피 피부발진 NC/Nga 생쥐의 비장에서 B 세포와 T세포를 순수 분리하여 항알레르기 작용에 대한 in vitro 실험을 실시하였다. PGA-LM은 hFCs에 대한 세포독성 실험에서 모든 농도에서 세포독성을 나타내지 않았다. PGA-LM이 B 세포 분화 및 활성화에 미치는 영향을 관찰하기 위하여, NC/Nga 생쥐의 비장에서 순수 분리한 B 세포에 anti-CD40/rmIL-4로 자극한 결과, 대조군은 전사인자인 NF- ${\kappa}B$ 의 활성화로 IL- $1\beta$ , IL-6, 그리고 TNF- $\alpha$ mRNA의 발현이 증가되었다. 그러나 PGA-LM과 양성대조군인 rmIL-10 투여군은 염증사이토카인 IL- $1\beta$ , IL-6 그리고 TNF- $\alpha$ mRNA 유전자 발현이 감소하였고, IL-10 mRNA 유전자 발현은 증가하였으나 TGF- $\beta$ mRNA의 유전자 발현은 대조군과 큰 차이가 나타나질 않았다. 또한 CD4+ T 세포에 PGA-LM $100\;{\mu}g/ml$ 를 처리한 후 4일간 동시 배양하여 CD4+IFN- $\gamma$ +와 CD4+CD25+foxp3+ Treg 세포를 세포내 염색으로 분석한 결과에서는 CD4+IFN- $\gamma$ +인 Th1 세포의 증가와 CD4+CD25+foxp3+ Treg 세포를 증가시켜 알레르기반응에서 우위한 Th2 세포에서 Th1 세포로 전환시키는 면역조절 역할을 나타내었다. 이상의 결과로 NC/Nga 생쥐에서 PGA-LM은 염증유전자 발현을 억제시키고 IFN- $\gamma$ +의 증가 및 조절 T 세포의 유도로 아토피피부염의 피부발진을 치료하는 면역조절제로 사용될 수 있을 것으로 생각된다.


    Poly- $\gamma$ -glutamic acid ( $\gamma$ -PGA) was mixed natural flora of Bacillus subtilis, contaminated from cooked soybeans. Also, it was performed to find out the antiallergic activity by using NC/Nga mice, in vitro. The $\gamma$ -PGA (PGA-HM : PGA-high molecular weight), Molecular weight 300 kDa, was decomposed and made PGA-LM (PGA-low molecular weight) which has molecular weight below 30 kDa by sonication. Therefore, it was same result between PGA-HM and PGA-LM, and reported PGA-LM as basic result. We found that PGA-LM contains antiallergic efficacy that inhibit B cells and Th2 cells activation from isolated CD4+T cells in NC/Nga atopic dermatitis model mice, and not show a cytotoxicity in the hFCs. To investigate the effects of these PGA-LM in vitro, isolation of splenic B cell and CD4+ T cells in atopic dermatitis mice were used. To elucidate the role of PGA-LM in anti-CD40+ interleukin-4 (IL-4)-mediated B-cell activation, showed that the capacity of B cells to expression IL- $1\beta$ , IL-6, and TNF- $\alpha$ mRNA down-regulated, and IL-10 mRNA up-regulation by PGA-LM treatment, but it had no effect on TGF- $\beta$ expression. In addition to CD4+IFN- $\gamma$ + and CD4+CD25+foxp3+, the functions of PGA-LM in the development of the CD4+CD25+foxp3+ and CD4+IFN- $\gamma$ +cells, the phenotype and functions of PGA-LM induced CD4+CD25+foxp3+, and CD4+IFN- $\gamma$ +cells in CD4+T cells. These results suggested that PGA-LM could change cytokine production and generate CD4+CD25+foxp3+ Tregs in NC/Nga mice, and may be effective for immunotherapy in patients with AD.


  • 주제어

    $\gamma$-PGA .   CD4+CD25+foxp3+ Treg .   NC/Nga.  

  • 참고문헌 (54)

    1. Abbas, A. K., K. M. Murphy, and A. Sher. 1996. Functional diversity of helper T lymphocytes. Nature. 383: 787-793 
    2. Barnes, P. J. 2008. Immunology of asthma and chronic obstructive pulmonary disease. Nat. Rev. Immunol. 8: 183-192 
    3. Bellanti, J. A. 1998. Cytokines and allergic diseases: clinical aspects. Allergy Asthma Proc. 19: 337-341 
    4. Bellinghausen, I., G. Metz, A. H. Enk, S. Christmann, J. Knop, and J. Saloga. 1997. Insect venom immunotherapy induces interleukin-10 production and a Th2-to-Th1 shift, and changes surface marker expression in venom-allergic subjects. Eur. J. Immunol. 27: 1131-1139 
    5. Bordi, F., C. Cametti, and G. Paradossi. 1996. A comparative study of the high-frequency dielectric properties of poly (alpha-glutamate) and poly (gamma-glutamate) aqueous solutions. Biopolymers 40: 485-494 
    6. Kaneda, N., K. Kobayashi, H. Ichinose, F. Kishi, A. Nakazawa, Y. Kurosawa, K. Fujita, and T. Nagatsu. 1987. Isolation of a novel cDNA clone for human tyrosine hydroxylase: alternative RNA splicing produces four kinds of mRNA from a single gene. Biochem. Biophys. Res. Commun. 146: 971-975 
    7. Kuhn, J. and S. Binder. 2002. RT-PCR analysis of 5' to 3'- end-ligated mRNAs identifies the extremities of cox2 transcripts in pea mitochondria. Nucleic Acids Res. 30: 439-446 
    8. Mosmann, T. R. and R. L. Coffman. 1989. TH1 and TH2 cells: different patterns of lymphokine secretion lead to different functional properties. Annu. Rev. Immunol. 7: 145-173 
    9. Ruiz, R. G., J. F. Price, D. Richards, and D. M. Kemeny. 1990. Lack of relation between IgE in neonatal period and later atopy. Lancet. 336: 808 
    10. Shih, I. L. and Y. T. Van. 2001. The production of poly- (gamma-glutamic acid) from microorganisms and its various applications. Bioresour. Technol. 79: 207-225 
    11. Ting, C. N., M. C. Olson, K. P. Barton, and J. M. Leiden. 1996. Transcription factor GATA-3 is required for development of the T-cell lineage. Nature. 384: 474-478 
    12. Witney, A. A., D. L. Doolan, R. M. Anthony, W. R. Weiss, S. L. Hoffman, and D. J. Carucci. 2001. Determining liver stage parasite burden by real time quantitative PCR as a method for evaluating pre-erythrocytic malaria vaccine efficacy. Mol. Biochem. Parasitol. 118: 233-245 
    13. Elias, K. M., A. Laurence, T. S. Davidson, G. Stephens, Y. Kanno, E. M. Shevach, and J. J. O'Shea. 2008. Retinoic acid inhibits Th17 polarization and enhances FoxP3 expression through a Stat-3/Stat-5 independent signaling pathway. Blood. 111: 1013-1020 
    14. Pandiyan, P., L. Zheng, S. Ishihara, J. Reed, and M. J. Lenardo. 2007. CD4+CD25+Foxp3+ regulatory T cells induce cytokine deprivation-mediated apoptosis of effector CD4+ T cells. Nat. Immunol. 8: 1353-1362 
    15. Szabo, S. J., S. T. Kim, G. L. Costa, X. Zhang, C. G. Fathman, and L. H. Glimcher. 2000. A novel transcription factor, T-bet, directs Th1 lineage commitment. Cell. 100: 655-669 
    16. Punnonen, J., R. de Waal Malefyt, P. van Vlasselaer, J. F. Gauchat, and J. E. de Vries. 1993. IL-10 and viral IL-10 prevent IL-4-induced IgE synthesis by inhibiting the accessory cell function of monocytes. J. Immunol. 151: 1280-1289 
    17. Kanda, N. and Watanabe, S. 2002. Ketoconazole suppresses interleukin-4 plus anti-CD40-induced IgE class switching in surface IgE negative B cells from patients with atopic dermatitis. J. Invest. Dermatol. 119: 590-599 
    18. Lee, J. S., H. Poo, C. J. Kim, Y. H. Choi, C. Park, T. Y. Lee, and M. H. Sung. 2004. Effect of poly-gamma-glutamic acid as an adjuvant and immune response stimulatory factor in vitro and in vivo. The 16th Annual Meeting of the Korean Society for Molecular and Cellular Biology. pp. 14-15 
    19. Gross, S. S., E. A. Jaffe, R. Levi, and R. G. Kilbourn. 1991. Cytokine-activated endothelial cells express an isotype of nitric oxide synthase which is tetrahydrobiopterin-dependent, calmodulin-independent and inhibited by arginine analogs with a rank-order of potency characteristic of activated macrophages. Biochem. Biophys. Res. Commun. 178: 823-829 
    20. Kimata, H., A. Yoshida, C. Ishioka, I. Lindley, and H. Mikawa. 1992. Interleukin 8 (IL-8) selectively inhibits immunoglobulin E production induced by IL-4 in human B cells. J. Exp. Med. 176: 1227-1231 
    21. Robinson, D. S., Q. Hamid, S. Ying, A. Tsicopoulos, J. Barkans, A. M. Bentley, C. Corrigan, S. R. Durham, and A. B. Kay. 1992. Predominant TH2-like bronchoalveolar Tlymphocyte population in atopic asthma. N. Engl. J. Med. 326: 298-304 
    22. Sakaguchi, S. and F. Powrie. 2007. Emerging challenges in regulatory T cell function and biology. Science 317: 627-629 
    23. Leung, D. Y. and T. Bieber. 2003. Atopic dermatitis. Lancet. 361: 151-160 
    24. Miltenyi, S., W. Muller, W. Weichel, and A. Radbruch. 1990. High gradient magnetic cell separation with MACS. Cytometry 11: 231-238 
    25. Zheng, W. and R. A. Flavell. 1997. The transcription factor GATA-3 is necessary and sufficient for Th2 cytokine gene expression in CD4 T cells. Cell 89: 587-596 
    26. Magnan, A. O., L. G. Mely, C. A. Camilla, M. M. Badier, F. A. Montero-Julian, C. M. Guillot, B. B. Casano, S. J. Prato, V. Fert, P. Bongrand, and D. Vervloet. 2000. Assessment of the Th1/Th2 paradigm in whole blood in atopy and asthma. Increased IFN-gamma-producing CD8(+) T cells in asthma. Am. J. Respir. Crit. Care Med. 161: 1790-1796 
    27. Larche, M., D. S. Robinson, and A. B. Kay. 2003. The role of T lymphocytes in the pathogenesis of asthma. J. Allergy Clin. Immunol. 111: 450-463 
    28. Perez-Camero, G., F. Congregado, J. J. Bou, and S. Munoz-Guerra. 1999. Biosynthesis and ultrasonic degradation of bacterial poly(gamma-glutamic acid). Biotechnol. Bioeng. 63: 110-115 
    29. Fontenot, J. D. and A. Y. Rudensky. 2005. A well adapted regulatory contrivance: regulatory T cell development and the forkhead family transcription factor Foxp3. Nat. Immunol. 6: 331-337 
    30. Komai-Koma, M. and P. C. Wilkinson. 1997. TGF-beta stimulates but IFN-gamma inhibits growth-related activation of locomotion of human B cells. J. Immunol. 158: 3125-3129 
    31. Martin, A. and M. Clynes. 1993. Comparison of 5 microplate colorimetric assays for in vitro cytotoxicity testing and cell proliferation assays. Cytotechnology. 11: 49-58 
    32. Obst, M. and A. Steinbuchel. 2004. Microbial Degradation of Poly(amino acid)s. Biomacromolecules 5: 1166-1176 
    33. Heine, G., K. Anton, B. M. Henz, and M. Worm. 2002. 1alpha,25-dihydroxyvitamin D3 inhibits anti-CD40 plus IL-4-mediated IgE production in vitro. Eur. J. Immunol. 32: 3395-3404 
    34. Agnello, D., C. S. Lankford, J. Bream, A. Morinobu, M. Gadina, J. J. O'Shea, and D. M. Frucht. 2003. Cytokines and transcription factors that regulate T helper cell differentiation: new players and new insights. J. Clin. Immunol. 23: 147-161 
    35. Brunkow, M. E., E. W. Jeffery, K. A. Hjerrild, B. Paeper, L. B. Clark, S. A. Yasayko, J. E. Wilkinson, D. Galas, S. F. Ziegler, and F. Ramsdell. 2001. Disruption of a new forkhead/winged-helix protein, scurfin, results in the fatal lymphoproliferative disorder of the scurfy mouse. Nat. Genet. 27: 68-73 
    36. Fiorentino, D. F., M. W. Bond, and T. R. Mosmann. 1989. Two types of mouse T helper cell. IV. Th2 clones secrete a factor that inhibits cytokine production by Th1 clones. J. Exp. Med. 170: 2081-2095 
    37. Wittig, H. J., J. Belloit, I. De Fillippi, and G. Royal. 1980. Age-related serum immunoglobulin E levels in healthy subjects and in patients with allergic disease. J. Allergy Clin. Immunol. 66: 305-313 
    38. Hori, S., T. Nomura, and S. Sakaguchi. 2003. Control of regulatory T cell development by the transcription factor Foxp3. Science. 299: 1057-1061 
    39. Vercelli, D., H. H. Jabara, K. Arai, T. Yokota, and R. S. Geha. 1989. Endogenous interleukin 6 plays an obligatory role in interleukin 4-dependent human IgE synthesis. Eur. J. Immunol. 19: 1419-1424 
    40. Fontenot, J. D., M. A. Gavin, and A. Y. Rudensky. 2003. Foxp3 programs the development and function of CD4+ CD25+ regulatory T cells. Nat. Immunol. 4: 330-336 
    41. Szabo, S. J., B. M. Sullivan, C. Stemmann, A. R. Satoskar, B. P. Sleckman, and L. H. Glimcher. 2002. Distinct effects of T-bet in TH1 lineage commitment and IFN-gamma production in CD4 and CD8 T cells. Science. 295: 338-342 
    42. Hahm, J. H., T. Y. Lee, J. S. Lee, C. Park, M. H. Sung, and H. Poo. 2004. Antitumor effect of Poly-glutamic acid by modulating cytokine production and NK cell activity. International Meeting of the Federation of Korean Microbiological Societies p. 21-22 
    43. Choi, H. J. and M. Kunioka. 1995. Preparation conditions and swelling equilibria of hydrogel prepared by [gamma]- irradiation from microbial poly([gamma]-glutamic acid). Radiation Physics and Chemistry 46: 175-179 
    44. Mosmann, T. R., H. Cherwinski, M. W. Bond, M. A. Giedlin, and R. L. Coffman. 1986. Two types of murine helper T cell clone. I. Definition according to profiles of lymphokine activities and secreted proteins. J. Immunol. 136: 2348-2357 
    45. Sakaguchi, S. 2005. Naturally arising Foxp3-expressing CD25+CD4+ regulatory T cells in immunological tolerance to self and non-self. Nat. Immunol. 6: 345-352 
    46. Worm, M., J. M. Krah, R. A. Manz, and B. M. Henz. 1998. Retinoic acid inhibits CD40 + interleukin-4-mediated IgE production in vitro. Blood. 92: 1713-1720 
    47. Curotto de Lafaille, M. A., N. Kutchukhidze, S. Shen, Y. Ding, H. Yee, and J. J. Lafaille. 2008. Adaptive Foxp3+ regulatory T cell-dependent and -independent control of allergic inflammation. Immunity. 29: 114-126 
    48. Kiniwa, M., M. Gately, U. Gubler, R. Chizzonite, C. Fargeas, and G. Delespesse. 1992. Recombinant interleukin-12 suppresses the synthesis of immunoglobulin E by interleukin-4 stimulated human lymphocytes. J. Clin. Invest. 90: 262-266 
    49. Leung, D. Y. 2000. Atopic dermatitis: new insights and opportunities for therapeutic intervention. J. Allergy Clin. Immunol. 105: 860-876 
    50. Cooper, K. D. 1994. Atopic dermatitis: recent trends in pathogenesis and therapy. J. Invest. Dermatol. 102: 128-137 
    51. Gurbay, A., C. Garrel, M. Osman, M. J. Richard, A. Favier, and F. Hincal. 2002. Cytotoxicity in ciprofloxacin-treated human fibroblast cells and protection by vitamin E. Hum Exp Toxicol 21: 635-641 
    52. Jabara, H. H., S. M. Fu, R. S. Geha, and D. Vercelli. 1990. CD40 and IgE: synergism between anti-CD40 monoclonal antibody and interleukin 4 in the induction of IgE synthesis by highly purified human B cells. J. Exp. Med. 172: 1861-1864 
    53. Kang, H. S., M. J. Lee, H. Song, S. H. Han, Y. M. Kim, J. Y. Im, and I. Choi. 2001. Molecular identification of IgEdependent histamine-releasing factor as a B cell growth factor. J. Immunol. 166: 6545-6554 
    54. Skehan, P., R. Storeng, D. Scudiero, A. Monks, J. McMahon, D. Vistica, J. T. Warren, H. Bokesch, S. Kenney, and M. R. Boyd. 1990. New colorimetric cytotoxicity assay for anticancer-drug screening. J. Natl. Cancer Inst. 82: 1107-1112 
  • 이 논문을 인용한 문헌 (1)

    1. Jang, Soon-Nam ; Kim, Kum-Lan ; Yun, Mi-Young ; Kang, Sang-Mo 2010. "The Effect of $\gamma$-PGA on NC/Nga Mice, a Mouse Model for Mite Antigen-induced Atopic Dermatitis" 한국미생물·생명공학회지 = Korean journal of microbiology and biotechnology, 38(1): 53~63     

 저자의 다른 논문

  • 장순남 (1)

    1. 2010 "집먼지 진드기 항원으로 아토피 피부염을 유발한 NC/Nga 생쥐에 미치는 $\gamma$-PGA의 효과" 한국미생물·생명공학회지 = Korean journal of microbiology and biotechnology 38 (1): 53~63    
  • 김금란 (3)

  • 강상모 (63)

 활용도 분석

  • 상세보기

    amChart 영역
  • 원문보기

    amChart 영역

원문보기

무료다운로드
유료다운로드

유료 다운로드의 경우 해당 사이트의 정책에 따라 신규 회원가입, 로그인, 유료 구매 등이 필요할 수 있습니다. 해당 사이트에서 발생하는 귀하의 모든 정보활동은 NDSL의 서비스 정책과 무관합니다.

원문복사신청을 하시면, 일부 해외 인쇄학술지의 경우 외국학술지지원센터(FRIC)에서
무료 원문복사 서비스를 제공합니다.

NDSL에서는 해당 원문을 복사서비스하고 있습니다. 위의 원문복사신청 또는 장바구니 담기를 통하여 원문복사서비스 이용이 가능합니다.

이 논문과 함께 출판된 논문 + 더보기