Molecular And Cellular Mechanisms Of T Cell Immune Responses Induced By Recombinant Mycobacterium Bovis BCG

Mycobacterium bovis BCG is the most widely used vaccine all over the world, and represents one of the most promising live vectors to deliver foreign antigens to the immune system. However, until now, little is known about the mechanisms of interaction between BCG or recombinant BCG(rBCG) and host immune system, and the detailed characteristics of cell ?mediated immunity induced by rBCG or BCG. To address these questions, the rBCG expressing MalE protein of Escherichia coil (rBCG ?MalE) was used as model strain to study the mechanisms by which MalE protein is presented by major histocompatibility complex class I (MHC I )molecules to T cells, to define the functional diversity of T cell epitopes of expressed MalE from rBCG, and to demonstrate the characteristics and regulation mechanisms of Th responses initiated by rBCG MalE. The epitope repertoire of expressed MalE from rBCG was analyzed in vitro by antigen presentation assay using dendritic cells (DCs) as antigen presenting cell (APC) pulsed with rBCG ?MalE. ?161 ? BCG ?wt or purified MalE protein. Four T cell epitopes -of MalE protein presented on the surface of those DCs pulsed with rBCG MalE and purified MalE protein were recognized by CD4~ T hybridomas specific for MalE peptides p68?2, p100?14, p151? 165 and p277?91,respectively, whereas DCs pulsed with BCG ?wt failed to stimulate these T hybridomas. The results also showed that rBCG ?MalE induced in vivo T cell proliferation and IFN ? responses against MalE protein and its peptides or PPD antigen while BCG ?wt only triggered responses specific for PPD. rBCG MalE functionally expressed the four T cell epitopes of MalE protein and epitope mapping from mice immunized with rBCG e MalE showed that no novel epitopes or cryptic epitopes were revealed in rBCG, and the p68?2 was immunodominant epitope and the others were subdominant epitopes. Phagocytic cells with APC functions play an important role in resistance to bacterial infection in turn that they can stimulate T cell responses enhancing bacterial clearance. Macrophages (M $Zf) are privileged host cells for intracellular bacteria and thus represent a large reservoir of antigenic material, but DCs are much more potent APC. Therefore, in T cell immunity to bacteria the role of these two phagocytic cells is not clearly established; Here, we investigated in vivo the relative contribution of M ~ and DC subsets to the antibacterial T cell response to Mycobacterium bovis BCG. Twelve hours after i. v. administration of rBCG . MalE, the rate of infection in the spleen was comparable for M ~ and DC. However, the presence of immunogenic MalE peptides/MHC II complexes was det&cted ex vivo on DC, but not on M ~, using T cell hybridomas specific for the MalE protein. Likewise ,upregulation of CD4O, B7. 1 and B7. 2 molecules and production of IL?2 p40 following infection was only observed for DC, confirming the exclusive role of DC in stimulating T cell responses. CD8a~ and CD8cZ spleen DC were ?162 ? equally potent antigen presenting cells in vivo, but the production of IL ?12 was mainly associated with the CD8a~ subset. Altogether, these data indicated .that in vivo DC playeda primary role not only in acquired immunity to mycobacteria in the early times of infection, but also in innate immunity through IL?2 secretion.