Biomedical subjects
Publications and source records attributed to Stephen J McSorley.
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A number of studies have documented suppression of lymphocyte activation in mice infected with Salmonella. Here, we describe incomplete activation of CD4+ T cells following intravenous injection of specific peptide and LPS into Salmonella-infected mice. Although antigen-specific CD4+ T cells were activated by peptide/LPS to increase surface CD69 expression, they did not produce IL-2 or TNF-alpha. Suppression of cytokine production did not require prolonged exposure of the T cells to the Salmonella-infected environment, was not antigen specific, but was dependent upon the presence of LPS during stimulation. These data suggest that Salmonella-infected mice are exquisitely sensitive to the generation of a suppressive environment following innate immune stimulation with LPS. In agreement with this interpretation, repeated low-dose administration of LPS caused uncontrolled replication of attenuated Salmonella in vivo.
The mammalian immune response to Salmonella has long been a subject of scientific study. Indeed, many of the general aspects of bacterial pathogenesis and host immune defense have been well described. However, a lack of clarity remains concerning important aspects of the host immune response to Salmonella, particularly with regard to the induction of an immune response in the intestinal mucosa. A major limitation has been the general lack of knowledge about specific antigenic targets that are recognized by both the innate and adaptive immune response in the intestine. Progress towards the identification of these targets is critical for the development of a detailed model of immunity to Salmonella and will lead to a better understanding of mucosal immune responses to other intracellular pathogens.
T cell activation by dendritic cells (DCs) is critical to the initiation of adaptive immune responses and protection against pathogens. Here, we demonstrate that a specialized DC subset in Peyer's patches (PPs) mediates the rapid activation of pathogen specific T cells. This DC subset is characterized by the expression of the chemokine receptor CCR6 and is found only in PPs. CCR6(+) DCs were recruited into the dome regions of PPs upon invasion of the follicle associated epithelium (FAE) by an enteric pathogen and were responsible for the rapid local activation of pathogen-specific T cells. CCR6-deficient DCs were unable to respond to bacterial invasion of PPs and failed to initiate T cell activation, resulting in reduced defense against oral infection. Thus, CCR6-dependent regulation of DCs is responsible for localized T cell dependent defense against entero-invasive pathogens.
Despite the recognized role of the T-bet transcription factor in the differentiation of Th1 cells, T-bet-deficient mice can develop small numbers of IFN-gamma-producing CD4 T cells. Although these are not sufficient to allow normal handling of some pathogens, T-bet-deficient mice do resolve infection with the intracellular pathogen Listeria monocytogenes. In contrast, we report that expression of T-bet is required for resistance to Salmonella infection. T-bet-deficient mice succumbed to infection with attenuated Salmonella and did not generate IFN-gamma-producing CD4 T cells or isotype-switched Salmonella-specific Ab responses. Spleen cells from Salmonella-infected T-bet-deficient mice secreted increased levels of IL-10, but not IL-4, upon in vitro restimulation. A Salmonella-specific TCR transgenic adoptive transfer system was used to further define the involvement of T-bet expression in the development of Salmonella-specific Th1 cells. Wild-type Salmonella-specific CD4 T cells activated in T-bet-deficient recipient mice displayed no defect in clonal expansion, contraction, or IFN-gamma production. In contrast, T-bet-deficient, Salmonella-specific CD4 T cells activated in wild-type recipient mice produced less IFN-gamma and more IL-2 upon in vivo restimulation. Therefore, expression of T-bet by CD4 T cells is required for the development of Salmonella-specific Th1 cells, regulation of IL-10 production, and resistance to Salmonella infection.
Bacterial flagellins are important components of the motility apparatus used by many microbial pathogens. These proteins are also targets of the innate and adaptive immune response of the host during infection and autoimmune disease. Flagellin interacts with TLR-5 and leads to the generation of a pro-inflammatory response and activation of host dendritic cells in vivo. Furthermore, flagellin is recognized by antibody and CD4 T cells responses during Salmonella infection. Here, we review recent developments in the understanding of flagellin interactions with the host immune system.
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Despite the current availability of Salmonella vaccines, typhoid fever remains a significant public health problem in developing countries. A greater understanding of T-cell activation and the development of immunological memory during Salmonella infection should lead to the development of more effective prophylactic intervention. Here, we review recent literature on the initiation, expansion and memory development of T-cell responses using the mouse model of typhoid. We pay particular attention to strategies for tracking T-cell responses in vivo and ex vivo, and suggest models to integrate some these studies.
Many pathogens can establish a lethal infection from relatively small inocula, yet the effect of infectious dose upon CD4 T cell activation is not clearly understood. This issue was examined by tracking Salmonella flagellin-specific SM1 T cells in vivo, after i.v. and oral challenge of mice with virulent Salmonella typhimurium. SM1 T cells rapidly expressed activation markers and expanded in response to high-dose infection but remained completely unresponsive in mice challenged with low doses of Salmonella. SM1 T cells, in these mice, remained unresponsive, despite massive bacterial replication in vivo. Naive SM1 T cells in low-dose Salmonella-infected mice were activated rapidly after the injection of flagellin peptide, demonstrating that these T cells were fully capable of responding, ruling out the possibility of a bacterial-induced suppressive environment. The inability of flagellin-specific SM1 T cells to respond to low-dose infection was not due to Ag down-regulation, because flagellin expression was detected using a functional assay. Together, these data suggest that low-dose Salmonella infection can evade flagellin-specific CD4 T cell activation in vivo.
The clonal burst size of CD4 T cells is predicted to be less than that of CD8 T cells. In this study, we demonstrate that massive numbers of Ag-specific CD4 T cells respond during vaccination of mice with live attenuated Salmonella, reaching a peak frequency of approximately 50% of CD4 T cells. Salmonella-specific T cells persisted at high frequency for several weeks and could be detected in the memory population for months after infection. Surprisingly, the expansion of endogenous Salmonella-specific CD4 T cells prevented the persistence of adoptively transferred Salmonella-specific T cells in vivo, demonstrating interclonal competition for access to the memory compartment.
After in vivo immunization, Ag-specific T cells disappear from circulation and become sequestered in lymphoid tissue where they encounter Ag presented by dendritic cells. In the same site and just after Ag presentation, they "disappear" a second time and we investigated this process. Using a mouse model of T cell deletion (without Toll-like receptor (TLR) stimulation) vs survival (with TLR stimulation), Ag-specific T cells indeed became undetectable by flow cytometry, however were readily detected by immunohistochemistry. Thus, whether or not the activated T cells were destined to delete or survive, they were difficult to extract from lymphoid tissue and did not disappear but in fact were abundantly present. Nevertheless, profound differences were observed during this time period when tolerizing conditions were compared with immunizing conditions. TLR stimulation induced an increase in CD25 expression, acquisition of surface MHC class II, and abnormally high increases in forward and side scatter of the peptide-specific T cells. Using a modified adoptive transfer approach, we demonstrated by flow cytometry that in the presence of TLR stimulation the Ag-specific T cells were tightly coupled to dendritic cells, explaining the unusual increases in size and granularity. Ultimately, these events induced the specific T cells to differentiate into memory cells. We postulate that this is a stage where T cells are either conditioned to survive or to delete depending upon the activation status of the innate immune system.
Advances in immune visualization have enabled the physical tracking of immune responses in vivo. The adaptation of such technology to models of infectious disease holds the promise of a more detailed analysis of host-pathogen interactions in a natural setting. However, the visualization of pathogen-specific immune responses in vivo confronts challenges that are inherent to the study of infectious disease systems. Recent attempts to track pathogen-specific immune responses in vivo validate the usefulness and underline the complexity of this experimental strategy.
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Peptide:MHC II complexes derived from a fluorescent antigen were detected in vivo to identify the cells that present subcutaneously injected antigen to CD4 T cells. Skin-derived dendritic cells (DCs) that acquired the antigen while in the draining lymph nodes were the first cells to display peptide:MHC II complexes. Presentation by these cells induced CD69, IL-2 production, and maximal proliferation by the T cells. Later, DCs displaying peptide:MHC II complexes migrated from the injection site via a G protein-dependent mechanism. Presentation by these migrants sustained expression of the IL-2 receptor and promoted delayed type hypersensitivity. Therefore, presentation of peptide:MHC II complexes derived from a subcutaneous antigen occurs in two temporally distinct waves with different functional consequences.
Flagellin is secreted by many enteric bacteria and, upon reaching the basolateral membrane of the intestinal epithelium, activates Toll-like receptor 5-mediated innate immune signaling pathways. We hypothesized that any flagellin that gets beyond the epithelium might also regulate cells of the adaptive immune system. Here we demonstrate that the clonal expansion of naive DO11.10 CD4 T cells in response to OVA peptide (323-339) was enhanced 3- to 10-fold in the presence of purified bacterial flagellin in vivo. OVA-specific CD4 T cells were also shown to have undergone more cell division in vivo if flagellin was coinjected with OVA. Flagellin administration increased the expression of B7-1 on splenic dendritic cells, and coinjection of CTLA4-Ig, which is known to block B7 function in vivo, completely ablated the adjuvant effect on CD4 T cells. Therefore, a conserved bacterial protein produced by many intestinal microbes can modulate CD4 T cell activation in vivo. Such an adjuvant effect for flagellin has important implications for vaccine development and the generation of CD4 T cell responses to enteric bacteria.
A novel adoptive transfer system was used to track the fate of naive Salmonella-specific CD4 T cells in vivo. These cells showed signs of activation in the Peyer's patches as early as 3 hr after oral infection. The activated CD4 T cells then produced IL-2 and proliferated in the T cell areas of these tissues before migrating into the B cell-rich follicles. In contrast, Salmonella-specific CD4 T cells were not activated in the spleen and very few of these cells migrated to the liver, despite the presence of bacteria in both organs. These results show that the T cell response to pathogenic Salmonella infection is localized to the gut-associated lymphoid tissue and does not extend efficiently to the major sites of late infection.