RESUMO
The heat shock protein Hsp65 has been characterized previously in several mycobacterial species. This is the first report of the complete sequence of the coding region of the Mycobacterium avium homologue. The sequence was highly homologous to the Hsp65 of other mycobacterial species, as well as being related closely to the murine and human homologues. Recombinant Hsp65 (rHsp65) was expressed in Escherichia coli to high levels and the recombinant protein tested for its immunogenicity in a murine model of M. avium infection. Although mice infected with M. avium produced antibodies that reacted with rHsp65, they showed low proliferative T-cell responses and no cytokine production in response to the same antigen. However, immunization with rHsp65 in the adjuvant dimethydioctadecylammonium bromide (DDA), induced T cells that responded to native Hsp65 with proliferation and IFN-gamma production, indicating that the recombinant and native forms of the protein were antigenically similar. Therefore, the findings indicate that Hsp65 is not a dominant T-cell antigen during M. avium infection.
Assuntos
Chaperoninas , Complexo Mycobacterium avium/genética , Complexo Mycobacterium avium/imunologia , Adjuvantes Imunológicos , Sequência de Aminoácidos , Animais , Anticorpos Antibacterianos/sangue , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Proteínas de Bactérias/metabolismo , Células Cultivadas , Chaperonina 60 , Chaperoninas/química , Chaperoninas/genética , Chaperoninas/imunologia , Chaperoninas/metabolismo , Clonagem Molecular , Modelos Animais de Doenças , Humanos , Linfonodos/citologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Infecção por Mycobacterium avium-intracellulare/imunologia , Compostos de Amônio Quaternário/farmacologia , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/metabolismo , Alinhamento de Sequência , Subpopulações de Linfócitos T/imunologiaRESUMO
Infection of different strains of mice with Mycobacterium avium has revealed genetic control of the immunoglobulin isotype induced and of the balance between Th1 and Th2 cytokines. Female BALB/c or C57BL/10 mice were infected intranasally with 10(5) M. avium organisms. The antibody response was measured over 18 weeks by enzyme-linked immunosorbent assay and Western blotting, while numbers of cytokine-producing cells were assessed at 12 to 15 weeks by ELISPOT assay. Upon infection, C57BL/10 mice produced a clear Th1 response with strong gamma interferon (IFN-gamma) production, no interleukin-4 (IL-4), and almost entirely immunoglobulin G2a (IgG2a) antibody. In contrast, BALB/c mice developed T cells producing IL-4, as well as those producing IFN-gamma, while the antibody response was a mixture of IgG1 and IgG2a. Antibodies from BALB/c mice were also able to recognize a greater range of antigens than were C56BL/10 mice. B10D2 mice, which carry the BALB/c major histocompatibility complex haplotype on a C57BL/10 background, followed the C57BL/10 cytokine pattern. Mice infected with Listeria monocytogenes did not show a similar response dichotomy.
