Regulatory cells, by virtue of their capacity to control the vigor of immune responses are essential to the maintenance of host homeostasis. Several types of CD4 regulatory T cells exist some of which are induced in response to infectious challenge and some of which are judged as natural regulators (natural Treg). Inducible Treg cells such as Tr1 or TH3 cells can develop from conventional CD4 T cells that are exposed to specific stimulatory conditions. Natural Treg cells, however, arise during the normal process of maturation in the thymus, obey defined rules and express a specific set of markers. Recently, studies also indicate that a unique transcription factor Foxp3 is required for the generation of natural Treg cells and this represents, to date, their most specific marker. Natural Treg play a central role in the control of autoimmunity, a function that is associated with their capacity to recognize self-antigen. Whether or not they also recognize foreign antigens and the extent of their repertoire for such antigens remain unknown. We and others have shown that natural Treg also play a critical role in the outcome of microbial infections. Natural Treg help limit collateral tissue damage caused by vigorous antimicrobial immune responses. These cells can also limit the magnitude of effector responses which result in failure to adequately control infection. Furthermore, there are clear evidences that the efficiency of vaccines can also be hampered by the presence of natural Treg. Thus, strategies aimed to manipulate natural Treg cells function or number, have clearly high therapeutic potential. To develop rational strategies, there is an urgent need to better characterize the function of natural Treg during infections. Using two models of unicellular gastrointestinal parasitic infection (Cryptosporidium muris and Encephalitozoon cuniculi) and one model of cutaneous infection (Leishmania major) we are exploring the antigen specificity of natural Treg that accumulate at sites of infections as well as the conditions that favor their retention and function.? ? We have successfully developed a murine model of microsporia infection in the laboratory. E.cuniculi are obligate intracellular parasites that infect a wide range of hosts including mice. The primary mode of infection occurs through the GI route, mainly through contamination of water sources. The initial site of infection is the duodenum. Recently, it was shown that in contrast with mice infected intraperitoneally, control of oral infection relies mainly on intraepithelial CD4+ and CD8+ T lymphocytes. We confirmed that IFN-gamma was required to control the parasite following oral infection. Strong Th1 and Th17 responses are induced during infection in the GALT. While IFN-gamma has been shown to be necessary for efficient control of this infection, a role for IL-17 has not yet been addressed. Removal of Treg enhanced immune responses (both IFN-gamma and IL-17) against the pathogen without inducing a Th2 response. Conversely, the transfer of Treg from the Lamina propria of infected mice (but not from other sites) into WT mice prior to infection strongly impaired the expression of protective responses.? ? Recent data, from our laboratory and others, clearly demonstrate that Treg accumulate at sites of infection. Although the mechanisms underlying this accumulation remain largely unknown, the influx of Treg is likely to depend on their antigen specificity as well as on appropriate host-derived signals for their recruitment and local survival. Differences in chemokine responsiveness or receptor expression between Treg and effector T cells have been demonstrated in various models. However, most of the available data were obtained using Treg purified from lymphoid organs in mice or peripheral blood in humans. There is virtually no data on the signals and molecules that are involved in the traffic and retention of Treg at local sites of infection where regulation takes place. Using our Leishmania animal model we have evaluated the mechanisms favoring the homing of Treg at sites of infection. We had previously shown that the integrin CD103 was necessary for the retention of natural Treg at site of Leishmania infection. We have pursued these studies by demonstrating that the chemokine receptor CCR5 at the surface of Treg was also required for the accumulation of these cells at sites of L.major infection. The elimination of this molecule on Treg led to enhanced effector immune responses and better control of the infection. ? ? The targets of Treg control at sites of infection remain poorly understood. Intravital microscopy allows imaging of cells in live tissues and thus could shed light on Treg function. Previous intravital imaging of Treg has focused on dynamics of transferred transgenic cells within lymph nodes. Key findings include the lack of stable prolonged contact between Treg and effector T cells (Teff) as well as persistent contact between Treg and dendritic cells. In addition, previous work showed that Treg and effector T cells move in almost identical fashion in the pancreatic lymph node in a mouse model of diabetes. These findings call into question the importance of Treg-Teff contact in vivo and suggest a more important role for secreted cytokines or direct effects upon antigen presenting cells. Nevertheless, no studies have addressed the motility of Treg in tissue effector sites outside the lymph node. In addition, physiologic polyclonal T cell populations may behave differently from transferred transgenic cells.
The aim of this project is to examine the behavior of polyclonal Treg in the dermis during parasitic Leishmania major infection. Characterizing their movement in the dermis will provide insight into mechanisms by which Treg are acting in tissues.? We have developed a unique approach allowing the monitoring of a single lesion over a period of several months. Using a system in which Treg, effector T cells and parasites can be visualized by the expression of respectively eGFP, CFP and RFP we were able to show that Treg accumulate as early as a few days post infection and are in stable contact with infected macrophages. In contrast, contacts with effector T cells are rare and only transient. While a significant percentage of the Treg arrest near the infected cells, effector T cells do not stop. This work strongly supports the idea that one of the primary targets of Treg at sites of infection are infected macrophages
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