藤黄微球菌Rpf结构域多肽诱导小鼠免疫应答的实验研究

Immune response induced by Rpf domain from Micrococcus luteus in mice

目的:研究藤黄微球菌Rpf结构域多肽的免疫学特性。方法:用原核表达的藤黄微球菌Rpf结构域多肽免疫BALB/c小鼠3次,每次间隔2周。用ELISA法检测免疫小鼠血清中特异性抗体滴度。分离免疫小鼠的脾淋巴细胞,体外用抗原再刺激后,用MTT比色法检测免疫小鼠脾淋巴细胞的增值指数。ELISA方法检测淋巴淋巴细胞悬液中IFN-γ、IL-10和IL-12产生水平。另一部分免疫的小鼠经尾静脉感染MTB毒株H37Rv,4周后,计数脾脏细菌负荷数。结果:藤黄微球菌Rpf结构域多肽免疫小鼠血清特异性抗体滴度为1∶128000,淋巴细胞增殖指数为(2.10±0.12),明显高于生理盐水对照组(0.90±0.21)。ELISA方法检测Rpf结构域多肽免疫组IFN-γ,IL-10和IL-12水平为(1126±36)ng/L,(368±13)ng/L和(289±14)ng/L,明显高于生理盐水对照组(P<0.01)。与生理盐水免疫组(细菌负荷6.64±0.13)相比较,Rpf结构域多肽免疫的BALB/c小鼠,对攻击感染后抗MTB在脾脏中增殖有明显作用(细菌负荷为5.03±0.11,P<0.05)。结论:藤黄微球菌Rpf结构域多肽有可能作为新型结核疫苗的候选组分。

AIM： To investigate the immunobiology of Rpf domain from Micrococcus luteus. METHODS： BALB/c mice were immunized with Rpf domain three times at 2-week interval. ELISA was used to detect the title of the anti-Rpf domain antibody titer in the immunized mice sera. The spleen lymphocytes of the immunized mice were separated and the stimulation index （SI） was measured by MTT colorimetry. The levels of secreted IFN-γ, IL-10 and IL-12 upon specific antigen stimulation was detected by ELISA. The Rpf domain immunized BALB/c mice were intravenously infected with 10^5 CFU MTB H37Rv. The number of CFU in the spleens was determined four weeks after fina injection. RESULTS. The titer of the specific antibody in sera of the immunized BALB/c mice was 1 ： 128 000. The SI of Rpf domain immunized group （2.10 ±0.12） was significantly higher than that of saline immunized group （0.90 ± 0.2] ）. The lever of IFN-γ, IL-10 and IL-12 levels in culture supernatant of spleen lymphocytes from the fusion protein immunized mice was （1 126±36） ng/L, （368±13） ng/L and （289± 14） ng/L, respectively, which was markedly higher than that of saline immunized group （ P 〈 0.01 ）. Compared with normal saline immunized mice （6.64±0.13） four weeks after fina injection, dramatic reduction in MTB replication was observed in the spleen （5.03 ± 0.11 ） from the BALB/c mice immunized with fusion proteins. CONCLUSION： Rpf domain can be used as a candidate for a new TB vaccine.