scholarly journals P03.07 Analysis of scRNAseq from the human thymus nominates genes potentially missing from central tolerance of cytotoxic T cells

2021 ◽  
Vol 9 (Suppl 1) ◽  
pp. A15.2-A16
Author(s):  
L Blumenberg ◽  
G Atwal ◽  
A Dhanik
Keyword(s):  
T Cells ◽  
T Cell ◽  
Negative Selection ◽  
Cytotoxic T Cells ◽  
Peripheral Tolerance ◽  
Thymic Epithelial Cells ◽  
Central Tolerance ◽  
Human Thymus ◽  
Part Time ◽  
Self Antigens

BackgroundDuring thymic development, cytotoxic T cells that can bind to and attack self antigens undergo negative selection thus preventing damage to the self tissues. The sparse medullar thymic epithelial cells (mTECs) present in the thymus are responsible for presenting self antigens to T cells so that they can trigger apoptosis or differentiation into non-cytotoxic lineages if they bind too strongly.Materials and MethodsUnderstanding gene expression in mTECs is essential for understanding the shape of the human T cell receptor repertoire, which is key for current and emerging cancer immunotherapies. Recent availability of human thymus single cell RNAseq (scRNAseq) data provides an extremely high-resolution view into the pattern of expression within this critical cell type. To determine which epitopes have had to opportunity to be presented during T cell negative selection, we analyzed the human thymus scRNAseq dataset to establish which genes are expressed in mTECs and therefore subject to central tolerance.ResultsThe coverage of the whole transcriptome of a particular cell is generally sparse. It is therefore difficult to understand basic features of individual cells or cell types such as how many genes are expressed. We used cell- and read-level subsampling to estimate whether a sufficient number of cells and reads had been captured to support categorizing a gene as non-expressed in mTECs. We also examined the expression of the genes not expressed in mTECs in other healthy tissues, and found their expression was almost exclusively restricted to the testis (an immune-privileged site) and the liver (a site of peripheral tolerance)ConclusionsAltogether, these analyses establish a strategy for determining if a data set has sufficient depth to estimate the total number of genes expressed and secondly define a key list of genes that are not expressed during central tolerization of T cells, which represent a compelling list of possible cancer immunotherapy targets.Disclosure InformationL. Blumenberg: A. Employment (full or part-time); Significant; Regeneron Pharmaceuticals. G. Atwal: A. Employment (full or part-time); Significant; Regeneron Pharmaceuticals. A. Dhanik: A. Employment (full or part-time); Significant; Regeneron Pharmaceuticals.

scholarly journals Clonal deletion induced by either radioresistant thymic host cells or lymphohemopoietic donor cells at different stages of class I-restricted T cell ontogeny.

1992 ◽  
Vol 175 (5) ◽  
pp. 1277-1283 ◽  
Author(s):  
D E Speiser ◽  
H Pircher ◽  
P S Ohashi ◽  
D Kyburz ◽  
H Hengartner ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Positive Selection ◽  
Negative Selection ◽  
Host Cells ◽  
Thymic Epithelial Cells ◽  
Clonal Deletion ◽  
Mhc Molecules ◽  
Donor Cells ◽  
Self Antigens

Major histocompatibility complex (MHC) products and self-antigens expressed in the thymus determine the repertoire of mature alpha/beta T cells. While positive selection of self-MHC-restricted T cells is directed by MHC molecules expressed by thymic epithelial cells, negative selection depends to a large extent on self-antigens presented by lymphohemopoietic cells. However, radioresistant components of the thymus also influence negative selection, but it remains controversial whether this is accomplished by clonal deletion, clonal anergy, or other mechanisms. In this study, T cell development in mice expressing a transgenic T cell receptor (TCR) specific for lymphocytic choriomeningitis virus (LCMV) plus H-2Db was analyzed in the presence or absence of the viral antigen. A novel approach to analyze the thymic tissue requirements for negative selection was possible by comparing thymocyte selection in H-2Db versus H-2Dbm13 mice, since the latter allowed positive selection but not LCMV-specific deletion of transgenic TCR-expressing thymocytes. In irradiation bone marrow chimeras expressing the restriction element for negative selection (H-2Db) on host tissue, we show that radioresistant recipient cells in the thymus deleted developing T cells at an early stage of differentiation. In contrast, chimeras expressing H-2Db on lymphohemopoietic donor cells showed clonal deletion at a later stage during ontogeny.

scholarly journals Age-Related Changes in Thymic Central Tolerance

2021 ◽  
Vol 12 ◽  
Author(s):  
Jayashree Srinivasan ◽  
Jessica N. Lancaster ◽  
Nandini Singarapu ◽  
Laura P. Hale ◽  
Lauren I. R. Ehrlich ◽  
...  
Keyword(s):  
T Cells ◽  
Negative Selection ◽  
Thymic Epithelial Cells ◽  
Central Tolerance ◽  
Adult Transition ◽  
Treg Function ◽  
Age Related ◽  
And Function ◽  
Antigen Presenting ◽  
Age Related Changes

Thymic epithelial cells (TECs) and hematopoietic antigen presenting cells (HAPCs) in the thymus microenvironment provide essential signals to self-reactive thymocytes that induce either negative selection or generation of regulatory T cells (Treg), both of which are required to establish and maintain central tolerance throughout life. HAPCs and TECs are comprised of multiple subsets that play distinct and overlapping roles in central tolerance. Changes that occur in the composition and function of TEC and HAPC subsets across the lifespan have potential consequences for central tolerance. In keeping with this possibility, there are age-associated changes in the cellular composition and function of T cells and Treg. This review summarizes changes in T cell and Treg function during the perinatal to adult transition and in the course of normal aging, and relates these changes to age-associated alterations in thymic HAPC and TEC subsets.

scholarly journals The lymphoreticular system in triggering virus plus self-specific cytotoxic T cells: evidence for T help.

1978 ◽  
Vol 147 (3) ◽  
pp. 897-911 ◽  
Author(s):  
R M Zinkernagel ◽  
G N Callahan ◽  
A Althage ◽  
S Cooper ◽  
J W Streilein ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
T Cell ◽  
Cytotoxic T Cells ◽  
Phenotypic Expression ◽  
Cell Activity ◽  
Thymic Epithelial Cells ◽  
Cytotoxic T Cell ◽  
Immunological Interactions

The thymus determines the spectrum of the receptor specificities of differentiating T cells for self-H-2; however, the phenotypic expression of T cell's specificity for self plus virus is determined predominantly by the H-2 type of the antigen presenting cells of the peripheral lymphoreticular system. Furthermore, virus specific helper T cells are essential for the generation of virus-specific cytotoxic T cells. For cooperation between mature T cells and other lymphocytes to be functional in chimeras, thymic epithelial cells and lymphohemopoietic stem cells must share the I region; killer T-cell generation also requires in addition compatibility for at least one K or D region. These conclusions derive from the following experiments: A leads to (A X B)F1 chimeric lymphocytes do produce virus-specific cytotoxic T-cell activity for infected A but not for infected B cells; when sensitized in an acutely irradiated and infected recipient (A X B)F1 these chimeric lymphocytes respond to both infected A and B. Therefore the predominantly immunogenically infected cells of chimeras the radiosensitive and by donor stem cells replaced lymphoreticular cells. In this adoptive priming model (KAIA/DB leads to KAIA/DC) chimeric lymphocytes could be sensitized in irradiated and infected F1 against KA and DC but not against infected DB targets. In contrast KBIB/DA leads to KCIC/DA chimeras' lymphocytes could not be sensitized at all in appropriately irradiated and infected F1 recipients. Thus these latter chimeras probably lack functional I-specific T helper cells that are essential for the generation of T killer cells against infected D compatible targets. If T cells learn in the thymus to recognize H-21 or K, D markers that are not at least partially carried themselves in other cells of the lymphoreticular system immunological interactions will be impossible and this paradox situation results in phenotypic immune incompetence in vivo.

scholarly journals CCR4 promotes medullary entry and thymocyte–dendritic cell interactions required for central tolerance

2015 ◽  
Vol 212 (11) ◽  
pp. 1947-1965 ◽  
Author(s):  
Zicheng Hu ◽  
Jessica N. Lancaster ◽  
Chayanit Sasiponganan ◽  
Lauren I.R. Ehrlich
Keyword(s):  
T Cells ◽  
T Cell ◽  
Positive Selection ◽  
Time Lapse ◽  
Cell Receptor ◽  
Peripheral Tolerance ◽  
Central Tolerance ◽  
Self Tolerance ◽  
Thymic Medulla ◽  
Antigen Presenting

Autoimmunity results from a breakdown in central or peripheral tolerance. To establish central tolerance, developing T cells must enter the thymic medulla, where they scan antigen-presenting cells (APCs) displaying a diverse array of autoantigens. If a thymocyte is activated by a self-antigen, the cell undergoes either deletion or diversion into the regulatory T cell (T reg) lineage, thus maintaining self-tolerance. Mechanisms promoting thymocyte medullary entry and interactions with APCs are incompletely understood. CCR4 is poised to contribute to central tolerance due to its expression by post-positive selection thymocytes, and expression of its ligands by medullary thymic dendritic cells (DCs). Here, we use two-photon time-lapse microscopy to demonstrate that CCR4 promotes medullary entry of the earliest post-positive selection thymocytes, as well as efficient interactions between medullary thymocytes and DCs. In keeping with the contribution of thymic DCs to central tolerance, CCR4 is involved in regulating negative selection of polyclonal and T cell receptor (TCR) transgenic thymocytes. In the absence of CCR4, autoreactive T cells accumulate in secondary lymphoid organs and autoimmunity ensues. These studies reveal a previously unappreciated role for CCR4 in the establishment of central tolerance.

scholarly journals Autoreactive T cells bypass negative selection and respond to self-antigen stimulation during infection

2012 ◽  
Vol 209 (10) ◽  
pp. 1769-1779 ◽  
Author(s):  
Sarah Enouz ◽  
Lucie Carrié ◽  
Doron Merkler ◽  
Michael J. Bevan ◽  
Dietmar Zehn
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Negative Selection ◽  
Cell Activation ◽  
Organ Damage ◽  
Peptide Ligands ◽  
Peripheral Tolerance ◽  
High Avidity ◽  
Transgenic Cells

Central and peripheral tolerance prevent autoimmunity by deleting the most aggressive CD8+ T cells but they spare cells that react weakly to tissue-restricted antigen (TRA). To reveal the functional characteristics of these spared cells, we generated a transgenic mouse expressing the TCR of a TRA-specific T cell that had escaped negative selection. Interestingly, the isolated TCR matches the affinity/avidity threshold for negatively selecting T cells, and when developing transgenic cells are exposed to their TRA in the thymus, only a fraction of them are eliminated but significant numbers enter the periphery. In contrast to high avidity cells, low avidity T cells persist in the antigen-positive periphery with no signs of anergy, unresponsiveness, or prior activation. Upon activation during an infection they cause autoimmunity and form memory cells. Unexpectedly, peptide ligands that are weaker in stimulating the transgenic T cells than the thymic threshold ligand also induce profound activation in the periphery. Thus, the peripheral T cell activation threshold during an infection is below that of negative selection for TRA. These results demonstrate the existence of a level of self-reactivity to TRA to which the thymus confers no protection and illustrate that organ damage can occur without genetic predisposition to autoimmunity.

scholarly journals Tumor Evasion from T Cell Surveillance

2011 ◽  
Vol 2011 ◽  
pp. 1-19 ◽  
Author(s):  
Katrin Töpfer ◽  
Stefanie Kempe ◽  
Nadja Müller ◽  
Marc Schmitz ◽  
Michael Bachmann ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Immune Surveillance ◽  
Tumor Burden ◽  
Peripheral Tolerance ◽  
Neoplastic Progression ◽  
Large Tumor ◽  
Central Tolerance ◽  
Intact Immune System

An intact immune system is essential to prevent the development and progression of neoplastic cells in a process termed immune surveillance. During this process the innate and the adaptive immune systems closely cooperate and especially T cells play an important role to detect and eliminate tumor cells. Due to the mechanism of central tolerance the frequency of T cells displaying appropriate arranged tumor-peptide-specific-T-cell receptors is very low and their activation by professional antigen-presenting cells, such as dendritic cells, is frequently hampered by insufficient costimulation resulting in peripheral tolerance. In addition, inhibitory immune circuits can impair an efficient antitumoral response of reactive T cells. It also has been demonstrated that large tumor burden can promote a state of immunosuppression that in turn can facilitate neoplastic progression. Moreover, tumor cells, which mostly are genetically instable, can gain rescue mechanisms which further impair immune surveillance by T cells. Herein, we summarize the data on how tumor cells evade T-cell immune surveillance with the focus on solid tumors and describe approaches to improve anticancer capacity of T cells.

scholarly journals 255 Investigating VISTA’s role intrinsic to T cells in the tumor microenvironment

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A276-A276
Author(s):  
Cassandra Gilmour ◽  
Li Wang ◽  
Juan Dong ◽  
Sarah Stone ◽  
Keman Zhang ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Microenvironment ◽  
Tumor Immunity ◽  
Patient Survival ◽  
Cytotoxic T Cells ◽  
Peripheral Tolerance ◽  
Checkpoint Blockade

BackgroundCancer immunotherapies, specifically checkpoint blockade therapies, have demonstrated clinical importance for long term patient survival. One of the major limitations to checkpoint blockade therapies, is the low response rate: ~30% with anti-CTLA4 and anti-PD1 treatment. This may be due to heterogeneity of the patients‘ immune system and the tumor microenvironment including T cell inhibitions. There is a clear need to study this phenomenon and develop additional therapies for long term survival to include a broad range of patients. V-domain Immunoglobulin Suppressor of T-cell Activation (VISTA) is a suppressive protein expressed on many cell types in the tumor microenvironment including cytotoxic T cells. VISTA’s role on T cells has been described as maintaining quiescence and peripheral tolerance in a graft vs host disease model, but is not fully understood in context of the tumor microenvironment.MethodsWe use a series of invivo experiments, including T cell specific VISTA knock outs, to understand the role of VISTA on T cells in the tumor microenvironment.ResultsHere we show a series of in vivo experiments that suggest VISTA has a potent intrinsic role on T cells and therefore anti-tumor immunity. Using a T cell specific VISTA knock out, our results suggest that the absence of VISTA on T cells in combination with anti-CTLA4 and vaccine is a very powerful tumor suppressor compared to vaccine and anti-CTLA4 treatment alone. These results also indicate that the absence of VISTA alters the phenotype of cytotoxic T cells in several ways including the production of inflammatory cytokines.ConclusionsOur preliminary data provides foundation to study VISTA’s role intrinsic to T cells in the tumor microenvironment and how disrupting VISTA’s influence intrinsic to T cells may be advantageous for anti-tumor immunity and long term patient survival.Ethics ApprovalAll in vivo studies were reviewed and approved by Institutional Animal Care and Use Committee (Approval number 2019–2142).

scholarly journals Thymic Epithelial Cell-Derived IL-15 and IL-15 Receptor α Chain Foster Local Environment for Type 1 Innate Like T Cell Development

2021 ◽  
Vol 12 ◽  
Author(s):  
Huishan Tao ◽  
Lei Li ◽  
Nan-Shih Liao ◽  
Kimberly S. Schluns ◽  
Shirley Luckhart ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Development ◽  
Local Environment ◽  
Cell Development ◽  
Thymic Epithelial Cells ◽  
Thymic Epithelial Cell ◽  
Central Tolerance ◽  
Selection Of

Expression of tissue-restricted antigens (TRAs) in thymic epithelial cells (TECs) ensures negative selection of highly self-reactive T cells to establish central tolerance. Whether some of these TRAs could exert their canonical biological functions to shape thymic environment to regulate T cell development is unclear. Analyses of publicly available databases have revealed expression of transcripts at various levels of many cytokines and cytokine receptors such as IL-15, IL-15Rα, IL-13, and IL-23a in both human and mouse TECs. Ablation of either IL-15 or IL-15Rα in TECs selectively impairs type 1 innate like T cell, such as iNKT1 and γδT1 cell, development in the thymus, indicating that TECs not only serve as an important source of IL-15 but also trans-present IL-15 to ensure type 1 innate like T cell development. Because type 1 innate like T cells are proinflammatory, our data suggest the possibility that TEC may intrinsically control thymic inflammatory innate like T cells to influence thymic environment.

scholarly journals The thymus and central tolerance

2001 ◽  
Vol 356 (1409) ◽  
pp. 609-616 ◽  
Author(s):  
Jonathan Sprent ◽  
Hidehiro Kishimoto
Keyword(s):  
T Cells ◽  
Cell Differentiation ◽  
T Cell ◽  
Tolerance Induction ◽  
Cell Receptor ◽  
Immunological Tolerance ◽  
T Cell Differentiation ◽  
Central Tolerance ◽  
Mhc Molecules ◽  
Self Antigens

T–cell differentiation in the thymus generates a peripheral repertoire of mature T cells that mounts strong responses to foreign antigens but is largely unresponsive to self–antigens. This state of specific immunological tolerance to self–components involves both central and peripheral mechanisms. Here we review the process whereby many T cells with potential reactivity for self–antigens are eliminated in the thymus during early T–cell differentiation. This process of central tolerance (negative selection) reflects apoptosis and is a consequence of immature T cells receiving strong intracellular signalling through T–cell receptor (TCR) recognition of peptides bound to major histocompatibility complex (MHC) molecules. Central tolerance occurs mainly in the medullary region of the thymus and depends upon contact with peptide–MHC complexes expressed on bone–marrow–derived antigen–presenting cells (APCs); whether tolerance also occurs in the cortex is still controversial. Tolerance induction requires a combination of TCR ligation and co–stimulatory signals. Co–stimulation reflects interaction between complementary molecules on T cells and APCs and probably involves multiple molecules acting in consort, which may account for why deletion of individual molecules with known or potential co–stimulatory function has little or no effect on central tolerance. The range of self–antigens that induce central tolerance is considerable and, via low–level expression in the thymus, may also include tissue–specific antigens; central tolerance to these latter antigens, however, is likely to be limited to high–affinity T cells, leaving low–affinity cells to escape. Tolerance to alloantigens and the possibility of using central tolerance to promote acceptance of allografts are discussed.

Characterizing and Augmenting Peripheral Tolerance in in Utero Hematopoietic Cell Transplantation

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4540-4540
Author(s):  
John Samuel Riley ◽  
Lauren E. McClain ◽  
Grace Lee ◽  
Haiying Li ◽  
Alan W. Flake ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Regulatory T Cell ◽  
Hematopoietic Cell Transplantation ◽  
Peripheral Tolerance ◽  
Chimeric Mice ◽  
Central Tolerance ◽  
Tr1 Cells

Abstract Background: In utero hematopoietic cell transplantation (IUHCT) results in long-term, multilineage chimerism without myeloablation/immunosuppression. It has the potential to treat a number of congenital hematologic disorders, including as Sickle cell disease. Central tolerance, through which donor-reactive host T cells and host-reactive donor T cells are deleted, is instrumental to the induction of donor specific tolerance (DST) and the prevention of graft-versus-host-disease (GVHD). Central deletion, however, is incomplete, and donor-reactive host T cells remain following allogeneic IUHCT. Furthermore, central deletion alone cannot explain the absence of GVHD either following IUHCT using non-T cell depleted bone marrow (BM) or in chimeric animals that undergo postnatal donor lymphocyte infusion (DLI) with subsequent conversion to >90% allogeneic donor cell engraftment (Hayashi et al., Blood 2002;100:804-12). Studies in both mice and humans have demonstrated the importance of regulatory T cells, including traditional CD25+FoxP3+ Tregs and CD49b+LAG-3+ Type 1 regulatory T (Tr1) cells, in maintaining tolerance following postnatal allogeneic transplantation. We hypothesized that peripheral regulatory T cells are increased in chimeric animals following IUHCT and that they contribute to the establishment of DST and prevention of GVHD. Methods: 10x106 BM cells from 6-8 week old C57Bl/6 (B6) FoxP3GFP mice (H2Kb, CD45.2+) were injected intravenously (IV) into gestational day 14 Balb/c FoxP3GFP fetal recipients (H2Kd, CD45.2+). Mice were sacrificed at 2, 4, 8, and 24 weeks of age. Splenocytes were analyzed for the expression of CD4, H2Kb, H2Kd, CD49b, LAG-3, CD25, and FoxP3 to determine the percentage of traditional Tregs and Tr1 cells within the CD4+ population as well as for the expression of intracellular IL-10 - a key mediator of regulatory T cell function. A GVHD model was developed in which 10x106 B6 BM cells were injected IV into day of life 0 (P0) naive Balb/c mice. The ability of IUHCT-induced regulatory T cells to suppress GVHD was assessed in two groups: 1) 10x106 B6 BM cells injected into P0 mice that had undergone IUHCT and 2) 10x106 B6 BM cells co-injected with 5x106 CD4+ splenocytes harvested from 2 week old chimeric mice into P0 naive Balb/c mice. To evaluate the role of regulatory T cells following DLI in chimeric mice, 30x106 B6CD45.1 (H2Kb, CD45.1+) splenocytes were injected IV into 4 week old chimeric recipients. Mice were sacrificed at 48 hours and 2 weeks post-DLI, and regulatory T cells were measured as a percentage of CD45.2+CD4+ cells in the spleen. Results: IUHCT was associated with a significant increase in Tr1 cells, predominantly of donor origin, compared to naive controls (Figure 1). There was a corresponding increase in intracellular IL-10 expression in donor CD4+ splenocytes in chimeric animals compared to naive B6 FoxP3GFP controls at 2 weeks of age (MFI 7.07 vs. 3.26, p=0.0017). Elevated levels of regulatory T cells following IUHCT also had a functional effect in the GVHD model (Figure 2). Mice that had undergone IUHCT (group 1) demonstrated a trend toward improved survival compared to naive mice when injected with B6 BM at P0. Naive mice injected at P0 with B6 BM plus CD4+ splenocytes from chimeric mice (group 2) demonstrated significant improvement in survival compared to naive recipients of B6 BM alone (p=0.049). Although we detected no increase in traditional Tregs after IUHCT alone, we noted a 3-fold increase in traditional Tregs of host origin in chimeric animals at 48 hours post-DLI (25.5% vs. 8.3%, p<0.001). Conclusion: The induction of tolerance following IUHCT is important for maintaining successful long-term engraftment and preventing GVHD. We demonstrate an increase in nontraditional regulatory T cells following IUHCT which can minimize GVHD as well as an increase in traditional Tregs following DLI in an established model of complete allogeneic engraftment. These studies suggest an important role for regulatory T cell populations in the induction of tolerance following IUHCT. They are significant in light of a potential obstacle to the clinical translation of IUHCT: the window of central tolerance induction in humans occurs at a gestational age that will pose significant technical challenges. Augmentation of regulatory T cell-dependent peripheral tolerance is one approach by which IUHCT could be delayed to a technically feasible point later in gestation. Disclosures No relevant conflicts of interest to declare.

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