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目的观察不同浓度重组刚地弓形虫抑制蛋白(r Tg PRF)滴鼻免疫小鼠诱导的脾淋巴细胞免疫应答,探讨其最佳免疫剂量。方法 50只雌性BALB/c小鼠随机分为5组,免疫组分别用10、20、30μg和40μg r Tg PRF溶于20μl磷酸盐缓冲液(PBS)后滴鼻免疫,对照组用20μl PBS。分别于第0、14和21天滴鼻免疫小鼠,末次免疫后2周,颈椎脱臼处死小鼠。无菌取脾,采用CCK-8法测定脾淋巴细胞刺激指数(SI),酶联免疫吸附试验(ELISA)测定脾淋巴细胞上清液中IFN-γ、IL-2、IL-4和IL-10浓度。结果经r Tg PRF刺激,30μg和40μg免疫组小鼠脾淋巴细胞SI均高于对照组(P均<0.05),且30μg组显著高于40μg组(P<0.05)。与对照组比较,所有免疫组小鼠脾淋巴细胞培养上清中IFN-γ含量升高(P均<0.05),20、30μg和40μg组小鼠脾淋巴细胞培养上清中IL-2含量增加(P均<0.05),其中30μg组小鼠脾淋巴细胞培养上清中IFN-γ(P均<0.01)和IL-2含量最高(P均<0.01);所有免疫组小鼠脾淋巴细胞培养上清中IL-4和IL-10含量与对照组比较差异均无统计学意义(P均>0.05)。结论 r Tg PRF滴鼻免疫小鼠可诱导脾淋巴细胞增殖和Th1型细胞免疫应答,以30μg剂量免疫效果最佳。
Objective To observe the immune response of splenic lymphocytes induced by intranasal immunization of mice with different concentrations of recombinant T.gondii gondii receptor protein (r Tg PRF) and to explore the best immunization dose. Methods Fifty female BALB / c mice were randomly divided into five groups. The immunized groups were intranasally immunized with 10, 20, 30μg and 40μg r Tg PRF respectively in 20μl phosphate buffered saline (PBS). The control group was immunized with 20μl PBS. Mice were immunized intranasally on days 0, 14 and 21, respectively. Two weeks after the last immunization, mice were sacrificed by cervical dislocation. The spleen lymphocyte stimulating index (SI) was determined by CCK-8 assay and the levels of IFN-γ, IL-2, IL-4 and IL- 10 concentration. Results The SI of spleen lymphocytes in 30μg and 40μg immunized mice was significantly higher than that in control group (all P <0.05), and was significantly higher in 30μg and 40μg groups (P <0.05). Compared with the control group, the content of IFN-γ in the spleen lymphocyte culture supernatant of all the immunized mice increased (P <0.05), while the levels of IL-2 in the supernatant of 20, 30, and 40μg spleen lymphocytes increased (All P <0.05). The levels of IFN-γ (P <0.01) and IL-2 in the culture supernatant of the spleen lymphocytes of 30μg group were the highest (all P <0.01) The content of IL-4 and IL-10 in the supernatant was not significantly different from the control group (all P> 0.05). Conclusion The mice immunized with r Tg PRF intranasally can induce spleen lymphocyte proliferation and Th1-type cellular immune response, and the immunization with 30 μg dose is the best.