结核分枝杆菌融合蛋白亚单位疫苗对BCG初始免疫的加强效应及保护效力  

Protective Efficacy of Mycobacterium tuberculosis Fusion Protein Subunit Vaccine as A Booster to Enhance BCG Primed Immunity

作　　者：李青[1] 傅林锋[2] 王秉翔[2] 姜雯雯[3] 于红娟[3] 雒彧[1] 张颖[3,4] 祝秉东[3] 

机构地区：[1]兰州大学结核病研究中心暨病理生理学研究所,兰州730000 [2]兰州生物制品研究所,兰州730046 [3]兰州大学结核病研究中心暨病原生物学研究所,兰州730000 [4]美国霍普金斯大学布隆博格公共卫生学院分子微生物学与免疫学系

出　　处：《中国生物制品学杂志》2010年第3期280-285,共6页Chinese Journal of Biologicals

基　　金：国家科技重大专项(2008ZX-10003-01305);国家科技部863项目(2006AA02Z420);教育部新世纪优秀人才支持计划(NCET-07-0403)资助

摘　　要：目的探讨结核分枝杆菌融合蛋白亚单位疫苗对BCG初始免疫的加强效应及保护效力。方法将Ag85B、Ag85B-Mpt64190-198-HspX(AMH)、Ag85B-Mpt64190-198-Mtb8.4(AMM)以及AMH+AMM蛋白分别与佐剂二甲基三十六烷基铵(DDA)和卡介苗多糖核酸(BCG-PSN)混合,制备亚单位疫苗。第0周用BCG皮下免疫C57BL/6小鼠,第8、10周分别用各蛋白疫苗皮下加强免疫,以PBS和BCG(仅免疫1次)作为对照。末次免疫后4周,采血检测血清抗体水平,并分离脾淋巴细胞,检测分泌IFNγ的水平;末次免疫后12周,经尾静脉注射H37Rv株,6周后取肺,进行菌落计数,抗酸及HE染色观察。结果各亚单位疫苗加强组诱导产生的特异性抗Ag85BIgG抗体水平均明显高于BCG组;融合蛋白疫苗加强组免疫小鼠脾细胞经Ag85B和PPD刺激后,产生分泌IFNγ的淋巴细胞数明显多于BCG组;融合蛋白加强组免疫小鼠肺组织结核结节面积与BCG组比较,差异无统计学意义;仅AMM+AMH加强组免疫小鼠肺组织菌落计数明显低于BCG组,其抗酸染色阳性细菌数明显低于PBS、BCG及AMH、Ag85B加强组。结论AMH和AMM疫苗联合加强BCG免疫,能诱导小鼠特异性的细胞免疫和体液免疫应答,具有较强的免疫原性,可增强BCG的保护效力。Objective To investigate the protective efficacy of Mycobacterium tuberculosis fusion protein subunit vaccine as a booster to enhance BCG primed immunity.Methods Mycobacterium tuberculosis fusion protein subunit vaccines were prepared by mixing dimethyl-dioctyldecyl ammonium bromide（DDA）as an adjuvant and BCG polysaccharide nucleic acid（BCG-PSN）with proteins Ag85B,Ag85B-Mpt64190-198-HspX（AMH）,Ag85B-Mpt64190-198-Mtb8.4（AMM）and AMH＋AMM respectively.C57BL/6 mice were immunized s.c.with BCG at week 0,and boosted with various subunit vaccines respectively at weeks 8 and 10,using those immunized with one dose of PBS and with one dose of BCG as controls.The serum samples of mice were collected 4 weeks after the last immunization and determined for antibody levels.Meanwhile,the splenic lymphocytes of mice were isolated for determination of IFNγ level secreted.The mice were challenged i.v.with H37Rv strain 12 weeks after the last immunization,and their lungs were taken out 6 weeks after challenge for bacterial colony counting as well as acid-fast staining and HE staining.Results The anti-Ag85B IgG levels induced by various subunit vaccines were significantly higher than that by BCG.After stimulation with Ag85B and PPD,the counts of IFNγ-secreting lymphocytes of mice boosted with various subunit vaccines were significantly higher than those receiving only primary immunization with BCG,while the areas of tubercle showed no significant difference.Only the bacterial colony counts in lung tissues of mice boosted with AMM＋AMH were significantly lower than those of mice receiving only primary immunization with BCG.However,the counts of cells positive in acid-fast staining of mice boosted with AMM＋AMH were significantly lower than those with AMH and with Ag85B or receiving only primary immunization with PBS and with BCG.Conclusion The booster immunization with AMH＋AMM after primary immunization with BCG induced specific cellular and humoral immune responses in mice,which showed strong immunogenicity and

关 键 词：结核分枝杆菌 融合蛋白 亚单位疫苗 初始-强化免疫策略 

分 类 号：R378.911[医药卫生—病原生物学] R392.33[医药卫生—基础医学]