新型结核病疫苗AEH/Al/IC在小鼠中的免疫原性研究

Immunogenicity of a new TB vaccines AEH with aluminum hydroxide and poly(I:C) in mice

目的检测重组结核分枝杆菌(Mycobacterium tuberculosis,Mtb)融合蛋白Ag85B-ESAT6(Antigen 85 B,6-kDa early secretory antigenic target,AE)和热休克蛋白X(heat shock protein X,HspX)与氢氧化铝和聚肌胞苷酸(Polyinosinic-polycytidylic acid,poly I∶C)构建的新型结核病亚单位疫苗AEH/Al/IC在小鼠中的免疫原性。方法用疫苗(AEH/Al/IC)和佐剂(Al/Poly I∶C)分别免疫雌性BALB/c小鼠3剂次,每剂次间隔10 d。末次免疫后第10天,经ELISA检测小鼠血清中抗原特异性IgG、IgG1和IgG2a的抗体效价,经酶联免疫斑点试验(enzyme-linked immunospot,ELISPOT)检测小鼠脾细胞分泌抗原特异性IFN-γ和IL-2的细胞频数,经ELISA检测小鼠脾细胞抗原特异性IFN-γ、IL-2和TNF-α的分泌量。结果与佐剂组相比,疫苗组诱导小鼠产生了高水平的AE和HspX特异性抗体;疫苗组诱导小鼠产生的AE和HspX特异性IFN-γ和IL-2阳性脾淋巴细胞(斑点形成细胞spot forming cells,SFC)均高于佐剂组,差异有统计学意义(IFN-γ:P<0.05;IL-2:P<0.05);疫苗组小鼠脾细胞AE特异性IFN-γ、IL-2和TNF-α分泌量明显较佐剂组高,差异均有统计学意义(P<0.05),HspX特异性IFN-γ和TNF-α分泌量比佐剂组稍高,但差异均无统计学意义(P>0.05)。结论结核病亚单位疫苗AEH/Al/IC具有良好的免疫原性,有希望成为一种新型的结核病预防候选疫苗。

Objective To evaluate the immunogenicity of a novel subunit vaccine AEH,which is comprised of recombinant Mycobacterium tuberculosis fusion protein Ag85B-ESAT6(Antigen 85 B,6-kDa early secretory antigenic target,AE)and HspX(heat shock protein X),in conjunction with aluminum hydroxide(Al)and poly I∶C in mice.Methods Female BALB/c mice were immunized with the vaccine(AEH/Al/IC)or Al/poly I∶C for 3 doses 3 days apart.Serum samples were collected 10 d after the last injection and determined for IgG/IgG1/IgG2a titer by indirect ELISA,the number of spleen cells secreted IFN-γand IL-2 was detected by ELISPOT,the level of IFN-γ,IL-2 and TNF-αin spleen cells was detected by ELISA.Results The detection of antibody showed that the levels of anti-AE and anti-HspX IgG,IgG1 and IgG2a in vaccine group were higher than the adjuvant group.ELISPOT assay showed that the AE and HspX-specific IFN-γand IL-2 spot forming cells(SFC)produced by the spleen lymphocytes of the vaccine group were significant higher than the adjuvant group(IFN-γ:P<0.05,IL-2:P<0.05).The results of cytokine ELISA showed that the levels of AE-specific IFN-γ,IL-2 and TNF-αproduced by spleen cells of the vaccine group were significantly higher than the adjuvant group(P<0.05);HspX-specific IFN-γand TNF-αproduced by spleen cells of the vaccine group were slightly higher than the adjuvant group,but the difference was not statistically significant(P>0.05).Conclusion The data indicated that the AEH/Al/IC was highly immunogenic in mice,it may represent a promising candidate vaccine for the control of TB.