RESUMO
The aim of this study was to test the protective immunity against Toxoplasma gondii in mice induced by a chimeric protein rSAG1/2. The PCR-amplified SAG2 fragment (558 bp) digested with the restriction enzyme XhoI was inserted into the XhoI site of plasmid pGEX-6p-1, termed pGexSAG2. The PCR-amplified SAG1 fragment (1,008 bp) digested with restriction enzymes EcoRI and XhoI was cloned into the EcoRI/ XhoI sites of a separate plasmid pGEX-6p-1, termed pGexSAG1. The SAG2 fragment (HindIII/HindIII) excised from pGexSAG2 was inserted into the HindIII site of pGexSAG1 and a chimeric vector constructed, pGexSAG1/2. The fusion proteins GST-SAG1/2, GST-SAG1 and GST-SAG2 were expressed in BL21 Star (DE3) Escherichia coli and purified by GSTrap FF columns. After removing the GST tag, the recombinant proteins rSAG1/2, rSAG1 and rSAG2 were independently collected and injected into different groups of mice to evaluate their protective capability. The highest proliferation of splenocytes stimulated with tachyzoite sonicate antigen (TsoAg) was observed in BALB/c mice which had received two intraperitoneal injections of rSAG1/2. Maximum production of IFN-gamma was also found in the culture supernatants of TsoAg-stimulated splenocytes from rSAG1/2-immunized mice. Finally, 73% (11/15) of mice immunized with rSAG1/2 survived at least 28 days after a lethal challenge of 1 x 10(3) live tachyzoites which killed all non-immunized mice within 10 days. Moderate survival rates were observed in mice immunized with either rSAG1 (60%) or rSAG2 (53%). These results show that the chimeric protein rSAG1/2 can elicit a Th1-associated protection against T. gondii infections in mice.
Assuntos
Antígenos de Protozoários/imunologia , Proteínas de Protozoários/imunologia , Células Th1/imunologia , Toxoplasma/imunologia , Toxoplasmose Animal/prevenção & controle , Sequência de Aminoácidos , Animais , Antígenos de Protozoários/química , Antígenos de Protozoários/genética , Antígenos de Protozoários/metabolismo , Sequência de Bases , Escherichia coli/genética , Escherichia coli/metabolismo , Feminino , Glutationa Transferase/genética , Glutationa Transferase/metabolismo , Imunização , Interferon gama/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Proteínas de Protozoários/química , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Vacinas Protozoárias/administração & dosagem , Vacinas Protozoárias/imunologia , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/metabolismo , Toxoplasma/genética , Toxoplasma/patogenicidade , Toxoplasmose Animal/imunologia , Toxoplasmose Animal/mortalidade , Toxoplasmose Animal/parasitologiaRESUMO
A monoclonal antibody TGCM5 (isotype G, subclass 1, kappa light chain) against SAG2, a major surface antigen of Toxoplasma gondii tachyzoites, was produced in quantity and its Fab-containing idiotype (Id) was injected into rabbits for production of rabbit anti-Id sera. The rabbit anti-Id IgG (aId-IgG) with a SAG2 internal image was obtained by purification with a protein A column. Significant lymphocyte proliferation induced by tachyzoite lysate antigen was observed in BALB/c mice that received two injections of aId-IgG, but not in non-immunized control mice. Large amounts of gamma interferon were detected in supernatants of splenocyte cultures prepared from mice immunized with aId-IgG. Of the mice immunized with aId-IgG, 75-87.5% survived at least 28 days after a lethal challenge of 1 x 10(3) live tachyzoites, which killed all non-immunized control mice within 10 days. The results showed that the SAG2 internal image presented by aId-IgG indeed elicited a protection against the infection of T. gondii in mice.
