The role of the CD8+ T cell in the immune response to Brucella melitensis in mice

Prior to these studies, the literature indicated an essential role for CD8+ T cell-mediated cytotoxicity in control of brucellosis (B.abortus ). The initial effort in these studies was to develop a cytotoxicity assay that could be used to identify individual B.melitensis proteins that activate cytotoxic T cells. This effort failed. Therefore, studies were undertaken to assess the role of the CD8+ T cell in B.melitensis immunity. Overall, male CD8 knockout mice showed vaccine-induced protection from infection indistinguishable from that observed in control mice. However, limited individual CD8 knockout mice showed exacerbation of infection. To eliminate possible variation induced by stress, female mice were used in subsequent experiments. Female Fas mutant and perforin and CD8 knockout mice clear a primary B.melitensis infection normally, and, compared to control mice, show a similar humoral immune response, equivalent expression of markers of T cell memory, T cell proliferation and interferon-gamma production. Therefore, it is clear that cytotoxicity does not play a key role in the immune response to B.melitensis. Nonetheless, further studies were undertaken to fully elucidate the role of the CD8+ T cell. Unseparated spleen cells and CD8 and CD4+ T cells from immunized or virulent Brucella-exposed BALB/c mice were stimulated in vitro and incubated with Brucella-infected J774 cells. Only T cells from virulent B.melitensis-exposed mice induced reduction in the bacterial load. This effect was abrogated by treatment with anti-γ-IFN. This indicates that T cell-derived γ-IFN can mediate suppression of intracellular B.melitensis proliferation, however, immunization may not induce this effect. However, a final experiment showed that CD8 and CD4+ T cells and unseparated spleen cells from immunized mice specifically produce γ-IFN in vitro. This strongly indicates that immunization induces γ-IFN-producing, memory CD8+ T cells. This assay can now be used to evaluate B. melitensis proteins for their ability to stimulate γ-IFN production from CD8+ T cells. When key proteins are identified, they can be delivered as subunits for the possible eventual construction of a vaccine that specifically induces γ-IFN production from CD8+ T cells in vivo