함초에서 분리한 다당체에 의한 큰포식세포 활성화
Activation of Macrophage by Polysaccharide Isolated from Salicornia herbacea L.
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함초 (Salicornia herbacea L.)는 국내의 해안가에서 자라는 식물로 생리활성기능에 대해서는 잘 알려져 있지 않지만, 면역계통을 활성화 시키는 기능이 있음이 보고되었다. 이 연구에서 함초에서 분리한 다당체가 비특 이적 면역반응에 관여하는 큰포식세포를 활성화 시킴을 확인하였다. 함초가 큰포식세포를 활성화시키는 기전을 밝혀내기 위하여 inducible nitric oxide synthase (iNOS)를 이용한 큰 포식세포 면역형광염색을, iNOS mRNA의 변화를 보기 위한 역전사효소-중합효소연쇄반응 (reverse transcriptasepolymerase chain reaction :RT-PCR)을 실시하였다. 함초에서 추출한 다당체(Salicornia polysaccharide, SPS)를 생쥐 큰포식세포주인 RAW 264.7 세포에 24시간 처 리한 결과, 용량의존적으로 일산화질소 생성을 증가 시켰다. iNOS에 면역형광염색을 실시한 결과, 세포질에서 소 량 발현되던 iNOS 단백질이 SPS를 처리한 후에는 다량 발현됨을 알 수 있었다. 또한 RT-PCR을 실시한 결과 SPS를 처리한 RAW264.7세포에서 iNOS mRNA의 생성이 용량의존적으로 증가함을 알 수 있었다. iNOS의 유전 자 발현에 중요한 역할을 하는 nuclear factor-κB/Rel (NF-κB/Rel)의 활성을 SPS가 현저하게 증가시킴을 확인하였다. 결론적으로 함초에서 분리한 다당체의 작용은 iNOS 유전자의 발현을 통하여 일산화질소의 생성을 증가시키며, 이러한 큰포식세포의 활성화는 이 다당체를 면역증강제로 활용할 가능성을 시사해 준다.
Abstract
Salicornia herbacea L. is one of the halophytes that can grow in salt marshes, or salt fields along the seashores in Korea. The objective of this study is to investigate the mechanism by which Salicornia Polysaccharide, (SPS) activates macrophages. To analyze macrophage activation and iNOS gene expression, we performed nitrite generation assay, immunohistochemistry, and reverse transcriptase-polymerase chain reaction. A polysaccharide isolated from the Salicornia herbacea L. significantly induces nitric oxide (NO). Immunohistochemical staining of inducible NO synthase (iNOS) showed that the increase of NO was due to the induction of iNOS production. RT-PCR analysis showed that SPS produced significant induction of iNOS gene expression. Immunohistochemical staining of p65 showed that SPS produced strong induction of NF-κB/Rel nuclear translocation. Electrophoretic mobility shift assay further confirmed the activation of NF-κB/Rel by SPS. In conclusion, we demonstrate that SPS stimulates the macrophages to express iNOS gene via the activation of NF-κB/Rel.
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