Papel de HSP70 de Toxoplasma gondii como antígeno vacinal e como ferramenta diagnóstica de toxoplasmose em camundongos BALB/c e C57BL/6

Abstract

Toxoplasma gondii is an obligate intracellular protozoan parasite of warm-blooded animals that can express the heat shock protein of 70 kDa (TgHSP70) during stage conversion. TgHSP70 is a highly antigenic protein and its presence indicates a danger signal to the host. The aim of this work was to verify the role of TgHSP70 as an alternative tool for active toxoplasmosis diagnosis, detecting the protein, specific antibodies and immune complexes (IC) in the experimental infection with T. gondii. Also, we observed the localization of this protein in T. gondii tachyzoites and its role in immunization procedures. Recombinant TgHSP70 was purified from E. coli and used to immunize chickens and Calomys callosus in order to produce specific antibodies anti-TgHSP70. It was observed by immunofluorescence that TgHSP70 is present in the cytoplasm of tachyzoites and could be associated to the inner layer of plasma membrane. It was also demonstrated by immunoblotting and ELISA that this protein is not secreted by T. gondii. Immunization of BALB/c and C57BL/6 mice only with TgHSP70 did not induce protection to challenge with ME-49 and RH strains of T. gondii after 30 of the last immunization. However, it was observed that spleen cells of non-immunized mice proliferate when stimulated in vitro with TgHSP70, indicating unspecific stimulation effect. Moreover, BALB/c and C57BL/6 mice were infected with T. gondii ME-49 strain and treated with dexamethasone (DXM) for toxoplasmosis reactivation. C57BL/6 mice treated with DXM presented higher parasite load in the brain than treated BALB/c mice. In addition, the treatment reduced the inflammatory changes in the brain of both lineages, although, these lesions were higher in C57BL/6 mice. It was verified by qPCR that infection induces TgHSP70 expression in the brain mainly in C57BL/6 mice, and also increases the specific protein concentration in sera from both lineages, but only leads IC formation in BALB/c mice. Finally, these data suggest that circulating TgHSP70 could be used as diagnostic tool for active infection in susceptible C57BL/6 mice and also that TgHSP70, in the other hand, could be used as adjuvant in immunization procedures.