RESUMO
Using a consensus epitope prediction approach, three rotavirus (RV) peptides that induce cytokine secretion by CD4 T cells from healthy volunteers were identified. The peptides were shown to bind HLA-DRB1*0101 and then used to generate MHC II tetramers. RV specific T cell lines specific for one of the three peptides studied were restricted by MHC class II molecules and contained T cells that bound the tetramer and secreted cytokines upon activation with the peptide. The majority of RV and Flu tetramer(+) CD4 T cells in healthy volunteers expressed markers of antigen experienced T cells, but only RV specific CD4 T cells expressed intestinal homing receptors. CD4 T cells from children that received a RV vaccine, but not placebo recipients, were stained with the RV-VP6 tetramer and also expressed intestinal homing receptors. Circulating RV-specific CD4 T cells represent a unique subset that expresses intestinal homing receptors.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Antígenos de Histocompatibilidade Classe II/imunologia , Integrinas/genética , Intestinos/imunologia , Receptores CCR/genética , Receptores Virais/genética , Infecções por Rotavirus/imunologia , Rotavirus/imunologia , Adolescente , Adulto , Criança , Feminino , Antígenos de Histocompatibilidade Classe II/química , Antígenos de Histocompatibilidade Classe II/genética , Humanos , Integrinas/imunologia , Intestinos/virologia , Masculino , Receptores CCR/imunologia , Receptores Virais/imunologia , Rotavirus/genética , Infecções por Rotavirus/genética , Infecções por Rotavirus/virologia , Especificidade da Espécie , Proteínas Virais/genética , Proteínas Virais/imunologia , Adulto JovemRESUMO
In vivo replication of rotaviruses is generally limited to enterocytes. Because of this restriction, most blood circulating rotavirus-specific B cells are hypothesized to originate in Peyer's patches and should express the intestinal homing receptor alpha4beta7. To test this hypothesis in humans, we used a flow cytometry assay that identifies antigen-activated (IgD-) B cells (CD19+) that express surface rotavirus-specific immunoglobulin. With this assay we could detect rotavirus-specific B cells in both children and adults with an acute rotavirus (RV) infection. Staining with an anti-alpha4beta7 monoclonal antibody, we could determine that B cells that express rotavirus-specific surface immunoglobulin predominantly express alpha4beta7. The response of rotavirus-specific antibody-secreting cells in the peripheral blood of children and adults with acute rotavirus infection was also studied by ELISPOT. The antibody-secreting cells of children were mainly of the IgM isotype, while the antibody-secreting cells of adults were predominantly of the IgA and IgG isotype. alpha4beta7+ and alpha4beta7- subsets of peripheral blood mononuclear cells were purified using paramagnetic beads and then tested in the ELISPOT assay. Rotavirus-specific antibody-secreting cells were predominantly present in the alpha4beta7+ subpopulation. The flow cytometry assay we have described will permit future studies to characterize the phenotype of virus-specific B cells and could be useful in the study of the immunogenicity and protective efficacy of RV vaccines and the identification of markers of protective immunity.