| Schistosomiasis is one kind of tropical disease which harmfulness is next to the malaria. This study researched the link between the special component of schistosoma japonicum antigen and expression of costimulatory molecules in modulating Th1/Th2 polarization in murine schistosomiasis, explained the molecular basis of Th2 immune deviation on the host infected schistosoma japonicum, searched for the key signal molecule modulating host Th1/Th2 polarization, provided a new way for contronling effectively the pathogenesis of the egg granuloma even liver fibrosis and provided the new strategy of the vaccine of schistosomiasis.1. Relation of costimulatory molecules expression profile and pathological changes of egg granuloma during the chronic stage in mice infected Schistosoma japonicum.For this purpose, we observed the associativity between expression of costimulatory molecules on spleen CD4+ T lymphocytes and liver granuloma cells of mice infected with Schistosoma japonicum and egg granuloma in murine liver by FCM at different disease stage. The result shows that dynamic change of the costimulatory molecules expression profile closely related with pathological changes of egg granuloma. High expression of CD80 and 4-1BB followed by the intensity inflammation during the acute stage and the expression level of CD86, ICOS, CD40, OX40 up-regulated with egg granuloma formation. Our data implicated that CD80, 4-1BB participate in modulation of pathological change in the acute stage and high expression of CD86, ICOS, CD40, OX40 companied by according with egg granuloma formation and liver fibrosis. Immunohistochemistry showed the expression level of costimulatory molecules on the liver granuloma enhanced during the infection, suggesting that the expression of costimulatory molecules related with continuous stimuli of egg antigen.2. Effection of costimulatory molecules on polarizing Th1/Th2 responses in mice infected schistosoma japonicumSensitization spleen lymphocytes of mice are harvested and stimulated by SEA, then the cytokines in the culture supernantans are measured by ELISA at different stage of infection. The result shows that Th1 cytokine IFN-γincrease highly in the early stage, however the level of IFN-γdecrease but the level of Th2 cytokines (IL-4,IL-5,IL-10 and IL-13) increase in the chronic stage. The test in vivo demonstrated that the level of IgG and its subtype (IgG1, IgG2a) changed depended on the stage of infection, especially IgG1 up-regulated markedly at 7w, and slightly lower at 12w, the level stayed very high during the chronic stage, at the same time Th2 differentiation index and the ratio of IgG1/IgG2a also increased followed with the disease. These data support that the Th1 immune response reached the high peak at the acute stage, then reduced gradually, but Th2 immune response up-regulated, and the host immune switch to Th2 profile.The change trend of cytokines and serum antibodies were determined by the level of costimulatory molecules on the murine spleen CD4+ T cell. CD80, 4-1BB play a role in Th1 polarization, and ICOS, CD86, CD40, OX40 induced Th2 polarization.3. Effection of functional antigen components of schistosoma japonicum and the expression of costimulatory molecules on Th1/Th2 polarizationDifferent components of soluble Schistosome japonicum egg antigen and adult worm antigen were identified with sera antibody at different disease stage by SDS-PAGE and Western blotting. Results show that the sera of different stage can recognize different antigen components, especially 10 KDa,18 KDa,41 KDa,47 KDa of soluble egg antigen and 13 KDa,37 KDa,57 KDa of adult worm antigen can be recognized by 6w~20w post-infection sera antibody and the issue of immune response is dependent on the disease stage and correlated with the time of schistosome immune deflection, suggesting these protein components are major function antigens and the key to cause immune diversion to Th2 dominance.Murine spleen lymphocytes were collected and stimulated by SEA and Sj16, then the expression of costimulatory molecules on CD4+ T cell were analyzed by flow cytometry and the level of cytokines in culture supernantans were measured by ELISA. The results display high-expression level of Th1 cytokines (IFN-γ) were induced by SEA and Sj16 at 4w and 7w, high-expression level of Th2 cytokines (IL-4, IL-5, IL-10, IL-13) in the chronic disease stage; the level of costimulatory molecules (ICOS, CD86, CD40, OX40) on murine spleen CD4+ T cell up-regulated at 7w, especially the level of ICOS and CD86 elevated sharply and this expression continue high during the chronic stage. The expression level of ICOS and CD86 corresponding with dynamic changes of Th2 cytokines suggests the up-regulating level of ICOS, CD86, CD40, OX40 depend on stimuli of SEA and Sj16. Compared SEA with Sj16, no statistical significant difference were found. Both SEA and Sj16 can lead murine spleen cell to secret Th2 cytokines and switch host immune response to Th2 profile. There were key component antigens in the worm and egg of Schistosome japonicum which can induce ICOS and CD86 to increase and drift immune response to Th2 profile.4. Effection on blocking ICOS, CD86, CD80 costimulatory signals in mice infected Schistosome japonicum on the Th1/Th2 polarization in vitroAnti-ICOSmAb, anti-CD86mAb , anti-CD80mAb antibodies were used to observed the influence of blocking costimulatory signals in the expression level of Th1/Th2 cytokines induced by murine spleen cell infected Schistosoma japonicum at 4w, 7w, 9w. The results shown the other antibodies down regulate IL-4 and IL-10 at 7w and 9w except anti-CD80mAb, indicating blocking ICOS and CD86 costimulatory signals may modulate Th1/Th2 polarization at 7w and 9w. This study suggests that blocking the signals of costimulatory molecules can regulate Th1/Th2 polarization in the Schistosomiasis, further provide a new way to control the formation of schistosoma egg granuloma and prevent liver fibrosis.In conclusion, our data display the dynamic expression characterisation of costimulatory molecules on the muring splenic CD4+ T cell infected schistosoma japonicum are following: CD80, 4-1BB may play a key role in Th1 polarization, whereas ICOS, CD86, CD40, OX40 may play a key role in Th2 polarization; explore the mechanism of costimulatory molecules on regulating the formation of egg granuloma and expression level of costimulatory molecules (ICOS, CD86, CD40, OX40) at post-infection, especially ICOS and CD86, accompanied by Th2 immune deviation and pathogenesis in mice; analysis the relation of Th2 immune deviation and the component antigen of schistosoma, explain the basis of Th2 deviation in the host infected the schistosoma japonicum; found blocking the costimulatory molecule signal of ICOS and CD86 can regulate Th1/Th2 polarization; provide a new way to control effectively the egg granuloma and the new strategy of the vaccine of schistosoma japonicum. |
