Medullary Thymic Epithelial Cells (mTECs)
mTECs are cells of the thymic medulla. Unlike most other antigen-presenting cells (APCs), mTECs express MHC class I and MHC class II molecules. They employ noncanonical peptide-loading pathways, mainly originating from intracellular sources. A unique feature of mTECs is their ability to express self-antigens typically characteristic of other tissues in the body, thereby teaching the immune system to recognize host tissues without attacking them. This process is called promiscuous gene expression (PGE), and the transcription factor AIRE plays a critical regulatory role.
Promiscuous Gene Expression Is Less Random Than Previously Assumed
For a long time, it was unclear whether the expression of antigens corresponding to one or several peripheral tissues (tissue-restricted antigens, TRA) in individual mTECs was random or followed specific patterns. Recent studies show that patterns of TRA expression recur, with each antigen group restricted to specific mTEC subsets. This phenomenon may relate to temporary developmental stages in mTECs or their differentiation. Adding more significance to this hypothesis, new data indicate substantial diversity among specialized mTEC types.
Unexpected Diversity Among mTECs
Based on MHC class II expression differences, mTECs can be subdivided into immature (mTEClow) and mature (mTEChi) populations. Approximately half of the mature mTECs express AIRE (AIRE+ mTEChi) and were once thought to represent the final stage of mTEC development, prone to apoptosis. However, more recent studies revealed the existence of post-AIRE cells that lose expression of AIRE and MHC class II, making them indistinguishable from immature mTECs. Additionally, a newly identified subset of mTECs—comprising 5–10% of all such cells—displays traits resembling various peripheral tissues: intestinal tuft cells, gut and liver epithelium, muscle, and endocrine cell types.
These so-called “mimetic” cells exhibit properties akin to the tissues they mimic. Many—if not all—mimetic cells share features with post-AIRE mTECs. It is thus tempting to propose that at least some AIRE+ mTEChi cells represent transitional developmental stages destined for subsequent differentiation into mimetic cells. Although mimetic cells express low levels of MHC class II, they can still participate in antigen presentation through other APCs, such as dendritic cells. For instance, mice engineered to express artificial antigens, specifically in mimetic mTECs, displayed tolerance to those antigens. Conversely, mice that lacked “endocrine mTEC” developed serum autoantibodies against endocrine-rich tissues such as the thyroid gland and stomach. Besides their role in immunological tolerance, mimetic mTECs influence innate lymphoid cell homeostasis and age-related thymic changes.
Thymic Dendritic Cells
CD11c+ thymic cells include plasmacytoid dendritic cells and conventional dendritic cells (cDC), which subdivide into two subtypes: cDC1 (SIRPα–CD8+XCR1+) and cDC2 (SIRPα+CD8–XCR1–). These subtypes mirror the cDC1–cDC2 division described in secondary lymphoid organs. The cDC1 population develops locally in the thymus and is hence regarded as a resident cell, whereas cDC2 are believed to migrate from peripheral sites, though this premise requires further clarification. Most cDCs reside in the thymic medulla. Owing to limited data on plasmacytoid and monocyte-derived dendritic cells in the thymus, the discussion here focuses on cDC1 and cDC2 subtypes.
Presentation of Circulating Antigens
Research suggests cDC2 is more efficient at capturing blood-borne antigens than cDC1, though the magnitude of these differences may vary by experimental context and antigen properties. Small molecules can passively diffuse through vessel barriers, thus reaching dendritic cells, whereas large molecules may not. Recent work has identified transendothelial thymic dendritic cells lying near medullary microvessels. These cells extend processes across the endothelium into the bloodstream. They primarily bear cDC2 characteristics and are permanently situated at the interface between the thymus and the blood. Transendothelial thymic dendritic cells play a crucial role in capturing blood-derived antigens and are engaged in the adverse selection of thymocytes—essential for establishing central tolerance. Similar functionalities have been observed with cDC2 in the cerebral cortex, where they cluster around blood vessels. cDC2 is especially adept at presenting endocytosed antigens to CD4+ T-cells via MHC class II, while cDC1 is superior at cross-presenting antigens to CD8+ T-cells.
Cooperation Between cDC and mTECs
Antigens produced by mTECs may be presented directly by the same mTECs or transferred to thymic dendritic cells for presentation. Evidence indicates that antigen transfer occurs unidirectionally—from mTECs to cDCs but not vice versa. In particular, cDC1 actively acquires mTEC-derived material, while cDC2 participates equally when all TECs uniformly express antigens. A crucial element in antigen transfer is the chemokine axis XCR1–XCL1: cDC1 expresses the XCR1 receptor, and mTEChi secrete the chemokine XCL1, which attracts cDC1 into proximity with AIRE+ mTECs. Among cDC1, only mature CCR7+ cells effectively acquire and present antigens from mTECs, suggesting that both XCL1 and other chemokines (CCL19, CCL21) cooperate to facilitate the passage of mTEC-derived antigens to mature cDC1. Though the mechanics of these events remain incompletely defined, researchers propose that various dendritic cell subtypes interact with specific mTEC subsets, capturing antigens from distinct sources. Given that these cells lack typical APC functions, It remains uncertain how mimetic mTECs transfer their antigens.
cDC1 or cDC2: Division of Labor or Redundancy?
Different dendritic cell subtypes may present different sets of self-antigens. If each type performs unique duties in central tolerance, the absence of one subtype should theoretically alter the fate of autoreactive thymocytes and the shape of the T-cell repertoire. However, evidence shows that cDC1 deficiency has minimal impact on the CD8+ T-cell or CD4+ Treg repertoire and the scale of adverse thymocyte selection—suggesting a degree of functional redundancy. At the same time, other studies indicate that cDC1 is critical for presenting mTEC-derived antigens and stimulating Treg cells. Investigations into cDC2 remain challenging without methods to remove them entirely. Partial cDC2 depletion (about 2/3) has been shown to delay the elimination of CD4+ T-cells but does not markedly shift the Treg-cell repertoire. Multiple APC types present the same antigens, imparting substantial redundancy. Indeed, when an antigen is membrane-bound, cDC1, cDC2, and mTEC contribute to deleting CD4+ and CD8+ T-cells. If mTECs secrete an antigen, cDC primarily handles its presentation after the antigen diffuses within the tissue fluid. Surprisingly, CD4+ T-cells and CD8+ T-cells generally rely on cDC2 despite earlier suggestions that cDC1 has a superior cross-presentation mechanism. In that specific experimental model, the speed of antigen uptake possibly overshadowed differences in cross-presentation proficiency.
Thymic B Cells
Thymic B cells are as frequent as dendritic cells in the thymus. Unlike resting peripheral B cells, they express high levels of MHC class II and co-stimulatory molecules. Approximately one-third of thymic B cells carry class-switched immunoglobulins. Thymic B cells resemble germinal center-like cells or precursors to memory B cells. A unique feature of thymic B cells is the expression of AIRE, which is absent in peripheral B cells. Although only 2% of thymic B cells express AIRE, its influence on their functions warrants further investigation. Thymic B cells may contribute to T-cell tolerance.
Thymic B Cells and Immunological Tolerance
Thymic B cells play a crucial role in central tolerance. MHC class II expression exclusively on B cells is sufficient for deleting superantigen-reactive T cells and the adverse selection of thymocytes. In B-cell-deficient mice, the number of CD4SP cells increases while the frequency of thymic Treg cells decreases. However, the nature of the autoantigens presented by thymic B cells remains unclear.
Autoantigen Capture by the B-Cell Receptor (BCR)
BCR-mediated capture of autoantigens enables thymic B cells to present tolerogenic antigens effectively. When thymic B cells express BCRs specific for an autoantigen, they more efficiently mediate the adverse selection of CD4+ T cells recognizing that antigen. Clonal expansion occurs in thymic B cells; some become autoreactive or polyreactive, which may result from intrinsic developmental processes in the thymus rather than the recruitment of autoreactive cells from the periphery.
Class Switching and Tolerogenicity of Thymic B Cells
Interestingly, while various immunoglobulin types circulate in large quantities, thymic B cells uniquely present epitopes derived from immunoglobulins, potentially due to their enhanced ability to load epitopes onto MHC class II, a process amplified upon B-cell activation. Studies in mice predisposed to type 1 diabetes suggest that impaired class switching in thymic B cells compromises central tolerance, increasing diabetes incidence.
Endogenous Autoantigens in Thymic B Cells
Over 70% of peptides associated with MHC class II on thymic B cells are derived from proteins produced within the cell. This suggests that the pMHC II ligandome in thymic B cells is enriched with autoantigens. AIRE may contribute to shaping this ligandome, though its impact is likely limited due to the small number of genes it activates. Thymic B cells resemble memory and germinal center B cells, with epitopes predominantly formed from endogenously activated autoantigens. These cells may mimic the germinal center B-cell ligandome, removing specific T cells from the repertoire and directing autoantigen-specific CD4+ T cells toward the T follicular helper compartment.
Broader Autoantigen Tolerance via Thymic B Cells
Recent findings suggest that thymic B cells are essential in maintaining tolerance to autoantigens, such as aquaporin-4 (AQP4), linked to autoimmune diseases like neuromyelitis optica. Although AQP4 is not a typical autoantigen for thymic B cells, its deletion in B cells of mice disrupts T-cell tolerance to this antigen, leading to a pathology resembling neuromyelitis optica.
Crosstalk with Autoreactive Thymocytes
The functions of APCs such as mTECs, cDCs, and thymic B cells are not dictated by rigid differentiation programs but emerge through dynamic interactions with developing thymocytes and other cells. Recent research highlights shared mechanisms governing the phenotypic adaptation of cDCs and thymic B cells.
Homeostatic Maturation of cDCs
Unlike peripheral dendritic cells, both cDC1 and cDC2 in the thymus contain a significant proportion of mature cells expressing high levels of MHCII and CD86. This phenotype persists in germ-free mice and does not rely on innate adaptor proteins, suggesting a “sterile” maturation process. cDC maturation in the thymus depends on CD40 signaling and interactions with autoreactive CD4SP thymocytes. Notably, transcriptional programs governing inflammatory and homeostatic maturation of cDCs are similar. Unlike microbe-stimulated cDCs, mature thymic cDCs retain antigen uptake and MHCII synthesis abilities, enhancing their capacity to present autoantigens.
Licensing Thymic B Cells
Like mTECs and cDCs, thymic B cells undergo reprogramming regulated by CD40 signals during interactions with autoreactive CD4SP thymocytes. This licensing process activates MHCII and co-stimulatory molecules while inducing AIRE expression. Thymic B cells may originate from intrathymic progenitors or migrate from the periphery, with both populations acquiring a licensed phenotype. Licensing of thymic B cells shares similarities with germinal center responses, including MHCII activation, CD80 expression, and immunoglobulin class switching. Although the thymus lacks true germinal centers, these findings support the hypothesis that thymic B cells represent a tolerogenic mirror of germinal center B cells.
Functional Programming via Tonic Inflammation
Through interactions with autoreactive thymocytes, mTECs, cDCs, and thymic B cells acquire enhanced APC functions. Each cell type exhibits unique features, but all utilize inflammatory signaling pathways. These pathways enable thymic APCs to develop tolerogenic properties while diversifying their pMHC ligandomes, incorporating autoantigens that could otherwise drive peripheral tissue inflammation.
Tuning mTEC via Toll-Like Receptor Signaling and Type III Interferon
mTECs express Toll-like receptors (TLRs), which fine-tune immune processes. Deleting the MyD88 signaling adaptor in these cells reduces the production of chemokines that recruit dendritic cells, altering the composition of thymic DCs and weakening antigen transfer from mTECs. It also diminishes Treg-cell generation, thereby affecting CD4+ T-cell composition. TLR signaling is pivotal for driving AIRE+ mTECs toward their terminal phenotype—progressing to post-AIRE keratinocyte-like mTECs. TLR activation likely derives from endogenous ligands. Since mTECs most prominently express TLR9, which detects “foreign” DNA, it is plausible that endogenous DNA released by apoptotic thymocytes can suffice to trigger TLR signals. Type III Interferon (IFN-III) enhances the antigen-presentation capacity of mTECs by boosting MHC class I expression. mTECs also produce IFN-III, further elevating their ability to establish CD8+ T-cell tolerance.
Activation of cDC2 by Type 2 Cytokines
The majority of thymic cDC2 cells, regardless of their maturation stage, express the activation marker CD301b and exhibit signs of exposure to type 2 cytokines such as IL-4, IL-5, IL-9, and IL-13. In mice lacking IL-4 and IL-13 cytokines or invariant natural killer T cells producing IL-4, the number of activated cDC2 cells is significantly reduced, which indicates the additive effects of IL-4 and IL-13 in cDC2 activation.
Future research should explore how genetic background influences the thymic cDC2 population and their ability to maintain tolerance. Type 2 cytokines are believed to enhance cDC2-mediated tolerance and contribute to the formation of pMHC ligandomes, enriching them with autoantigens associated with inflammatory responses in peripheral tissues.
B-Cell Modulation via Cytokines in the Thymus
Thymic B-cells display a marked interferon signature distinguishing them from germinal center B-cells in secondary lymphoid organs. Ablating the IFN-III receptor severely reduces the licensed thymic B-cell population, impairing Treg-cell formation in the thymus. This effect mirrors the phenotype of complete B-cell deficiency. It is unclear how interferon modulates licensed thymic B-cells—whether by modulating their survival, proliferation, activation, or class switching. However, class-switch regulation is likely orchestrated by IL-4, TGFβ, and IFN-γ signals. Recent work has identified a minor population of IgE+ plasma cells in the thymus of BALB/c mice. These cells require IL-4 secreted by invariant natural killer T-cells and serve as a source of IgE. Thymic plasma cells form locally, bypassing a germinal center reaction, and express AIRE, suggesting they may arise from licensed B-cells. Analogous cells have been found in newborn humans, providing early innate humoral immunity.
Conclusions
Single-cell–scale technologies have greatly expanded our understanding of antigen-presenting cells in the thymus. The discovery of mimetic mTECs adds new complexity to thymic biology. The role of thymic B-cells remains a double-edged sword. On the one hand, their capacity to display self-antigens supports central tolerance induction; on the other hand, aberrant B-cell differentiation into plasma cells may fuel autoimmunity (e.g., in myasthenia gravis or lupus). Studies show that in patients with myasthenia gravis, thymic changes involving structures resembling germinal centers might cause or consequence of disease. Age also shapes thymic function. In early life, unique Treg populations are established, whereas in adolescence, adverse selection intensifies, possibly hindering the emergence of Treg-cells bearing the same receptors later on. With aging, thymic tolerance weakens: PGE in mTECs declines, thymocyte motility may wane (limiting their antigen encounter), and the licensed B-cell population is reduced. These changes can promote age-related autoimmunity. Whether “rejuvenation” strategies such as stem cell transplantation can completely restore the thymus’ tolerogenic capacity remains an open question.
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Reference
Antigen presentation for central tolerance induction
