Construction of Helicobacter pylori Lpp20 DNA vaccine and primary study of its immunocompetence in mice / 中华微生物学和免疫学杂志

ABSTRACT

Objective To construct an eukaryotic expression plasmid PeDNA3.1 (+)/Lpp20 and to detect its expression in HeLa cells, and to observe the humoral and cellular immune responses in C57BL/6 mice induced by the Helicobacter pylori Lpp20 DNA vaccine injected intramuscularly. Methods The Lpp20 gene was amplified by PCR. PCR product was subcloned into the eukaryotic expression vector pcDNA3.1 (+)/ Lpp20, and the recombinant plasmid was transfected into HeLa cells using Liposome. After verifying that the Lpp20 antigen gene could be expressed in HeLa cells. Six weeks old C57BL/6 mice were immunized with pcDNA3.1 (+)/Lpp20 or pcDNA3.1 (+) or PBS buffer intramuscularly at 2-week interval for four times. ELISA was used for the quantitative detection of the specific IgG antibody in the sera of C57BL/6 mice and the cytokine IFN-γ in mice spleen lymphocyte culture medium after stimulating by Lpp20. The proliferation response of spleen cells was detected by MTT assay. The Lpp20 gene in muscle was identified by PCR. Results The significant specific antibody titers were detected by ELISA in DNA vaccine groups and the highest titer was 11024 after 6 weeks. The cytnkine IFN-γ in mice inoculated with pcDNA3.1 (+)/Lpp20 was increased and reached (410.36±56.23) pg/ml. A significant difference was tested between the experiment group and the control group[(25.26±10.85)pg/ml] ,P <0.01. The proliferation response of spleen cells of DNA vaccine group(SI 2.37±0.22) was significantly higher than those of mice injected with pcDNA3.1 (+) (SI1.53+0.47) ,P<0.01. Lpp20 gene could exist constantly in musculature cells of mice. Conclusion The eukaryotic expression recombinant pcDNA3.1 (+)/Lpp20 was successfully constructed. Strong humoral and cellular im-munity can be induced by DNA vaccine of pcDNA3.1(+)/Lpp20 in C57BL/6 mice, which might be helpful for further investigation concerning the immunoprotection of DNA vaccine.