Avaliação da resposta humoral e celular de camundongos BALB/C imunizados com a proteína rMSP1a obtida do isolado UFMG2 de Anaplasma marginale

Abstract

Bovine Anaplasmosis is a disease caused by the intra-erythrocytic rickettsia Anaplasma marginale, which is biologically transmitted by ticks and mechanically by hematophagous flies. Surface proteins (MSPs) of A. marginale, are important for the interaction of the pathogen with host and constitute potential vaccine targets against this pathogen. MSP1a and MSP2 have greater potential as immunogens, being consideredthe MSP1a a adhesin of bovine erythrocytes and tick cells. Isolated exogenous proteins administered into the body can have low biodistribution or are rapidly degraded or eliminated. The use of carriers would be an alternative for increasing the efficiency of distribution and presentation of the antigen to the immune system. The aim of thepresent study was to evaluate the humoral and cellular immune response of Balb/c mice immunized with the recombinant protein fragment MSP1a, originated from the A. marginale UFMG2 in different combinations. A fragment of 339bp MSP1a comprising the N-terminal region, was cloned into the expression vector pJexpress414. The expression product was purified and identity of the protein was confirmed by SDS-PAGE and Western Blot. Multiwalled carbon nanotubes (MWNTs) were used as a carrier molecule to the rMSP1a as antigen presentation to the immune system. Thirty-six Balb/c mice, aged 6 to 8 weeks, were divided into six groups of six animals each (non-immunized, PBS, rMSP1a, MWNT, MWNT+rMSP1a and complete vaccine (antigen of A. marginale cultivated in vitro + MWNT+rMSP1a) and immunized withthree doses, in 21 days intervals. Mice immunized with the protein rMSP1a, MWNT+rMSP1a and vaccine produced high levels of IgG anti-rMSP1a, showing that the binding of rMSP1a in MWNT did not interfere in its original conformation. However, the complete vaccine group presented antibody titers lower than groups rMSP1a and MWNT+rMSP1a. No response was detected response in the groupimmunized with free MWNT. Splenocytes of group MWNT+rMSP1a significantly proliferated upon stimulation with the protein and presented a higher percentage of CD4+/CD44 + and CD4 +/CD62 +. Significant levels of TNF- were observed in the group MWNT+rMSP1a compared to PBS group, and the highest concentration of IL-10 was observed in serum of the animals tested complete vaccine. These results suggestthat rMSP1a associated with MWNT may influence both the increased memory response as lymphocyte activation.