RESUMO
T helper cell-recognized epitopes were determined in chitinase of Onchocerca volvulus, a vaccine candidate protein. The proliferation of splenic T cells of mice immunized with recombinant protein was tested with a library of chitinase-peptides of 16 amino acids with termini overlapping by 12 amino acids, and a library of "designer peptides", i.e. sequences identified with three epitope-predicting algorithms. Fourteen epitope-bearing stretches were identified with the peptides of the overlapping library. Testing of the designer peptides partially confirmed these data and revealed additional epitopes. Five clusters of epitopes were identified for the creation of peptide or minigene DNA vaccines with good potency and potential range of MHC allele presentation.
Assuntos
Quitinases/química , Quitinases/imunologia , Epitopos de Linfócito T/imunologia , Onchocerca volvulus/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Sequência de Aminoácidos , Animais , Antígenos de Helmintos/biossíntese , Antígenos de Helmintos/química , Antígenos de Helmintos/genética , Antígenos de Helmintos/imunologia , Quitinases/biossíntese , Quitinases/genética , Clonagem Molecular , DNA Complementar/genética , DNA de Helmintos/genética , Epitopos de Linfócito T/genética , Antígenos H-2/genética , Proteínas de Helminto/biossíntese , Proteínas de Helminto/química , Proteínas de Helminto/genética , Proteínas de Helminto/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Onchocerca volvulus/enzimologia , Peptídeos/química , Peptídeos/genética , Peptídeos/imunologia , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Subpopulações de Linfócitos T/imunologia , Vacinas Sintéticas/genéticaRESUMO
The adjuvanticity of MALP-2, a 2-kDa synthetic lipopeptide with macrophage-stimulatory activity, was evaluated in BALB/c mice using beta-galactosidase (beta-gal) as model antigen. When co-administered with beta-gal by either the intranasal (i.n.) or i.p. route, MALP-2 (0.5 microg) was capable of increasing beta-gal-specific serum IgG titers by 675-3,560-fold (i.n.) and 64-128-fold (i.p.), respectively, as compared to immunization with beta-gal alone. Using MALP-2, almost maximal IgG responses were already stimulated following the first immunization, and the IgG titers were similar to those observed using 10 microg of cholera toxin B subunit (CTB) as adjuvant. The mucosal immune system was also effectively stimulated (p<0.05) when MALP-2 was administered by the i.n. route (36% and 23% of beta-gal-specific IgA in lung and vaginal lavages, respectively). The i.n. co-administration of MALP-2 stimulated a stronger cellular immune response than CTB, both in submandibular lymph nodes and spleen (p<0.05). The analysis of beta-gal-specific IgG isotypes and the profiles of cytokines secreted by in vitro re-stimulated cells showed that co-administration of MALP-2 triggered a dominant Th2-response pattern. A recruitment of B220(+) and MAC-1(+) cells with an up-regulated expression of MHC class I, CD80 (B7.1) and CD54 (ICAM-1) was observed in nasal associated lymphoid tissues from MALP-2 treated mice. Taken together, our results demonstrated that the synthetic lipopeptide MALP-2 represents a very promising adjuvant for the mucosal delivery of vaccine antigens.