Aplicação da apirase de batata e de polipeptídeo recombinante derivado no estudo da ATPDase 2 de Schistosoma mansoni e da esquistossomose mansoni

Abstract

A polypeptide belonging to the conserved domain B (r78-117) from the potato apyrase, homologue to the domain B (r156-195) from the S. mansoni ATPDase 2 isoform, was obtained as a 6xHis tag polypeptide (r-potDomain B). Polyclonal anti-r-potDomain B antibodies, when immobilized on Protein A-Sepharose, immunoprecipitated 67-91% of the ATPase and ADPase activities from the adult worm preparation. By Western blots, the immune serum recognized band of 63 and 55 kDa in both parasite preparation and immunoprecipitated complex, confirming the ATPDase 2 identity. Additionally, polyclonal anti-r-potDomain B antibodies inhibited 38-58% of the ATPase and ADPase activities, suggesting that the domain B from the S.mansoni ATPDase 2 is a new target for inhibitor design. Serum samples from patients with schistosomiasis were tested in ELISA using r-potDomain B as coating antigen. The seropositivity of the subclass IgG1 (27%, 8/30), IgG2 (27%, 8/30), IgG3 (50%, 15/30) or IgG4 (17%, 5/30) suggested that the domain B from the ATPDase 2 is potentially involved in the host immune response. The potato apyrase or r-potDomain B, emulsified in Freund Complete and Incomplete adjuvants, was inoculated in Swiss mice, increasing the IgG antibodies production, which were also reactive with soluble egg antigens (SEA). Protective immunity induced by each biomolecule was evaluated 60 days after challenge infection. No worm burden or egg number reduction was observed. Mice immunized with potato apyrase developed high IgG1 antibody levels, reactive with either biomolecules or SEA, while IgG2a antibody levels were produced in lower proportion, suggesting a tendency of a Th2 type immune response. The r-potDomain B was able to induce higher IgG2a antibody production, an immune response possibly associated to the Th1 type. Histological analyses of haematoxylin-eosin-sections obtained from mice immunized with potato apyrase, but not with r-potDomain B, showed a significant reduction in liver granuloma area as evidenced by the cells number reduction and substitution by collagen fibbers. Further studies of these biomolecules could contribute to a better understanding of host-parasite interactions, and may to be explored as new tools in schistosomiasis research.