结核分枝杆菌联合DNA疫苗初免-BCG加强的免疫效果观察

目的:研究并比较结核分枝杆菌免疫保护性抗原DNA(Ag85A和ESAT-6)疫苗联合免疫,BCG免疫以及联合DNA疫苗初免-BCG加强免疫等不同的免疫策略,诱导免疫应答效果观察.方法:健康雌性BALB/c小鼠24只,随机分成PBS 阴性对照组,DNA初免-BCG异源加强组,DNA(Ag85A和ESAT-6)初免DNA同源加强组和BCG阳性对照组,共进行3次免疫,初免2次,最后1次加强,间隔2周1次.PBS组3次均注射PBS 溶液;DNA/BCG组以质粒DNA免疫2次,最后1次以BCG加强免疫;DNA/DNA组3次均以质粒DNA进行免疫;BCG组则注射PBS溶液2次后以BCG免疫.末次免疫后4、6、8周分别分离血清测定总IgG水平,同时分离小鼠脾细胞,体外经TB-PPD刺激后进行淋巴细胞增殖实验(XTT法)并测定脾细胞培养上清中IFN-γ和IL-4水平.结果:DNA/BCG、DNA/DNA、BCG组体外经TB-PPD刺激后均检测到特异性IgG抗体产生,3组平均效价为1:120、1:160、1:80,DNA/DNA组的抗体效价高于另外2组;小鼠脾细胞体外经TB-PPD刺激后,均能产生特异性淋巴细胞增殖并诱生较强的IFN-γ反应,其中DNA/BCG组IFN-γ的分泌水平高于DNA/DNA组和BCG组(P<0.05).结论:联合DNA疫苗初免-BCG加强的免疫策略能在小鼠体内诱导较强的特异性细胞免疫反应,产生高水平的IFN-γ.

Immunogenicity of DNA vaccine prime-BCG boost against Mycobacterium tuberculosis

AIM:To compare the immunogenicity of vaccine strategies about combined DNA prime-BCG boost and DNA or BCG vaccination only. METHODS: BALB/c mice were divided into four groups each containing 6 mice, and all mice received three immunizations at 2 weeks interval. One group of mice was vaccinated with PBS (PBS group); one group was vaccinated twice at 0 week and 2 weeks with combined DNA expressing two mycobacterial antigens, ESAT-6 and antigen 85A respectively; and then boosted with BCG at 4 weeks (DNA/BCG group); another group was vaccinated twice with combined DNA at 0 week, 2 weeks and 4 weeks(DNA/DNA group); and the final group received PBS twice at 0 week and 2 weeks and then vaccinated with BCG at 4 weeks(BCG group). Specific IgG antibody in serum of mice was determined with indirect ELISA in 4 weeks, 6 weeks, 8 weeks respectively after final vaccination.The splenic lymphocytes of mice were separated and stimulated with PPD to measure their proliferation by XTT, and to evaluate the production of interferon-r(IFN-gamma) and IL-4 in cell suspensions of spleen cells by ELISA. RESULTS: PPD could stimulate specific IgG responses in DNA/BCG, DNA/DNA and BCG groups (DNA/DNA group>DNA/BCG group and BCG group), and the most significant response occurred in 12 weeks; the splenic lymphocyte proliferation reactions and IFN-gamma and IL-4 production was detectable in DNA/BCG, DNA/DNA and BCG groups and DNA/BCG group induced significant higher production, especially in 12 weeks. CONCLUSION: Priming with the combined DNA vaccines and boost with attenuated M. bovis vaccine (BCG) could enhance specific cell reactions and high level IFN-gamma compared with DNA or attenuated M. bovis vaccine alone.