Thymic Antigen-presenting Cells Research Articles

Antigen presentation for central tolerance induction.

The extent of central T cell tolerance is determined by the diversity of self-antigens that developing thymocytes 'see' on thymic antigen-presenting cells (APCs). Here, focusing on insights from the past decade, we review the functional adaptations of medullary thymic epithelial cells, thymic dendritic cells and thymic B cells for the purpose of tolerance induction. Their distinct cellular characteristics range from unconventional phenomena, such as promiscuous gene expression or mimicry of peripheral cell types, to strategic positioning in distinct microenvironments and divergent propensities to preferentially access endogenous or exogenous antigen pools. We also discuss how 'tonic' inflammatory signals in the thymic microenvironment may extend the intrathymically visible 'self' to include autoantigens that are otherwise associated with highly immunogenic peripheral environments.

Evaluating in vivo approaches for studying the roles of thymic DCs in T cell development in mice.

T cells express an enormous repertoire of T cell receptors, enabling them to recognize any potential antigen. This large repertoire undergoes stringent selections in the thymus, where receptors that react to self- or non-danger-associated- antigens are purged. We know that thymic tolerance depends on signals and antigens presented by the thymic antigen presenting cells, but we still do not understand precisely how many of these cells actually contribute to tolerance. This is especially true for thymic dendritic cells (DC), which are composed of diverse subpopulations that are derived from different progenitors. Although the importance of thymic DCs has long been known, the functions of specific DC subsets have been difficult to untangle. There remains insufficient systematic characterization of the ontogeny and phenotype of thymic APCs in general. As a result, validated experimental models for studying thymic DCs are limited. Recent technological advancement, such as multi-omics analyses, has enabled new insights into thymic DC biology. These recent findings indicate a need to re-evaluate the current tools used to study the function of these cells within the thymus. This review will discuss how thymic DC subpopulations can be defined, the models that have been used to assess functions in the thymus, and models developed for other settings that can be potentially used for studying thymic DCs.

The potential role of infusions of T regulatory cells in inducing and maintaining liver allograft tolerance.

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DM functions differently in thymic antigen presentation cell subsets in development of autoreactive CD4+ T cells

Abstract Autoreactive CD4+ T cells broadly exist in healthy individuals and potentially cause autoimmune diseases; however, why they can escape from thymic negative selection is still controversial. We observed that the activity of DM, a peptide epitope editor in MHC class II antigen presentation pathway, is differentially regulated in different thymic antigen presenting cells (APC). It is high in cTEC that involved in cortical positive selection, but low in mTEC, B cell, pDC and cDC that mediate negative selection in medulla, while it remains high in all types of peripheral APC. The molecular mechanism of differential regulation is still unknown. It is not directly inhibited by DO, a negative regulator of DM that directly competes DM catalytic site with peptide-MHC class II complex, since DM editing activity remains low in those thymic medulla APC in DO−/− mice. We further confirmed that presentation of autoantigen epitope MOG35–55 for thymic positive selection and in vitro activation of autoreactive CD4+ T cells in EAE, a mouse model of human multiple sclerosis disease, is DM dependent. This is also true for presentation of insulin epitope B9–23 to activate autoreactive CD4+ T cells restricted to type 1 diabetes associated HLA-DQ8. Our findings highlighted a fundamental mechanism that immune system differentially regulates DM activity in thymus to balance the efficiency of negative selection to minimize autoimmunity and the maximum of T cell repertoire to defend infection; however, the DM-dependency property of certain autoantigen epitopes accidently caused the failure deletion of autoreactive CD4+ T cells.

CCR4 and CCR7 differentially regulate thymocyte localization with distinct outcomes for central tolerance

Abstract Central tolerance ensures autoreactive thymocytes are either eliminated via clonal deletion or are diverted to the regulatory T-cell lineage, thus minimizing T cell autoimmunity. To undergo central tolerance, thymocytes must enter the medulla to test their TCRs for autoreactivity against the diverse self-antigens displayed by medullary antigen presenting cells (APCs). While CCR7 is known to promote thymocyte medullary entry and negative selection against some self-antigens, our previous studies implicate CCR4 in these processes, raising the question of whether CCR4 and CCR7 play distinct or redundant roles in central tolerance. Here, we use synchronized positive selection assays and 2-photon timelapse microscopy to demonstrate that CCR4 and CCR7 are expressed sequentially after positive selection and promote medullary accumulation and central tolerance of distinct post-positive selection thymocyte subsets. CCR4 is upregulated within hours of positive selection signaling and promotes medullary entry and clonal deletion of immature post-positive selection thymocytes. In contrast, CCR7 is expressed several days later and promotes medullary entry and negative selection of mature thymocytes. Unlike CCR7 ligands, which are expressed by medullary thymic epithelial cells, CCR4 ligands are expressed by hematopoietic APCs, and we find that CCR4 enforces thymocyte tolerance to self-antigens presented by activated thymic APCs. Our findings demonstrate that CCR7 expression is neither necessary nor sufficient for thymocyte medullary entry and support a revised model of central tolerance in which CCR4 and CCR7 promote early and late stages of central tolerance, respectively, via interactions with distinct APC subsets. Supported by a grant from the NIH/NIAID (R01AI104870)

Curative islet and hematopoietic cell transplantation in diabetic mice without toxic bone marrow conditioning.

Mixed hematopoietic chimerism can promote immune tolerance of donor-matched transplanted tissues, like pancreatic islets. However, adoption of this strategy is limited by the toxicity of standard treatments that enable donor hematopoietic cell engraftment. Here, we address these concerns with a non-myeloablative conditioning regimen that enables hematopoietic chimerism and allograft tolerance across fully mismatched major histocompatibility complex (MHC) barriers. Treatment with an αCD117 antibody, targeting c-Kit, administered with Tcell-depleting antibodies and low-dose radiation permits durable multi-lineage chimerism in immunocompetent mice following hematopoietic cell transplant. In diabetic mice, co-transplantation of donor-matched islets and hematopoietic cells durably corrects diabetes without chronic immunosuppression and no appreciable evidence of graft-versus-host disease (GVHD). Donor-derived thymic antigen-presenting cells and host-derived peripheral regulatory Tcells are likely mediators of allotolerance. These findings provide the foundation for safer bone marrow conditioning and cell transplantation regimens to establish hematopoietic chimerism and isletallograft tolerance.

Proteomic identification of MHC class I-associated peptidome derived from non-obese diabetic mouse thymus and pancreas

The peptides repertoire presented to CD8+ T cells by major histocompatibility complex (MHC) class I molecules is referred to as the MHC I-associated peptidome (MIP), which regulates thymus development, peripheral survival and function during lifetime of CD8+ T cells. Type 1 diabetes (T1D) is an organ-specific autoimmune disease caused by pancreatic β cells destruction mediated primarily by autoreactive CD8+ T cells. Non-obese diabetic (NOD) mouse is an important animal model of T1D. Here, we deeply analyzed the MIP derived from NOD mice thymus and pancreas, and demonstrated that the thymus MIP source proteins partially shared with the MIP source proteins derived from NOD mice pancreas and β cell line. One H-2Kd restricted peptide SLC35B126–34 which was shared by MIP derived from both NOD mice pancreatic tissues and islet β-cell line, but absent in MIP from NOD thymus tissues, showed ability to stimulate IFN-γ secretion and proliferation of NOD mice splenic CD8+ T cells. The global view of the MHC I-associated self-peptides repertoire in the thymus and pancreas of NOD mice may serve as a biological reference to identify potential autoantigens targeted by autoreactive CD8+ T cells in T1D. Data are available via ProteomeXchange with identifier PXD031966. SignificanceThe peptides repertoire presented to CD8+ T cells by major histocompatibility complex (MHC) class I molecules is referred to as the MHC I-associated peptidome (MIP). The MIP presented by thymic antigen presenting cells (APCs) is crucial for shaping CD8+ T cell repertoire and self-tolerance, while the MIP presented by peripheral tissues and organs is not only involved in maintaining periphery CD8+ T cell survival and homeostasis, but also mediates immune surveillance and autoimmune responses of CD8+ T cells under pathological conditions. Type 1 diabetes (T1D) is an organ-specific autoimmune disease caused by the destruction of pancreatic β cells, mediated primarily by autoreactive CD8+ T cells. Non-obese diabetic (NOD) mouse is one of important animal models of spontaneous autoimmune diabetes that shares several key features with human T1D. The global view of the MHC I-associated self-peptides repertoire in the thymus and pancreas of NOD mice may serve as a good biological reference to identify potential autoantigens targeted by autoreactive CD8+ T cells in T1D. It has great significance for further clarifying the immune recognition and effect mechanism of autoreactive CD8+ T cells in the pathogenesis of T1D, and then developing antigen-specific immune intervention strategies.

Mechanisms of Direct and Indirect Presentation of Self-Antigens in the Thymus.

The inevitability of evolution of the adaptive immune system with its mechanism of randomly rearranging segments of the T cell receptor (TCR) gene is the generation of self-reactive clones. For the sake of prevention of autoimmunity, these clones must be eliminated from the pool of circulating T cells. This process occurs largely in the thymic medulla where the strength of affinity between TCR and self-peptide MHC complexes is the factor determining thymocyte fate. Thus, the display of self-antigens in the thymus by thymic antigen presenting cells, which are comprised of medullary thymic epithelial (mTECs) and dendritic cells (DCs), is fundamental for the establishment of T cell central tolerance. Whereas mTECs produce and present antigens in a direct, self-autonomous manner, thymic DCs can acquire these mTEC-derived antigens by cooperative antigen transfer (CAT), and thus present them indirectly. While the basic characteristics for both direct and indirect presentation of self-antigens are currently known, recent reports that describe the heterogeneity of mTEC and DC subsets, their presentation capacity, and the potentially non-redundant roles in T cell selection processes represents another level of complexity which we are attempting to unravel. In this review, we underscore the seminal studies relevant to these topics with an emphasis on new observations pertinent to the mechanism of CAT and its cellular trajectories underpinning the preferential distribution of thymic epithelial cell-derived self-antigens to specific subsets of DC. Identification of molecular determinants which control CAT would significantly advance our understanding of how the cellularly targeted presentation of thymic self-antigens is functionally coupled to the T cell selection process.

How Thymocyte Deletion in the Cortex May Curtail Antigen-Specific T-Regulatory Cell Development in the Medulla.

CD4+ T cell responses to self-antigens are pivotal for immunological self-tolerance. Activation of Foxp3– T-conventional (T-conv) cells can precipitate autoimmune disease, whereas activation of Foxp3+ T-regulatory (T-reg) cells is essential to prevent autoimmune disease. This distinction indicates the importance of the thymus in controlling the differentiation of self-reactive CD4+ T cells. Thymocytes and thymic antigen-presenting cells (APC) depend on each other for normal maturation and differentiation. In this Hypothesis and Theory article, we propose this mutual dependence dictates which self-antigens induce T-reg cell development in the thymic medulla. We postulate self-reactive CD4+ CD8– thymocytes deliver signals that stabilize and amplify the presentation of their cognate self-antigen by APC in the thymic medulla, thereby seeding a niche for the development of T-reg cells specific for the same self-antigen. By limiting the number of antigen-specific CD4+ thymocytes in the medulla, thymocyte deletion in the cortex may impede the formation of medullary T-reg niches containing certain self-antigens. Susceptibility to autoimmune disease may arise from cortical deletion creating a “hole” in the self-antigen repertoire recognized by T-reg cells.

Cellular and molecular mediators of thymic DC homeostasis and activation

Abstract Central tolerance in the thymus ensures that autoreactive thymocytes undergo negative selection or diversion to the regulatory T cell (Treg) lineage. Thymic antigen presenting cells (APCs) such as medullary thymic epithelial cells (mTECs) and dendritic cells (DCs) present myriad self-peptides to thymocytes to induce central tolerance. We and others have previously shown that thymic cDCs can be subdivided into activated CCR7+MHC-IIhi and resting CCR7− MHC-IIlo cDC1 and cDC2 subsets with distinct functional properties, suggesting the thymic DC compartment may be more heterogeneous than previously described. Here, we performed single-cell RNA sequencing (scRNAseq) of hematopoietic APCs isolated from adult mouse thymi to comprehensively identify thymic DC subsets with distinct gene expression profiles. Our data reveal 11 transcriptionally distinct cDC subsets. We identify 6 cDC1 subsets, 3 cDC2 subsets, 2 activated CCR7+ “cDC3” subsets distinguished by expression of cDC1 and cDC2 markers, and 4 plasmacytoid DC clusters. Through functional assays, we find that the XCR1+ cDC3s are the most efficient at presenting mTEC-derived peptides on MHC-I and MHC-II, while 2 distinct cDC2 subsets are the most efficient at generating Tregs in vitro. Additional scRNA-seq and flow cytometry analyses of thymic DCs from mice in which thymocyte maturation was blocked at different developmental stages reveal candidate cellular and molecular crosstalk signals that regulate homeostasis and/or activation of pDC and cDC subsets. Our findings reveal hitherto unknown functional heterogeneity within the thymic DC compartment and identify potential interactions that support DC homeostasis and activation which in turn could impact central tolerance. Supported by NIH P01 AI139449-02

The ICOS-ICOSL pathway tunes thymic selection.

Negative selection of developing T cells plays a significant role in T‐cell tolerance to self‐antigen. This process relies on thymic antigen‐presenting cells which express both self‐antigens and cosignaling molecules. Inducible T‐cell costimulator (ICOS) belongs to the CD28 family of cosignaling molecules and binds to ICOS ligand (ICOSL). The ICOS signaling pathway plays important roles in shaping the immune response to infections, but its role in central tolerance is less well understood. Here we show that ICOSL is expressed by subsets of thymic dendritic cells and medullary thymic epithelial cells as well as thymic B cells. ICOS expression is upregulated as T cells mature in the thymus and correlates with T‐cell receptor signal strength during thymic selection. We also provide evidence of a role for ICOS signaling in mediating negative selection. Our findings suggest that ICOS may fine‐tune T‐cell receptor signals during thymic selection contributing to the generation of a tolerant T‐cell population.

Recognition of Multiple Hybrid Insulin Peptides by a Single Highly Diabetogenic T-Cell Receptor.

The mechanisms underlying the major histocompatibility complex class II (MHCII) type 1 diabetes (T1D) association remain incompletely understood. We have previously shown that thymocytes expressing the highly diabetogenic, I-Ag7-restricted 4.1-T-cell receptor (TCR) are MHCII-promiscuous, and that, in MHCII-heterozygous mice, they sequentially undergo positive and negative selection/Treg deviation by recognizing pro- and anti-diabetogenic MHCII molecules on cortical thymic epithelial cells and medullary hematopoietic antigen-presenting cells (APCs), respectively. Here, we use a novel autoantigen discovery approach to define the antigenic specificity of this TCR in the context of I-Ag7. This was done by screening the ability of random epitope–GS linker–I-chain fusion pools to form agonistic peptide–MHCII complexes on the surface of I- chain-transgenic artificial APCs. Pool deconvolution, I-Ag7-binding register-fixing, TCR contact residue mapping, and alanine scanning mutagenesis resulted in the identification of a 4.1-TCR recognition motif XL(G/A)XEXE(D/E)X that was shared by seven agonistic hybrid insulin peptides (HIPs) resulting from the fusion of several different chromogranin A and/or insulin C fragments, including post-translationally modified variants. These data validate a novel, highly sensitive MHCII-restricted epitope discovery approach for orphan TCRs and suggest thymic selection of autoantigen-promiscuous TCRs as a mechanism for the murine T1D–I-Ag7-association.

Murine Roseolovirus Induces Autoimmune Gastritis and Disrupts Thymic Antigen Presenting Cells.

Abstract Murine roseolovirus (MRV) is a beta-herpesvirus that is a natural murine pathogen and is genetically highly related to the human roseoloviruses, HHV6 and HHV7. The human roseoloviruses have been associated with autoimmune disease but demonstrating causality has been difficult due to the ubiquitous and chronic nature of roseolovirus infections. Neonatal MRV infection resulted in transient thymic and peripheral CD4+ T cell depletion, including CD4+Foxp3+ regulatory T cells (Tregs). We found that neonatal MRV infection induced autoimmune gastritis in adult mice. Our studies showed that autoimmune gastritis occurs in the absence of detectable virus in gastric mucosa. Ganciclovir (GCV) inhibited MRV replication as well as MRV-mediated CD4+ T cell depletion and development of autoimmune gastritis. This suggests that MRV replication is necessary for altering immunologic tolerance early in life. Interestingly, neonatal MRV infection resulted in the production of a wide array of autoantibodies in adult mice. The inflammatory infiltrate into the gastric mucosa was characterized by an increase in T cells, neutrophils and eosinophils. Furthermore, the CD4+ T cells in the gastric mucosa exhibited Th2 and Th17 differentiation. The development of autoimmune gastritis was CD4+ T cell and IL-17 dependent, while B cells were only necessary early in life. Neonatal MRV infection resulted in a transient decrease in medullary thymic epithelial and thymic dendritic cells numbers, as well as reduced expression of AIRE and tissue restricted self-antigens in the thymus. These findings strongly suggest that infection with a roseolovirus early in life results in disruption of central tolerance and development of autoimmune disease.

HLA-DR15 Molecules Jointly Shape an Autoreactive T Cell Repertoire in Multiple Sclerosis

SummaryThe HLA-DR15 haplotype is the strongest genetic risk factor for multiple sclerosis (MS), but our understanding of how it contributes to MS is limited. Because autoreactive CD4+ T cells and B cells as antigen-presenting cells are involved in MS pathogenesis, we characterized the immunopeptidomes of the two HLA-DR15 allomorphs DR2a and DR2b of human primary B cells and monocytes, thymus, and MS brain tissue. Self-peptides from HLA-DR molecules, particularly from DR2a and DR2b themselves, are abundant on B cells and thymic antigen-presenting cells. Furthermore, we identified autoreactive CD4+ T cell clones that can cross-react with HLA-DR-derived self-peptides (HLA-DR-SPs), peptides from MS-associated foreign agents (Epstein-Barr virus and Akkermansia muciniphila), and autoantigens presented by DR2a and DR2b. Thus, both HLA-DR15 allomorphs jointly shape an autoreactive T cell repertoire by serving as antigen-presenting structures and epitope sources and by presenting the same foreign peptides and autoantigens to autoreactive CD4+ T cells in MS.

GILT in thymic epithelial cells facilitates central CD4 T cell tolerance to a tissue-restricted, melanoma-associated self antigen

Abstract Central tolerance prevents autoimmunity, but also limits T cell responses to potentially immunodominant tumor epitopes with limited expression in healthy tissues. In peripheral antigen presenting cells (APCs), gamma-interferon-inducible lysosomal thiol reductase (GILT) is critical for MHC class II-restricted presentation of disulfide bond-containing proteins, including the self and melanoma antigen tyrosinase-related protein 1 (TRP1). The role of GILT in thymic antigen processing and generation of central tolerance has not been investigated. We found that GILT enhanced the negative selection of TRP1-specific thymocytes. GILT expression was enriched in thymic APCs capable of mediating deletion, namely medullary thymic epithelial cells (mTECs) and dendritic cells, while TRP1 expression was restricted solely to mTECs. GILT facilitated MHC class II-restricted presentation of endogenous TRP1 by pooled thymic APCs. Using bone marrow chimeras, GILT expression in thymic epithelial cells (TECs), but not hematopoietic cells, was sufficient for complete deletion of TRP1-specific thymocytes. An increased frequency of TRP1-specific regulatory T (Treg) cells was present in chimeras with increased deletion of TRP1-specific thymocytes. Only chimeras that lacked GILT in both TECs and hematopoietic cells had a high conventional T:Treg cell ratio and were protected from melanoma challenge. Thus, GILT expression in thymic APCs, and mTECs in particular, preferentially facilitates MHC class II-restricted presentation, negative selection and increased Treg cells, resulting in a diminished anti-tumor response to a tissue-restricted, melanoma-associated self antigen.

The aged thymus is impaired in supporting T cell central tolerance

Abstract During aging, the thymus undergoes involution, decreasing dramatically in size and mass, and becoming increasingly disorganized. Notably, thymic epithelial cells decline with age, especially within the compartment of mature medullary thymic epithelial cells (mTECs) that express and present tissue-restricted antigens to thymocytes. Age-associated changes in the composition of hematopoietic antigen-presenting cells (APCs) have also been observed, but are not well characterized. Given that both mTECs and APCs present diverse self-antigens to thymocytes, we hypothesized that central tolerance is impaired in the aged thymus. Using live-cell two-photon microscopy, we determined that CD4-single positive (CD4SP) thymocytes, regardless of thymocyte age, had reduced motility and accumulation within the medulla of an aged thymus. We report age-associated changes in both the localization and composition of thymic APC subsets. To test whether this altered thymic environment functionally impacted central tolerance, we used thymic slice assays to measure tolerance induction of CD8− and CD4SP thymocytes encountering a range of model self-antigens. These assays indicated that central tolerance in the aged thymus was impaired, particularly against low-avidity auto-antigens. Our findings indicate that newly generated T cells selected in an aged thymic environment have not undergone a rigorous screening process against autoimmunity.

Recirculating IL-1R2+ Tregs fine-tune intrathymic Treg development under inflammatory conditions.

The vast majority of Foxp3+ regulatory T cells (Tregs) are generated in the thymus, and several factors, such as cytokines and unique thymic antigen-presenting cells, are known to contribute to the development of these thymus-derived Tregs (tTregs). Here, we report the existence of a specific subset of Foxp3+ Tregs within the thymus that is characterized by the expression of IL-1R2, which is a decoy receptor for the inflammatory cytokine IL-1. Detailed flow cytometric analysis of the thymocytes from Foxp3hCD2xRAG1GFP reporter mice revealed that the IL-1R2+ Tregs are mainly RAG1GFP- and CCR6+CCR7-, demonstrating that these Tregs are recirculating cells entering the thymus from the periphery and that they have an activated phenotype. In the spleen, the majority of IL-1R2+ Tregs express neuropilin-1 (Nrp-1) and Helios, suggesting a thymic origin for these Tregs. Interestingly, among all tissues studied, the highest frequency of IL-1R2+ Tregs was observed in the thymus, indicating preferential recruitment of this Treg subset by the thymus. Using fetal thymic organ cultures (FTOCs), we demonstrated that increased concentrations of exogenous IL-1β blocked intrathymic Treg development, resulting in a decreased frequency of CD25+Foxp3+ tTregs and an accumulation of CD25+Foxp3- Treg precursors. Interestingly, the addition of IL-1R2+ Tregs, but not IL-1R2- Tregs, to reaggregated thymic organ cultures (RTOCs) abrogated the IL-1β-mediated blockade, demonstrating that these recirculating IL-1R2+ Tregs can quench IL-1 signaling in the thymus and thereby maintain thymic Treg development even under inflammatory conditions.

Investigating T Cell Receptor Signals In Situ by Two-Photon Microscopy of Thymocytes Expressing Genetic Reporters in Low-Density Chimeras.

T cell development is a dynamic process accompanied by extensive thymocyte migration, cellular interactions, and T cell receptor (TCR) signaling. In particular, thymic selection processes that ensure a functional, self-tolerant repertoire require TCR interactions with self-peptide presented by major histocompatibility complex molecules expressed by specialized thymic antigen-presenting cells. The quantity and quality of these TCR signals influence T cell fate. Two-photon microscopy, which enables live imaging of cells in intact tissue, has emerged as a powerful tool to gain insights into thymocyte migration and TCR signaling during T cell development in situ. Here we describe the generation of non-irradiated, low-density chimeric mice by neonatal injection of adult bone marrow engineered to express fluorescent TCR signaling reporters for imaging by two-photon microscopy. We also describe how the thymic lobes from chimeric mice are prepared for live imaging of thymocyte behavior and TCR signaling events. While we focus on imaging TCR signals associated with T cell development in the thymus, these techniques can also be adapted to study TCR signaling in mature T cells in peripheral lymphoid organs.

Transcriptomes of antigen presenting cells in human thymus.

Antigen presenting cells (APCs) in the thymus play an essential role in the establishment of central tolerance, i.e. the generation of a repertoire of functional and self-tolerant T cells to prevent autoimmunity. In this study, we have compared the transcriptomes of four primary APCs from human thymus (mTECs, CD19+ B cells, CD141+ and CD123+ DCs). We investigated a set of genes including the HLA genes, genes encoding transcriptional regulators and finally, tissue-enriched genes, i.e, genes with a five-fold higher expression in a particular human tissue. We show that thymic CD141+ DCs express the highest levels of all classical HLA genes and 67% (14/21) of the HLA class I and II pathway genes investigated in this study. CD141+ DCs also expressed the highest levels of the transcriptional regulator DEAF1, whereas AIRE and FEZF2 expression were mainly found in primary human mTECs. We found expression of “tissue enriched genes” from the Human Protein Atlas (HPA) in all four APC types, but the mTECs were clearly dominating in the number of uniquely expressed tissue enriched genes (20% in mTECs, 7% in CD19+ B cells, 4% in CD123+ DCs and 2% in CD141+ DCs). The tissue enriched genes also overlapped with reported human autoantigens. This is, to our knowledge, the first study that performs RNA sequencing of mTECs, CD19+ B cells, CD141+ and CD123+ DCs isolated from the same individuals and provides insight into the transcriptomes of these human thymic APCs.

Perturbation of Thymocyte Development Underlies the PRRS Pandemic: A Testable Hypothesis.

Porcine reproductive and respiratory syndrome virus (PRRSV) causes immune dysregulation during the Critical Window of Immunological Development. We hypothesize that thymocyte development is altered by infected thymic antigen presenting cells (TAPCs) in the fetal/neonatal thymus that interact with double-positive thymocytes causing an acute deficiency of T cells that produces “holes” in the T cell repertoire allowing for poor recognition of PRRSV and other neonatal pathogens. The deficiency may be the result of random elimination of PRRSV-specific T cells or the generation of T cells that accept PRRSV epitopes as self-antigens. Loss of helper T cells for virus neutralizing (VN) epitopes can result in the failure of selection for B cells in lymph node germinal centers capable of producing high affinity VN antibodies. Generation of cytotoxic and regulatory T cells may also be impaired. Similar to infections with LDV, LCMV, MCMV, HIV-1 and trypanosomes, the host responds to the deficiency of pathogen-specific T cells and perhaps regulatory T cells, by “last ditch” polyclonal B cell activation. In colostrum-deprived PRRSV-infected isolator piglets, this results in hypergammaglobulinemia, which we believe to be a “red herring” that detracts attention from the thymic atrophy story, but leads to our second independent hypothesis. Since hypergammaglobulinemia has not been reported in PRRSV-infected conventionally-reared piglets, we hypothesize that this is due to the down-regulatory effect of passive maternal IgG and cytokines in porcine colostrum, especially TGFβ which stimulates development of regulatory T cells (Tregs).