Magali Irla

Autoantigen-specific interactions with CD4+ thymocytes control mature medullary thymic epithelial cell cellularity.

Medullary thymic epithelial cells (mTECs) are specialized for inducing central immunological tolerance to self-antigens. To accomplish this, mTECs must adopt a mature phenotype characterized by expression of the autoimmune regulator Aire, which activates the transcription of numerous genes encoding tissue-restricted self-antigens. The mechanisms that control mature Aire(+) mTEC development in the postnatal thymus remain poorly understood. We demonstrate here that, although either CD4(+) or CD8(+) thymocytes are sufficient to sustain formation of a well-defined medulla, expansion of the mature mTEC population requires autoantigen-specific interactions between positively selected CD4(+) thymocytes bearing autoreactive T cell receptor (TCR) and mTECs displaying cognate self-peptide-MHC class II complexes. These interactions also involve the engagement of CD40 on mTECs by CD40L induced on the positively selected CD4(+) thymocytes. This antigen-specific TCR-MHC class II-mediated crosstalk between CD4(+) thymocytes and mTECs defines a unique checkpoint in thymic stromal development that is pivotal for generating a mature mTEC population competent for ensuring central T cell tolerance.

MHC class II–restricted antigen presentation by plasmacytoid dendritic cells inhibits T cell–mediated autoimmunity

Although plasmacytoid dendritic cells (pDCs) express major histocompatibility complex class II (MHCII) molecules, and can capture, process, and present antigens (Ags), direct demonstrations that they function as professional Ag-presenting cells (APCs) in vivo during ongoing immune responses remain lacking. We demonstrate that mice exhibiting a selective abrogation of MHCII expression by pDCs develop exacerbated experimental autoimmune encephalomyelitis (EAE) as a consequence of enhanced priming of encephalitogenic CD4+ T cell responses in secondary lymphoid tissues. After EAE induction, pDCs are recruited to lymph nodes and establish MHCII-dependent myelin-Ag–specific contacts with CD4+ T cells. These interactions promote the selective expansion of myelin-Ag–specific natural regulatory T cells that dampen the autoimmune T cell response. pDCs thus function as APCs during the course of EAE and confer a natural protection against autoimmune disease development that is mediated directly by their ability to present of Ags to CD4+ T cells in vivo.

Control of central self-tolerance induction by autoreactive CD4+ thymocytes

Medullary thymic epithelial cells (mTECs) are crucial for the selection of a T-cell-receptor (TCR) repertoire purged of self-reactive specificities, because these cells activate a promiscuous gene-expression program that leads to the synthesis of a wide array of peripheral tissue-restricted self-antigens. This review summarizes recent progress in our understanding of the cellular interactions, ligands, receptors and signal-transduction pathways that control mature-mTEC development. The particular focus is on new findings supporting the model that mature-mTEC development in the postnatal thymus depends on nuclear factor-kappaB (NF-kappaB) signaling induced by CD40-CD40 ligand, and receptor-activator-of-NF-kappaB (RANK)-RANK ligand interactions, and that these signals are delivered in the context of antigen-specific interactions between CD4(+) thymocytes carrying autoreactive TCRs and mTECs displaying cognate autoantigen-MHC-class-II complexes.

Anti-CD154 mAb and Rapamycin Induce T Regulatory Cell Mediated Tolerance in Rat-to-Mouse Islet Transplantation

Background Anti-CD154 (MR1) monoclonal antibody (mAb) and rapamycin (RAPA) treatment both improve survival of rat-to-mouse islet xenograft. The present study investigated the effect of combined RAPA/MR1 treatment on rat-to-mouse islet xenograft survival and analyzed the role of CD4+CD25+Foxp3+ T regulatory cells (Treg) in the induction and maintenance of the ensuing tolerance. Methodology/Principal Findings C57BL/6 mice were treated with MR1/RAPA and received additional monoclonal anti-IL2 mAb or anti CD25 mAb either early (0–28 d) or late (100–128 d) post-transplantation. Treg were characterised in the blood, spleen, draining lymph nodes and within the graft of tolerant and rejecting mice by flow cytometry and immunohistochemistry. Fourteen days of RAPA/MR1 combination therapy allowed indefinite islet graft survival in >80% of the mice. Additional administration of anti-IL-2 mAb or depleting anti-CD25 mAb at the time of transplantation resulted in rejection (100% and 89% respectively), whereas administration at 100 days post transplantation lead to lower rejection rates (25% and 40% respectively). Tolerant mice showed an increase of Treg within the graft and in draining lymph nodes early post transplantation, whereas 100 days post transplantation no significant increase of Treg was observed. Rejecting mice showed a transient increase of Treg in the xenograft and secondary lymphoid organs, which disappeared within 7 days after rejection. Conclusions/Significances These results suggest a critical role for Treg in the induction phase of tolerance early after islet xenotransplantation. These encouraging data support the need of developing further Treg therapy for overcoming the species barrier in xenotransplantation.

A General Survey of Thymocyte Differentiation by Transcriptional Analysis of Knockout Mouse Models

The thymus is the primary site of T cell lymphopoiesis. To undergo proper differentiation, developing T cells follow a well-ordered genetic program that strictly depends on the heterogeneous and highly specialized thymic microenvironment. In this study, we used microarray technology to extensively describe transcriptional events regulating αβ T cell fate. To get an integrated view of these processes, both whole thymi from genetically engineered mice together with purified thymocytes were analyzed. Using mice exhibiting various transcriptional perturbations and developmental blockades, we performed a transcriptional microdissection of the organ. Multiple signatures covering both cortical and medullary stroma as well as various thymocyte maturation intermediates were clearly defined. Beyond the definition of histological and functional signatures (proliferation, rearrangement), we provide the first evidence that such an approach may also highlight the complex cross-talk events that occur between maturing T cells and stroma. Our data constitute a useful integrated resource describing the main gene networks set up during thymocyte development and a first step toward a more systematic transcriptional analysis of genetically modified mice.

Thymic Crosstalk Coordinates Medulla Organization and T-Cell Tolerance Induction

The thymus ensures the generation of a functional and highly diverse T-cell repertoire. The thymic medulla, which is mainly composed of medullary thymic epithelial cells (mTECs) and dendritic cells (DCs), provides a specialized microenvironment dedicated to the establishment of T-cell tolerance. mTECs play a privileged role in this pivotal process by their unique capacity to express a broad range of peripheral self-antigens that are presented to developing T cells. Reciprocally, developing T cells control mTEC differentiation and organization. These bidirectional interactions are commonly referred to as thymic crosstalk. This review focuses on the relative contributions of mTEC and DC subsets to the deletion of autoreactive T cells and the generation of natural regulatory T cells. We also summarize current knowledge regarding how hematopoietic cells conversely control the composition and complex three-dimensional organization of the thymic medulla.

Three-Dimensional Visualization of the Mouse Thymus Organization in Health and Immunodeficiency

Lymphoid organs exhibit complex structures tightly related to their function. Surprisingly, although the thymic medulla constitutes a specialized microenvironment dedicated to the induction of T cell tolerance, its three-dimensional topology remains largely elusive because it has been studied mainly in two dimensions using thymic sections. To overcome this limitation, we have developed an automated method for full organ reconstruction in three dimensions, allowing visualization of intact mouse lymphoid organs from a collection of immunolabeled slices. We validated full organ reconstruction in three dimensions by reconstructing the well-characterized structure of skin-draining lymph nodes, before revisiting the complex and poorly described corticomedullary organization of the thymus. Wild-type thymi contain ∼200 small medullae that are connected to or separated from a major medullary compartment. In contrast, thymi of immunodeficient Rag2−/− mice exhibit only ∼20 small, unconnected medullary islets. Upon total body irradiation, medullary complexity was partially reduced and then recovered upon bone marrow transplantation. This intricate topology presents fractal properties, resulting in a considerable corticomedullary area. This feature ensures short distances between cortex and medulla, hence efficient thymocyte migration, as assessed by mathematical models. Remarkably, this junction is enriched, particularly in neonates, in medullary thymic epithelial cells expressing the autoimmune regulator. The emergence of a major medullary compartment is induced by CD4+ thymocytes via CD80/86 and lymphotoxin-α signals. This comprehensive three-dimensional view of the medulla emphasizes a complex topology favoring efficient interactions between developing T cells and autoimmune regulator–positive medullary thymic epithelial cells, a key process for central tolerance induction.

Repression of Arginase-2 Expression in Dendritic Cells by MicroRNA-155 Is Critical for Promoting T Cell Proliferation

Arginine, a semiessential amino acid implicated in diverse cellular processes, is a substrate for two arginases—Arg1 and Arg2—having different expression patterns and functions. Although appropriately regulated Arg1 expression is critical for immune responses, this has not been documented for Arg2. We show that Arg2 is the dominant enzyme in dendritic cells (DCs) and is repressed by microRNA-155 (miR155) during their maturation. miR155 is known to be strongly induced in various mouse and human DC subsets in response to diverse maturation signals, and miR155-deficient DCs exhibit an impaired ability to induce Ag-specific T cell responses. By means of expression profiling studies, we identified Arg2 mRNA as a novel miR155 target in mouse DCs. Abnormally elevated levels of Arg2 expression and activity were observed in activated miR155-deficient DCs. Conversely, overexpression of miR155 inhibited Arg2 expression. Bioinformatic and functional analyses confirmed that Arg2 mRNA is a direct target of miR155. Finally, in vitro and in vivo functional assays using DCs exhibiting deregulated Arg2 expression indicated that Arg2-mediated arginine depletion in the extracellular milieu impairs T cell proliferation. These results indicate that miR155-induced repression of Arg2 expression is critical for the ability of DCs to drive T cell activation by controlling arginine availability in the extracellular environment.

Ag-presenting CpG-activated pDCs prime Th17 cells that induce tumor regression

Plasmacytoid dendritic cells (pDC) rapidly and massively produce type I IFN and other inflammatory cytokines in response to foreign nucleic acids, thereby indirectly influencing T-cell responses. Moreover, antigen (Ag)-presenting pDCs directly regulate T-cell differentiation. Depending on the immune environment, pDCs exhibit either tolerogenic or immunogenic properties. Here, we show that CpG-activated pDCs promote efficient Th17 differentiation. Indeed, Th17 responses are defective in mice selectively lacking MHCII on pDCs upon antigenic challenge. Importantly, in those mice, the frequency of Th17 cells infiltrating solid tumors is impaired. As a result, the recruitment of infiltrating leukocytes in tumors, including tumor-specific cytotoxic T lymphocytes (CTL), is altered and results in increased tumor growth. Importantly, following immunization with tumor Ag and CpG-B, MHCII-restricted Ag presentation by pDCs promotes the differentiation of antitumor Th17 cells that induce intratumor CTL recruitment and subsequent regression of established tumors. Our results highlight a new role for Ag presenting activated pDCs in promoting the development of Th17 cells and impacting on antitumor immunity.

IDO-orchestrated crosstalk between pDCs and Tregs inhibits autoimmunity.

Plasmacytoid dendritic cells (pDCs) have been shown to both mediate and prevent autoimmunity, and the regulation of their immunogenic versus tolerogenic functions remains incompletely understood. Here we demonstrate that, compared to other cells, pDCs are the major expressors of Indoleamine-2,3-dioxygenase (IDO) in steady-state lymph nodes (LNs). IDO expression by LN pDCs was closely dependent on MHCII-mediated, antigen-dependent, interactions with Treg. We further established that IDO production by pDCs was necessary to confer suppressive function to Tregs. During EAE development, IDO expression by pDCs was required for the generation of Tregs capable of dampening the priming of encephalitogenic T cell and disease severity. Thus, we describe a novel crosstalk between pDCs and Tregs: Tregs shape tolerogenic functions of pDCs prior to inflammation, such that pDCs in turn, promote Treg suppressive functions during autoimmunity.