Korean Journal of Microbiology (미생물학회지)

The Microbiological Society of Korea (한국미생물학회)
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Effect of Recombinant CagL Immunization on the Gastric Diseases Induced by Helicobacter pylori in Mongolian gerbils

CagL 재조합 단백질 접종후에 Mongolian gerbil에서 나타나는 Helicobacter pylori 감염에 대한 반응

  • Bak, Eun-Jung (Oral Cancer Research Institute, Yonsei University College of Dentistry) ;
  • Jang, Sung-Il (Research Center for Orofacial Hard Tissue Regeneration, College of Dentistry, Yonsei University) ;
  • Choi, Yun-Hui (Research Center for Orofacial Hard Tissue Regeneration, College of Dentistry, Yonsei University) ;
  • Kim, Jin-Moon (Research Center for Orofacial Hard Tissue Regeneration, College of Dentistry, Yonsei University) ;
  • Kim, Ae-Ryun (Research Center for Orofacial Hard Tissue Regeneration, College of Dentistry, Yonsei University) ;
  • Kim, Ji-Hye (Research Center for Orofacial Hard Tissue Regeneration, College of Dentistry, Yonsei University) ;
  • Woo, Gye-Hyeong (Department of Clinical Laboratory Science, Semyung University) ;
  • Yoo, Yun-Jung (Research Center for Orofacial Hard Tissue Regeneration, College of Dentistry, Yonsei University) ;
  • Lee, Sung-Haeng (Department of Cellular and Molecular Medicine, Chosun University) ;
  • Cha, Jeong-Heon (Oral Cancer Research Institute, Yonsei University College of Dentistry)
  • 박은정 (연세대학교 치과대학 구강종양연구소) ;
  • 장성일 (연세대학교 치과대학 악안면 경조직 재생 연구센터) ;
  • 최윤희 (연세대학교 치과대학 악안면 경조직 재생 연구센터) ;
  • 김진문 (연세대학교 치과대학 악안면 경조직 재생 연구센터) ;
  • 김애련 (연세대학교 치과대학 악안면 경조직 재생 연구센터) ;
  • 김지혜 (연세대학교 치과대학 악안면 경조직 재생 연구센터) ;
  • 우계형 (세명대학교 임상병리학과) ;
  • 유윤정 (연세대학교 치과대학 악안면 경조직 재생 연구센터) ;
  • 이성행 (조선대학교 의과대학 세포분자의과학교실) ;
  • 차정헌 (연세대학교 치과대학 구강종양연구소)
  • Received : 2012.05.03
  • Accepted : 2012.05.11
  • Published : 2012.06.30
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Abstract

Helicobacter pylori is an important factor of chronic gastritis, digestive ulcer, and stomach cancer. CagL, a virulence factor of H. pylori, is well-known as a pilus protein which acts as adhesion to host cell and a component of Type 4 secretion system. In this study, we evaluated the protective response of recombinant CagL protein (rCagL) using Mongolian gerbil animal model for H. pylori infection. The cagL gene was cloned from 26695 H. pylori followed by over-expression and purification of the protein in E. coli. Mongolian gerbils were immunized with rCagL protein mixed with aluminum adjuvant via intramuscular injections once a week during 4 weeks. At a week after the last immunization, the Mongolian gerbils were administrated with H. pylori 7.13 strain into the stomach and sacrificed to measure antibody titer on rCagL by ELISA and bacterial colonization in the stomach, and to examine the histopathological changes and cytokine expression at 6 week after challenge. Antibody titers on recombinant protein were significantly increased from a week after the first immunization. There was no significant change of the number of bacterial colony between control group and immunized group. The relative stomach weight was significantly decreased in immunized group, but the significant change of histopathological assessment was not observed in the stomach. Cytokine expression such as IL-$1{\beta}$ and KC also was not significantly different between control and immunized groups. These results indicate that rCagL could effectively induce the formation of the specific IgG antibodies. However, bacterial colonization and histopathological lesions could not be inhibited by the immunization in the stomach, indicating not enough protection against H. pylori infection. We consider that along with CagL other adequate antigens could be needed stimulating immune response and inducing protective effects against gastric disease, and also a better adjuvant could be considered.

Helicobacter pylori는 만성 위염, 소화성 궤양, 위암의 중요한 역학적 인자중 하나이다. H. pylori의 독성인자중 CagL은 숙주 세포와 H. pylori의 제 4형 분비기관(Type 4 secretion system)을 연결하는 adhesin으로 작용하는 섬모 단백질로 H. pylori가 발병하는데 중요한 역할을 하는 것으로 알려져 있다. 이번 연구는 저빌에 H. pylori를 감염시킨 동물 모델을 이용하여 CagL 재조합 단백질을 면역화시켰을 때 나타나는 효과를 평가하였다. 재조합 CagL은 클론되었고, 과발현시켜 정제하여 준비하였다 저빌은 H. pylori 감염 대조군과 H. pylori 감염 CagL 재조합 단백질 접종군으로 분류하였고, 접종시 알루미늄 애쥬번트를 사용하였다. 일주일 간격으로 4회 근육내 접종하였고, 마지막 접종 일주일 후, 모든 저빌에 H. pylori 7.13 균주를 $1{\times}10^9\;bacteria/500{\mu}l$ 농도로 위내 투여하였다. H. pylori 감염 6주째 모든 저빌을 희생하여 혈청 IgG 반응평가를 위한 ELISA를 실시하였고, 위에서는 집락화된 H. pylori의 수평가, 병리조직학적 평가 및 사이토카인 유전자발현을 조사하였다. CagL 재조합 단백질접종 일주일 후부터 H. pylori 감염 CagL 재조합 단백질 접종군의 혈청내 IgG 항체형성이 유의적으로 증가하였다. 위에서의 집락화된 세균수는 두군의 차이가 없었다. 저빌 체중에 대한 위무게 비율는 H. pylori 감염 CagL 재조합 단백질 접종군이 유의적으로 감소하였으나 병리조직학적 평가에서는 유의적인 차이는 확인하지 못하였다. 위에서의 IL-$1{\beta}$와 KC (IL-8 homologues)의 유전자발현 정도도 두 군사이에 유의적인 차이는 없었다. 이번 결과는 CagL 재조합 단백질의 접종은 IgG 항체형성은 효과적으로 자극하였지만 면역화된 숙주에서 세균 집락화의 감소 및 병변형성의 방어까지는 유도하지 못한 것으로 나타났으며, 앞으로 H. pylori 감염에 대해 유효한 면역 반응 및 질병 방어 효과를 나타내기 위해서 CagL을 포함한 다른 종류의 재조합 항원 사용 및 보조적으로 전신 면역 및 점막 면역을 효과적으로 유도하기 위해 안정성있는 애쥬번트의 사용을 고려해야 할 것으로 사료된다.

Keywords

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