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Induction of boosted immune response in mice by leptospiral surface proteins expressed in fusion with DnaK.

Atzingen MV, Rodriguez D, Siqueira GH, Leite LC, Nascimento AL - Biomed Res Int (2014)

Bottom Line: Currently available veterinarian vaccines do not induce long-term protection against infection and do not provide cross-protective immunity.Our results demonstrate that leptospiral proteins fused with DnaK have elicited an enhanced immune response in mice when compared to the effect promoted by the individual proteins.The boosted immune effect was demonstrated by the production of total IgG, lymphocyte proliferation, and significant amounts of IL-10 in supernatant of splenocyte cell cultures.

View Article: PubMed Central - PubMed

Affiliation: Centro de Biotecnologia, Instituto Butantan, Avenida Vital Brazil 1500, 05503-900 São Paulo, SP, Brazil.

ABSTRACT
Leptospirosis is an important global disease of human and veterinary concern. Caused by pathogenic Leptospira, the illness was recently classified as an emerging infectious disease. Currently available veterinarian vaccines do not induce long-term protection against infection and do not provide cross-protective immunity. Several studies have suggested the use of DnaK as an antigen in vaccine formulation, due to an exceptional degree of immunogenicity. We focused on four surface proteins: rLIC10368 (Lsa21), rLIC10494, rLIC12690 (Lp95), and rLIC12730, previously shown to be involved in host-pathogen interactions. Our goal was to evaluate the immunogenicity of the proteins genetically fused with DnaK in animal model. The chosen genes were amplified by PCR methodology and cloned into pAE, an E. coli vector. The recombinant proteins were expressed alone or in fusion with DnaK at the N-terminus. Our results demonstrate that leptospiral proteins fused with DnaK have elicited an enhanced immune response in mice when compared to the effect promoted by the individual proteins. The boosted immune effect was demonstrated by the production of total IgG, lymphocyte proliferation, and significant amounts of IL-10 in supernatant of splenocyte cell cultures. We believe that this approach could be employed in vaccines to enhance presentation of antigens of Leptospira to professional immune cells.

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Related in: MedlinePlus

Analysis of IgG isotype profile in serum of mice immunized with the recombinant proteins. Sera from BALB/c mice immunized with the recombinant proteins alone or in fusion with DnaK were analyzed by ELISA. IgG, IgG1, and IgG2a titers were evaluated in each case. Sera from PBS injected mice were employed as negative control. Statistical analyses were performed by two-tailed t-test, comparing the titer obtained with leptospiral surface protein alone with the corresponding DnaK fusion protein.
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fig4: Analysis of IgG isotype profile in serum of mice immunized with the recombinant proteins. Sera from BALB/c mice immunized with the recombinant proteins alone or in fusion with DnaK were analyzed by ELISA. IgG, IgG1, and IgG2a titers were evaluated in each case. Sera from PBS injected mice were employed as negative control. Statistical analyses were performed by two-tailed t-test, comparing the titer obtained with leptospiral surface protein alone with the corresponding DnaK fusion protein.

Mentions: In order to studythe type of immune response triggered in mice by the individual DnaK, Lsa21, rLIC10494, Lp95 C-terminus region, rLIC12730, and the corresponding fusion proteins with DnaK, we performed the ELISA using subclass-specific antibodies IgG1 and IgG2a. The results show that DnaK alone elicits a strong statistically significant IgG immune response, with a predominance of IgG1 (Figure 4), when compared to PBS-immunized control (data not shown). Lsa21 alone was not able to induce an efficient immune response in mice, but the DnaK-Lsa21 protein had an improved, statistically significant production of both IgG total and IgG2a subclass, when compared to Lsa21 alone (Figure 4). Analysis of proteins rLIC10494, Lp95 C-terminus, and rLIC12730 alone and with its DnaK fusion showed an increase, statistically significant titer values of total IgG. The values obtained with IgG subclasses were not statistically significant.


Induction of boosted immune response in mice by leptospiral surface proteins expressed in fusion with DnaK.

Atzingen MV, Rodriguez D, Siqueira GH, Leite LC, Nascimento AL - Biomed Res Int (2014)

Analysis of IgG isotype profile in serum of mice immunized with the recombinant proteins. Sera from BALB/c mice immunized with the recombinant proteins alone or in fusion with DnaK were analyzed by ELISA. IgG, IgG1, and IgG2a titers were evaluated in each case. Sera from PBS injected mice were employed as negative control. Statistical analyses were performed by two-tailed t-test, comparing the titer obtained with leptospiral surface protein alone with the corresponding DnaK fusion protein.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4109591&req=5

fig4: Analysis of IgG isotype profile in serum of mice immunized with the recombinant proteins. Sera from BALB/c mice immunized with the recombinant proteins alone or in fusion with DnaK were analyzed by ELISA. IgG, IgG1, and IgG2a titers were evaluated in each case. Sera from PBS injected mice were employed as negative control. Statistical analyses were performed by two-tailed t-test, comparing the titer obtained with leptospiral surface protein alone with the corresponding DnaK fusion protein.
Mentions: In order to studythe type of immune response triggered in mice by the individual DnaK, Lsa21, rLIC10494, Lp95 C-terminus region, rLIC12730, and the corresponding fusion proteins with DnaK, we performed the ELISA using subclass-specific antibodies IgG1 and IgG2a. The results show that DnaK alone elicits a strong statistically significant IgG immune response, with a predominance of IgG1 (Figure 4), when compared to PBS-immunized control (data not shown). Lsa21 alone was not able to induce an efficient immune response in mice, but the DnaK-Lsa21 protein had an improved, statistically significant production of both IgG total and IgG2a subclass, when compared to Lsa21 alone (Figure 4). Analysis of proteins rLIC10494, Lp95 C-terminus, and rLIC12730 alone and with its DnaK fusion showed an increase, statistically significant titer values of total IgG. The values obtained with IgG subclasses were not statistically significant.

Bottom Line: Currently available veterinarian vaccines do not induce long-term protection against infection and do not provide cross-protective immunity.Our results demonstrate that leptospiral proteins fused with DnaK have elicited an enhanced immune response in mice when compared to the effect promoted by the individual proteins.The boosted immune effect was demonstrated by the production of total IgG, lymphocyte proliferation, and significant amounts of IL-10 in supernatant of splenocyte cell cultures.

View Article: PubMed Central - PubMed

Affiliation: Centro de Biotecnologia, Instituto Butantan, Avenida Vital Brazil 1500, 05503-900 São Paulo, SP, Brazil.

ABSTRACT
Leptospirosis is an important global disease of human and veterinary concern. Caused by pathogenic Leptospira, the illness was recently classified as an emerging infectious disease. Currently available veterinarian vaccines do not induce long-term protection against infection and do not provide cross-protective immunity. Several studies have suggested the use of DnaK as an antigen in vaccine formulation, due to an exceptional degree of immunogenicity. We focused on four surface proteins: rLIC10368 (Lsa21), rLIC10494, rLIC12690 (Lp95), and rLIC12730, previously shown to be involved in host-pathogen interactions. Our goal was to evaluate the immunogenicity of the proteins genetically fused with DnaK in animal model. The chosen genes were amplified by PCR methodology and cloned into pAE, an E. coli vector. The recombinant proteins were expressed alone or in fusion with DnaK at the N-terminus. Our results demonstrate that leptospiral proteins fused with DnaK have elicited an enhanced immune response in mice when compared to the effect promoted by the individual proteins. The boosted immune effect was demonstrated by the production of total IgG, lymphocyte proliferation, and significant amounts of IL-10 in supernatant of splenocyte cell cultures. We believe that this approach could be employed in vaccines to enhance presentation of antigens of Leptospira to professional immune cells.

Show MeSH
Related in: MedlinePlus