重组日本血吸虫四跨膜蛋白第二亲水基团对小鼠免疫保护作用的研究

目的探讨重组日本血吸虫四跨膜蛋白第二亲水基团融合蛋白（GST-TSP2HD）抗血吸虫感染免疫保护效果。方法采用Glutathione sepharose 4B胶亲和层析法大量制备GST-TSP2HD蛋白。实验组以GST-TSP2HD融合蛋白的福氏佐剂抗原免疫C57BL/6J小鼠,同时设置佐剂对照组及GST蛋白免疫对照组,每2周加强免疫1次。加强免疫2次后每只小鼠经腹部皮肤感染（45±2）条血吸虫尾蚴。感染42 d后剖杀小鼠,经生理盐水静脉灌注收集成虫,计数减虫率,用KOH溶液消化肝组织收集虫卵,计算减卵率。组织切片观察小鼠肝组织病理变化,通过酶联免疫吸附试验检测小鼠血清中的抗体水平。结果与佐剂对照组相比,GST-TSP2HD融合蛋白免疫组的减虫率为27.10%,减卵率为32.30%;与GST蛋白免疫组相比,GST-TSP2HD融合蛋白免疫组可获得39.57%的减虫率,43.53%的减卵率。GST-TSP2HD融合蛋白免疫组肝组织中的虫卵肉芽肿数量明显减少,单个虫卵肉芽肿平均直径比佐剂对照组、GST蛋白免疫组分别下降了27.08%（P〈0.05）和40.53%（P〈0.01）,差异均有统计学意义。GST-TSP2HD能诱导高水平抗TSP2HD蛋白的特异性抗体IgG,其中IgG1、IgG2b和IgG2c亚类可能在小鼠抗血吸虫感染保护性免疫中发挥重要作用。结论日本血吸虫四跨膜蛋白是一个有效的日本血吸虫病疫苗候选分子。

Protective effect of recombinant TSP2 hydrophilic domain (TSP2HD) of Schistosoma japonicum in immunized mice

Objective To investigate the protective effect of the second hydrophilic domain of recombinant tetraspanin（TSP2HD） against the infection of Schistosoma japonicum.Methods The purified TSP2HD fusion proteins were prepared largely through affinity chromatography of Glutathione sepharose 4B gel.The C57BL/6J mice in a test group were immunized with recombinant GST-TSP2HD fusion protein mixed with Freund adjuvant,the mice in other two groups were immunized with GST protein mixed with Freund,and Freund alone,respectively,as control,and boosted per two weeks.After the second boosted immune,the mice were challenged with cercariae（45±2） of Schistosoma japonicum.On the 42nd day after the challenge,the mice were sacrificed and the adult worms were recovered by the portal vein perfusion with physiological saline,and the eggs were recovered from the liver tissue digested by potassium hydroxide（KOH）.The worm reduction and egg reduction rates were calculated.The pathological changes were observed by biopsy and H.E.staining.The IgG levels of serum were examined by Enzyme-Linked Immunosorbent Assay（ELISA）.Results Compared with the Freund control group,the worm reduction rate was 27.10% and the egg reduction rate was 32.30% in the test group;compared with the GST immunized group,the worm reduction rate was 39.57% and the egg reduction rate was 43.53% in the test group.The number of granuloma in the liver tissue of the test group reduced significantly,the mean diameter of single granuloma in the test group reduced by 27.08%（P〈0.05） compared with the Freund control group,and reduced by 40.53% compared with the GST immunized group（P〈0.01）.GST-TSP2HD induced a high level of specific IgG antibody against TSP2HD protein,and the subclass of IgG1,IgG2b,IgG2c might play an important role in the protective immune effect of mice against Schistosoma japonicum infection.Conclusion The tetraspanin of Schistosoma japonicum is an effective vaccine candidate against schistosomiasis.