日本血吸虫重组Bb(pGEX-Sj32)疫苗免疫BALB/c小鼠诱导免疫应答的动态研究

Dynamic study on immune responses of BALB/c mice induced by the recombinant Bb(pGEX-Sj32)vaccine against Schistosoma japonicum

目的动态观察日本血吸虫重组Bb（pGEX-Sj32）疫苗免疫BALB/c小鼠后诱导的免疫应答,探讨该疫苗的免疫机制。方法将日本血吸虫重组Bb（pGEX-Sj32）疫苗分别经口服（PO）和滴鼻（IN）途径免疫BALB/c小鼠,在免疫后0~20周,每隔2周每组随机剖杀4只小鼠,取脾并制备脾细胞悬液,用四甲基偶氮唑盐比色法（MTT）检测脾细胞增殖水平、流式细胞仪（FACsort）检测脾CD4＋和CD8＋T淋巴细胞亚群和脾细胞凋亡、双抗体夹心ELISA法检测血清及脾细胞培养上清液中细胞因子IL-10、IL-12和TNF-α水平和血清抗体IgG及其亚类、IgE和IgA的动态变化。结果当SjAWA和ConA刺激或不刺激时,PO组和IN组脾细胞增殖水平均于免疫后6周达最高水平,分别于6周和8周达最高脾细胞凋亡发生率,脾CD4＋T细胞亚群百分比分别在6周和8周达峰值,两组CD8＋T细胞亚群百分比在免疫后2~20周升高不显著,PO组脾细胞因子IL-10、IL-12和TNF-α水平均分别于10周、6周和8周达峰值,IN组分别在10周、8周和8周达峰值,口服组血清抗体IgG、IgG1、IgG2a、IgG2b、IgG3和IgA水平均于8周达峰值,而IN组除IgG和IgA于8周达峰值外,其余均于12周达最高水平,IgE于14周达峰值,两组血清细胞因子IL-10、IL-12和TNF-α水平分别在免疫后（PO组）14、6和8周及（IN组）8、6和8周达最高水平。结论该疫苗可诱导小鼠产生有效的免疫应答。

To observe the dynamic changes of immune responses in mice immunized with recombinant vaccine Bifidobacte- riurn bifidurn （pGEX-Sj32） of Schistosoma japonicurn and study the immune mechanisms of the recombinant vaccine, each mouse in this study was immunized with recombinant Bb （pGEX Sj32） vaccine by per os （PO group） and intranasally （IN group） with recombinant vaccine. During week 0-20 after immunization, splenocytes and eye blood sera of 4 mice from each group were separated at 2-week interval. Enzyme-linked immunosorbent assay （ELISA） was performed to detect the levels of antibodies IgG, IgG1, IgG2a, IgG2b, IgG3, IgE, IgA and cytokine interleukin（IL）-10, IL-12 and tumor necrosis factor （TNF）-a. Splenocyte proliferation was investigated by MTT colorimetric assay, subsets of CD4＋ and CD8＋T cells and apop- tosis of splenocytes by FACsort flow cytometry, respectively. The maximum proliferation efficiencies of both PO and IN group got on week 6 after immunization, while the apoptotic rates appears on 6th week without any stimulation as well as on 8th week with ConA. Rates of CD4＋T and CD8＋ T cells subsets reached to the peak on 6thand 8thweek in PO group after immunization while on 8th and 12th week in IN group, respectively. The level of TNF-α, IL 10 and IL-12 in splenocyte supernatant peaked on the 8t , 10th and 6th week in group PO, while got the maximum on the 8th , 10th and 8th week in group IN, respectively. Except for IgE reached peak on the 14th week, all mice developed sig- nificant peaks of specific IgG, IgG1, IgG2a, IgG2b, IgG3 and IgA ort the 8th week in PO group and those appeared on the 12th week （IgG and IgA peaked on week 8） in group IN, re-spectively, compared to 0 week （P〈0.05 or P〈0.01）. The level of IL 10, IL-12 and TNF-αwhich detected in serum peaked on the 14th , 6th and 8th week respectively in group PO, and got the maximum on the 8th , 6th and 8th week in group IN, respec- tively. The rBb （pGEX-Sj32） vaccine may induce mice to produce immune response effectively.