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《Chinese Journal of Cellular and Molecular Immunology》 2015-01
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Preparation of TAT-Ag85B protein vaccine and evaluation of its anti-Myobacterium tuberculosis effect

ZHANG Rongbo;WANG Wenyang;HU Dong;WANG Wan;XING Yingru;HE Jiang;DAI Jingjing;WU Jing;Department of Immunology and Laboratory Medicine,Medical School,Anhui University of Science and Technology;Institute of Infection and Immunology,Anhui University of Science and Technology;Department of Clinical Laboratory,Affiliated Hospital,Anhui University of Science and Technology;  
Objective To obtain a new Ag85 B protein fused with protein transduction domain(PTD) produced by a HIV-trans-activating transduction domain(TAT-PTD) expression system and investigate its protective effect against Myobacterium tuberculosis as a subunit vaccine.Methods The p ET28a-Ag85 B and p ET28a-TAT-Ag85 B plasmids were established and transformed into E.coli BL21(DE3) strains for recombinant protein expression and purification.Then three groups of BALB/c mice were subcutaneously vaccinated three times with Ag85 B protein,TAT-Ag85 B protein and PBS,respectively.One week after the last immunization,5 mice in each group were sacrificed for detecting serum specific anti-Ag85 B and IFN-γ/IL-2produced by spleen cells using ELISA.Simultaneously,the levels of CD80 and CD86 on macrophages which were stimulated by Ag85 B or TAT-Ag85 B protein were measured using flow cytometry.Subsequently,the rest of the mice were intravenously injected with virulent Myobacterium tuberculosis H37 Rv and their bacterial loads in the lung and spleen were determined 1,2,4 and 8 weeks after infection.Moreover,pulmonary pathological changes were observed by HE staining at 8 weeks after infection.Results Ag85 B and TAT-Ag85 B proteins were obtained successfully.Compared with Ag85 B,higher titers of Ig G antibodies and the levels of IFN-γ and IL-2 were induced by TAT-Ag85 B.Lower bacterial loads in the lung and spleen and smal er scale of pulmonary lesion were found in mice immunized with TAT-Ag85 B than those in Ag85B-treated mice.In addition,TAT-Ag85 B stimulated higher CD80 and CD86 expressions on macrophages.Conclusion TAT-Ag85 B protein is an efficient vaccine that induces a strong Th1 immune response and provides a good protection against Myobacterium tuberculosis infection.The mechanism of the subunit vaccine may be partial y explained as the enhanced capability of antigen-presentation of macrophages.
【Fund】: 国家自然科学基金(81302524 81202294 81172778);; 安徽省自然科学基金(1208085QH162 KJ2013A105 2013SQRL029ZD)
【CateGory Index】: R392
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