Interleukin-17-producing decidual CD4+ T cells are not deleterious for human pregnancy when they also produce interleukin-4.

BACKGROUND: Trophoblast expressing paternal HLA-C antigens resemble a semiallograft, and could be rejected by maternal CD4+ T lymphocytes. We examined the possible role in human pregnancy of Th17 cells, known to be involved in allograft rejection and reported for this reason to be responsible for miscarriages. We also studied Th17/Th1 and Th17/Th2 cells never investigated before. We defined for the first time the role of different Th17 subpopulations at the embryo implantation site and the role of HLA-G5, produced by the trophoblast/embryo, on Th17 cell differentiation. METHODS: Cytokine production by CD4+ purified T cell and T clones from decidua of normal pregnancy, unexplained recurrent abortion, and ectopic pregnancy at both embryo implantation site and distant from that site were analyzed for protein and mRNA production. Antigen-specific T cell lines were derived in the presence and in the absence of HLA-G5. RESULTS: We found an associated spontaneous production of IL-17A, IL-17F and IL-4 along with expression of CD161, CCR8 and CCR4 (Th2- and Th17-type markers) in fresh decidua CD4+ T cells during successful pregnancy. There was a prevalence of Th17/Th2 cells (producing IL-17A, IL-17F, IL-22 and IL-4) in the decidua of successful pregnancy, but the exclusive presence of Th17 (producing IL-17A, IL-17F, IL-22) and Th17/Th1 (producing IL-17A, IL-17F, IL-22 and IFN-γ) cells was found in the decidua of unexplained recurrent abortion. More importantly, we observed that Th17/Th2 cells were exclusively present at the embryo implantation site during tubal ectopic pregnancy, and that IL-4, GATA-3, IL-17A, ROR-C mRNA levels increased in tubal biopsies taken from embryo implantation sites, whereas Th17, Th17/Th1 and Th1 cells are exclusively present apart from implantation sites. Moreover, soluble HLA-G5 mediates the development of Th17/Th2 cells by increasing IL-4, IL-17A and IL-17F protein and mRNA production of CD4+ T helper cells. CONCLUSION: No pathogenic role of decidual Th17 cells during pregnancy was observed. Indeed, a beneficial role for these cells was observed when they also produced IL-4. HLA-G5 could be the key feature of the uterine microenvironment responsible for the development of Th17/Th2 cells, which seem to be crucial for successful embryo implantation.

Neurons are MHC class I-dependent targets for CD8 T cells upon neurotropic viral infection.

Following infection of the central nervous system (CNS), the immune system is faced with the challenge of eliminating the pathogen without causing significant damage to neurons, which have limited capacities of renewal. In particular, it was thought that neurons were protected from direct attack by cytotoxic T lymphocytes (CTL) because they do not express major histocompatibility class I (MHC I) molecules, at least at steady state. To date, most of our current knowledge on the specifics of neuron-CTL interaction is based on studies artificially inducing MHC I expression on neurons, loading them with exogenous peptide and applying CTL clones or lines often differentiated in culture. Thus, much remains to be uncovered regarding the modalities of the interaction between infected neurons and antiviral CD8 T cells in the course of a natural disease. Here, we used the model of neuroinflammation caused by neurotropic Borna disease virus (BDV), in which virus-specific CTL have been demonstrated as the main immune effectors triggering disease. We tested the pathogenic properties of brain-isolated CD8 T cells against pure neuronal cultures infected with BDV. We observed that BDV infection of cortical neurons triggered a significant up regulation of MHC I molecules, rendering them susceptible to recognition by antiviral CTL, freshly isolated from the brains of acutely infected rats. Using real-time imaging, we analyzed the spatio-temporal relationships between neurons and CTL. Brain-isolated CTL exhibited a reduced mobility and established stable contacts with BDV-infected neurons, in an antigen- and MHC-dependent manner. This interaction induced rapid morphological changes of the neurons, without immediate killing or impairment of electrical activity. Early signs of neuronal apoptosis were detected only hours after this initial contact. Thus, our results show that infected neurons can be recognized efficiently by brain-isolated antiviral CD8 T cells and uncover the unusual modalities of CTL-induced neuronal damage.

Specific immobilization of influenza A virus on GaAs (001) surface.

In the quest for the development of an all-optical biosensor for rapid detection and typing of viral pathogens, we investigate biosensing architectures that take advantage of strong photoluminescence emission from III-V quantum semiconductors (QS). One of the key elements in the development of such a biosensor is the ability to attach various analytes to GaAs--a material of choice for capping III-V QS of our interest. We report on the study of biofunctionalization of GaAs (001) with polyethylene-glycol (PEG) thiols and the successful immobilization of influenza A virus. A diluted solution of biotinylated PEG thiols in OH-terminated PEG thiols is used to form a network of sites for the attachment of neutravidin. Biotinylated polyclonal influenza A antibodies are applied to investigate the process of the immobilization of inactivated influenza A virus. The successful immobilization is demonstrated using atomic force microscopy and fluorescence microscopy measurements.

Tracking antigen-specific CD8+ T cells in the rat using MHC class I multimers.

Studies of the quantitative and qualitative aspects of anti-microbial, anti-tumoral or autoreactive immune responses have been greatly facilitated by the possibility to stain antigen-specific CD8(+) T cells using fluorescently labeled multimeric major histocompatibility complex (MHC) class I/peptide complexes. So far, this technology has been developed for human and mouse, but not yet in the rat. Here, we describe the generation of the first rat MHC multimer. We produced a rat RT1(l) Pro5 MHC Pentamer combined with the immunodominant peptide for Borna disease virus (BDV), in order to study the characteristics of the antiviral CD8(+) T cell response. BDV is an RNA virus that can cause persistent infections of the central nervous system (CNS), often associated with prominent brain inflammation. In adult Lewis rats, of the RT1(l) MHC haplotype, BDV infection leads to severe immune-mediated neurological symptoms. The pathogenic role of the immune response is due primarily to antiviral CD8(+) T cells, many of them being specific for an immunodominant epitope located in the BDV nucleoprotein (N(230-238)). Ex vivo flow cytometry analyses revealed that 3 to 12% of CD8(+) T cells found in the brains of BDV-infected rats stained positively with the BDV-Pentamer. Interestingly, the frequency of Pentamer-positive cells increased up to 3.3 fold after a short resting period in culture. Virus-specific CD8(+) T cells were mainly detected in the brain and were virtually undetectable in peripheral lymphoid organs. This novel rat Pro5 MHC Pentamer represents an attractive tool for the detection, isolation and characterization of antigen-specific CD8(+) T cell responses in the rat.

LF 15-0195 treatment protects against central nervous system autoimmunity by favoring the development of Foxp3-expressing regulatory CD4 T cells.

Experimental autoimmune encephalomyelitis (EAE) is an instructive model for the human demyelinating disease multiple sclerosis. Lewis (LEW) rats immunized with myelin-basic protein (MBP) develop EAE characterized by a single episode of paralysis, from which they recover spontaneously and become refractory to a second induction of disease. LF 15-0195 is a novel molecule that has potent immunosuppressive effects in several immune-mediated pathological manifestations, including EAE. In the present study, we show that a 30-day course of LF 15-0195 treatment not only prevents MBP-immunized LEW rats from developing EAE but also preserves their refractory phase to reinduction of disease. This effect is Ag driven since it requires priming by the autoantigen during the drug administration. In contrast to other immunosuppressive drugs, short-term treatment with this drug induces a persistent tolerance with no rebound of EAE up to 4 mo after treatment withdrawal. This beneficial effect of LF 15-0195 on EAE does not result from the deletion of MBP-specific Vbeta8.2 encephalitogenic T cells. In contrast, this drug favors the differentiation of MBP-specific CD4 T cells into Foxp3-expressing regulatory T cells that, upon adoptive transfer in syngeneic recipients, prevent the development of actively induced EAE. Finally, we demonstrate that the tolerance induced by LF 15-0195 treatment is not dependent on the presence of TGF-beta. Together, these data demonstrate that short-term treatment with LF 15-0195 prevents MBP-immunized LEW rats from EAE by favoring the development of Foxp-3-expressing regulatory CD4 T cells.

LF 15-0195 inhibits the development of rat central nervous system autoimmunity by inducing long-lasting tolerance in autoreactive CD4 T cells.

Experimental autoimmune encephalomyelitis (EAE) is a T cell-dependent autoimmune disease induced in susceptible animals by a single immunization with myelin basic protein (MBP). LF 15-0195 is a novel immunosuppressor that has been shown to have a potent immunosuppressive effect in several pathological manifestations. The purpose of this study was to investigate the effect of this drug on the induction and progression of established rat EAE and to dissect the mechanisms involved. We show that LF 15-0195 administration at the time of MBP immunization reduces the incidence and severity of EAE in Lewis rats. This drug also inhibits ongoing and passively induced EAE, indicating that LF 15-0195 affects already differentiated pathogenic lymphocytes. Compared with lymph node cells from untreated rats, lymphocytes from MBP-immunized rats treated with LF 15-0195 proliferated equally well in response to MBP in vitro, while their ability to produce effector cytokines and to transfer EAE into syngeneic recipients was significantly reduced. This phenomenon is stable and long-lasting. Indeed, neither IL-12 nor repeated stimulation with naive APC and MBP in vitro rendered MBP-specific CD4 T cells from protected rats encephalitogenic. In conclusion, LF 15-0195 treatment suppresses EAE by interfering with both the differentiation and effector functions of autoantigen-specific CD4 T cells.

LF 15-0195 prevents from the development and inhibits the progression of rat experimental autoimmune myasthenia gravis.

Experimental autoimmune myasthenia gravis (EAMG) is a T cell-dependent antibody-mediated neuromuscular autoimmune disease induced in susceptible rats by a single immunisation with torpedo acetylcholine receptor (AChR). Here, we report that subcutaneous administration of a novel immunosuppressant, LF 15-0195, is effective in inhibiting the induction and the progression of rat EAMG-suggesting that this drug may be used for preventive and curative treatment. The beneficial effect of LF 15-0195 is accompanied by decreased production of pathogenic autoantibodies and inhibition of the differentiation of antigen specific T cells into effector lymphocytes. These finding suggest that LF 15-0195 is a promising therapeutic for this autoimmune disease.