scholarly journals Peripheral but Not Central Tolerance Regulates RBC Autoreactive CD4+ T Cells in a Novel Mouse

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3441-3441
Author(s):  
Sut Ling Wong ◽  
Amanda L Richards ◽  
James C. Zimring ◽  
Krystalyn E. Hudson
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd4 T Cells ◽  
T Cell Tolerance ◽  
Cell Tolerance ◽  
Tolerance Mechanisms ◽  
Ki 67 ◽  
Central Tolerance

Abstract Background: Elucidating the mechanisms of T cell tolerance towards autoantigens can be critical for therapeutic efforts to re-establish tolerance in autoimmune diseases or break tolerance in tumor immunotherapy. Tolerance mechanisms have been well described towards tissue-restricted antigens and include deletion, anergy, or regulatory cells; however, T cell tolerance mechanisms against RBC-restricted antigens are poorly understood. Herein, we utilized the HOD mouse to investigate T cell tolerance mechanisms to RBC-specific antigens. The HOD mouse expresses a triple fusion protein consisting of hen egg lysosyme (H EL), ovalbumin (O VA), and human blood group molecule, D uffy (HOD) driven by a RBC-specific promoter. A TCR transgenic mouse that recognizes an OVA peptide presented by MHI II (OT-II mouse) was crossed with the HOD mouse to allow analysis of T cells autoreactive to an RBC specific antigen. Methods: To evaluate central tolerance, OVA specific T cells were analyzed from thymii from 6-8 weeks old OT-II+ HOD+ mice (OT-II+ HOD- littermates were used as controls for T cells in absence of autoantigen). Splenic CD4+ T cells from both OT-II+ HOD+ and control OT-II+ HOD- mice were CFSE-labeled and analyzed 1) in vitro by co-culturing with splenocytes pulsed with whole soluble OVA or 2) in vivo by adoptively transferring into B6.Thy1.1 recipients, followed by transfusion of HOD RBC. Proliferative responses of cells were measured by CFSE dilution analyzed with flow cytometry. Results: There is no detectable autoantibody production against the HOD antigen in OT-II+ HOD+ mice. In the thymus, similar frequencies and total nbers of OVA-specific CD4+ single positive (SP) T cells were observed between OT-II+ HOD+ and OT-II+ HOD- mice. This was not because OT-II+ HOD+ mice were incapable of deleting autoreactive T cells, since OT-II+ HOD+ mice i.v injected with soluble whole OVA, known to induce deletion, had a significant decrease in the CD4+ SP T cells. Phenotypic analysis of mature CD4+ T cells in the spleen revealed fewer OVA-specific CD4+ T cells in OT-II+ HOD+ compared to OT-II+ HOD- mice (p=0.009). The CD4+ T cells present in OT-II+ HOD+ expressed higher levels of markers associated with T cell activation and proliferation such as CD69, CD44, Ki-67 and also decreased levels of L-selection. Moreover, these cells expressed elevated levels of markers associated with T cell anergy and tolerance, including PD-1, CD5 and LAG-3. Finally, OT-II+ HOD+ mice exhibited increased numbers of CD25+ Foxp3+ Tregs (p=0.007). In 2 of 2 experiments (3 mice/group), adoptively transferred CD4+ cells from OT-II+ HOD+ did not proliferate in response to HOD RBCs while OT-II+ HOD- exhibited robust proliferation. In contrast to the in vivo results, CD4+ cells from OT-II+ HOD+ proliferated when co-cultured with splenocytes + whole soluble OVA in vitro. Conclusions : We describe a model in which regulation of CD4+ T cells autoreactive for self-RBC antigen can be studied. Although a significant number of RBC autoreactive T cells are present, and appear to have seen antigen, been activated (i.e. increased CD69 and CD44), and proliferated (increased Ki-67); these cells are nevertheless anergic as indicated by adoptive transfer studies. The ability to proliferate in vitro but not in vivo in response to HOD RBC suggests that removal from the normal in vivo environment removes inhibitory factors, consistent with peripheral tolerance mechanisms in vivo. Indeed, increased Tregs were observed in autoreactive mice compared to control mice. In contrast, thymic deletion seems to play little or no role in this case. Together, these findings identify peripheral and not central tolerance as major control mechanisms of RBC autoreactive CD4+ T cells. Mechanisms may include alterations in expression of PD-1, CD5, and LAG-3 by autoreactive cells. Disclosures Zimring: BloodworksNW: Patents & Royalties: Patent Application filed on technology in this abstract - no royalties; Immucor Inc.: Research Funding.

Regulatory T Cells Are Not Required for Prevention of RBC-Specific Autoantibody Generation

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 568-568
Author(s):  
Krystalyn E. Hudson ◽  
James C. Zimring
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Cd4 T Cells ◽  
T Cell Tolerance ◽  
Cell Tolerance ◽  
Tolerance Mechanisms ◽  
Antibody Treatment ◽  
Isotype Control

Introduction: Loss of humoral tolerance to red blood cell (RBC) antigens may lead to the generation of pathogenic autoantibodies and result in autoimmune hemolytic anemia (AIHA), a severe and potentially fatal disease. Failure of tolerance to RBC antigens occurs with considerable frequency (1-3 cases/1,000 adults) and prevalence of AIHA is as high as 30% in persons with compromised B and/or T cell tolerance mechanisms. However, RBC-specific tolerance mechanisms are poorly understood. To elucidate the immune tolerances to RBC autoantigens, we utilized HOD mice. The HOD mouse expresses an RBC-specific transgene consisting of hen egg lysozyme (HEL), ovalbumin (OVA), and Duffy. Using the HOD model, we previously demonstrated B cell tolerance to RBC-specific HOD antigen is incomplete; however, T cell tolerance is stringent. HOD mice have similar detectable frequencies of HOD-specific CD4+ T cells compared to B6 mice. Although present, autoreactive HOD-specific CD4+ T cells are non-functional. Circumventing T cell tolerance by adoptive transfer, HOD mice make high titer anti-HOD autoantibodies in vivo. Thus, despite the presence of autoreactive B cells, no HOD-reactive antibodies are detectable unless CD4+ T cells are given, indicating T cell tolerance is a stopgap to autoimmunity. Methods: Leukocytes from C57BL/6 (B6) and HOD mice were harvested and OVA-specific CD4+ T cell responses were assessed by tetramer-pulldown assays with pooled tetramers I-Ab-OVA 329-337/326-334. Isolated cells were stained for surface and intracellular markers and analyzed via flow cytometry. For in vivo analysis, mice were treated with 300ug anti-CD25 (clone PC-61) depleting antibody or isotype control; a subset of antibody-treated mice was immunized with OVA/CFA. Antibodies bound to HOD RBCs were determined by direct antibody test. Anti-HOD antibodies were quantified by indirect immunofluorescence using HOD RBCs as targets. Results: Tetramer pull-down assays revealed similar numbers of OVA-reactive CD4+ T cells from HOD and B6 mice (mean 56 and 40, respectively, p = 0.3). However, cell surface and intracellular marker staining demonstrated that HOD mice had higher numbers of OVA-tetramer reactive CD4+ T cells that express regulatory markers CD25 and FoxP3, and exhaustion marker PD1 as compared to control B6 mice. Inhibitory CTLA4 expression was not detectable on OVA-reactive CD4+ T cells from HOD or B6 mice. To test whether regulatory T cells were required for RBC-specific immune tolerance, HOD and B6 mice were treated with CD25 depleting antibody or isotype control antibody. Anti-CD25 antibody treated mice had a significant reduction of CD25+ cells 4 days post treatment (p < 0.001, 2 independent experiments). Similarly, there was a significant reduction in FoxP3+CD25+CD4+ T cells (Tregs) in anti-CD25 treated mice (p < 0.001), compared to isotype. Mice received weekly injections of anti-CD25 or isotype antibody to maintain depletion for one month. A subset of mice received an OVA/CFA immunization. Sustained CD25+ depletion did not result in anti-HOD autoantibody generation. Further, there was no change in the endogenous frequency of OVA-reactive CD4+ T cells between HOD and B6 mice, regardless of antibody treatment. Similarly, HOD mice treated with depletion (or isotype) antibody and immunized with OVA/CFA did not make detectable anti-HOD autoantibodies. Consistent with lack of detectable autoantibodies, no expansion of OVA-tetramer reactive CD4+ T cells was observed in HOD mice. In contrast, B6 mice (treated with anti-CD25 or isotype antibody) had a detectable expansion of OVA-specific CD4+ T cells as a result of immunization. Conclusions: The data demonstrate a phenotypic difference between the OVA-reactive CD4+ T cells from HOD and B6 mice, with an increase in number of Tregs detectable in HOD mice. Administration of anti-CD25 antibody significantly reduced the number of overall CD25+ cells and Tregs. Prolonged depletion of these cellular subsets did not elicit autoantibodies in HOD mice. Further, immunization of CD25 depleted mice with a strong immune stimulus (OVA/CFA, known to expand OVA-reactive T cells in B6 mice), did not induce anti-HOD autoantibodies nor did it expand OVA-specific autoreactive CD4+ T cells in HOD mice. Together, these data demonstrate that CD25+ cells are not required for the maintenance of RBC-specific T cell tolerance and suggest a role for other regulatory mechanisms. Disclosures No relevant conflicts of interest to declare.

scholarly journals Loss of Zbtb32 in NOD mice does not significantly alter T cell responses.

F1000Research ◽  
2018 ◽  
Vol 7 ◽  
pp. 318
Author(s):  
William D. Coley ◽  
Yongge Zhao ◽  
Charles J. Benck ◽  
Yi Liu ◽  
Chie Hotta-Iwamura ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Nod Mice ◽  
Diabetes Incidence ◽  
T Cell Tolerance ◽  
Cell Tolerance

Background: We previously identified the transcriptional regulator Zbtb32 as a factor that can promote T cell tolerance in the Non-Obese Diabetic (NOD) mouse, a model of Type 1 diabetes. Antigen targeted to DCIR2+ dendritic cells (DCs) in vivo inhibited both diabetes and effector T cell expansion in NOD mice. Furthermore, Zbtb32 was preferentially induced in autoreactive CD4 T cells stimulated by these tolerogenic DCIR2+ DCs, and overexpression of Zbtb32 in islet-specific T cells inhibited the diabetes development by limiting T cell proliferation and cytokine production. Methods: To further understand the role of Zbtb32 in T cell tolerance induction, we have now used CRISPR to target the Zbtb32 gene for deletion directly in NOD mice and characterized the mutant mice. We hypothesized that the systemic loss of Zbtb32 in NOD mice would lead to increased T cell activation and increased diabetes pathogenesis. Results: Although NOD.Zbtb32-/- male NOD mice showed a trend towards increased diabetes incidence compared to littermate controls, the difference was not significant. Furthermore, no significant alteration in lymphocyte number or function was observed. Importantly, in vitro stimulation of lymphocytes from NOD.Zbtb32-/- mice did not produce the expected hypersensitive phenotype observed in other genetic strains, potentially due to compensation by homologous genes. Conclusions: The loss of Zbtb32 in the NOD background does not result in the expected T cell activation phenotype.

scholarly journals Loss of Zbtb32 in NOD mice does not significantly alter T cell responses.

F1000Research ◽  
2018 ◽  
Vol 7 ◽  
pp. 318
Author(s):  
William D. Coley ◽  
Yongge Zhao ◽  
Charles J. Benck ◽  
Yi Liu ◽  
Chie Hotta-Iwamura ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Nod Mice ◽  
Diabetes Incidence ◽  
T Cell Tolerance ◽  
Cell Tolerance

Background:We previously identified the transcriptional regulator Zbtb32 as a factor that can promote T cell tolerance in the Non-Obese Diabetic (NOD) mouse, a model of Type 1 diabetes. Antigen targeted to DCIR2+dendritic cells (DCs)in vivoinhibited both diabetes and effector T cell expansion in NOD mice. Furthermore, Zbtb32 was preferentially induced in autoreactive CD4 T cells stimulated by these tolerogenic DCIR2+DCs, and overexpression of Zbtb32 in islet-specific T cells inhibited the diabetes development by limiting T cell proliferation and cytokine production.Methods:To further understand the role of Zbtb32 in T cell tolerance induction, we have now used CRISPR to target the Zbtb32 gene for deletion directly in NOD mice and characterized the mutant mice. We hypothesized that the systemic loss of Zbtb32 in NOD mice would lead to increased T cell activation and increased diabetes pathogenesis.Results:Although NOD.Zbtb32-/-male NOD mice showed a trend towards increased diabetes incidence compared to littermate controls, the difference was not significant. Furthermore, no significant alteration in lymphocyte number or function was observed. Importantly,in vitrostimulation of lymphocytes from NOD.Zbtb32-/-mice did not produce the expected hypersensitive phenotype observed in other genetic strains, potentially due to compensation by homologous genes.Conclusions:The loss of Zbtb32 in the NOD background does not result in the expected T cell activation phenotype.

scholarly journals Cd40-Independent Pathways of T Cell Help for Priming of Cd8+ Cytotoxic T Lymphocytes

2000 ◽  
Vol 191 (3) ◽  
pp. 541-550 ◽  
Author(s):  
Zhengbin Lu ◽  
Lingxian Yuan ◽  
Xianzheng Zhou ◽  
Eduardo Sotomayor ◽  
Hyam I. Levitsky ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd4 T Cells ◽  
Cytotoxic T Lymphocyte ◽  
Cell Communication ◽  
Cd8 T Cell ◽  
Cd4 Help ◽  
T Cell Help

In many cases, induction of CD8+ CTL responses requires CD4+ T cell help. Recently, it has been shown that a dominant pathway of CD4+ help is via antigen-presenting cell (APC) activation through engagement of CD40 by CD40 ligand on CD4+ T cells. To further study this three cell interaction, we established an in vitro system using dendritic cells (DCs) as APCs and influenza hemagglutinin (HA) class I and II peptide–specific T cell antigen receptor transgenic T cells as cytotoxic T lymphocyte precursors and CD4+ T helper cells, respectively. We found that CD4+ T cells can provide potent help for DCs to activate CD8+ T cells when antigen is provided in the form of either cell lysate, recombinant protein, or synthetic peptides. Surprisingly, this help is completely independent of CD40. Moreover, CD40-independent CD4+ help can be documented in vivo. Finally, we show that CD40-independent T cell help is delivered through both sensitization of DCs and direct CD4+–CD8+ T cell communication via lymphokines. Therefore, we conclude that CD4+ help comprises at least three components: CD40-dependent DC sensitization, CD40-independent DC sensitization, and direct lymphokine-dependent CD4+–CD8+ T cell communication.

scholarly journals Selective anergy of V beta 8+,CD4+ T cells in Staphylococcus enterotoxin B-primed mice.

1990 ◽  
Vol 172 (4) ◽  
pp. 1065-1070 ◽  
Author(s):  
Y Kawabe ◽  
A Ochi
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
T Cell Anergy ◽  
Specific Cytotoxicity ◽  
Enterotoxin B ◽  
Staphylococcus Enterotoxin B

The cellular basis of the in vitro and in vivo T cell responses to Staphylococcus enterotoxin B (SEB) has been investigated. The proliferation and cytotoxicity of V beta 8.1,2+,CD4+ and CD8+ T cells were observed in in vitro response to SEB. In primary cytotoxicity assays, CD4+ T cells from control spleens were more active than their CD8+ counterparts, however, in cells derived from SEB-primed mice, CD8+ T cells were dominant in SEB-specific cytotoxicity. In vivo priming with SEB abrogated the response of V beta 8.1,2+,CD4+ T cells despite the fact that these cells exist in significant number. This SEB-specific anergy occurred only in V beta 8.1,2+,CD4+ T cells but not in CD8+ T cells. These findings indicate that the requirement for the induction of antigen-specific anergy is different between CD4+ and CD8+ T cells in post-thymic tolerance, and the existence of coanergic signals for the induction of T cell anergy is suggested.

scholarly journals In Vivo Detection of Dendritic Cell Antigen Presentation to CD4+ T Cells

1997 ◽  
Vol 185 (12) ◽  
pp. 2133-2141 ◽  
Author(s):  
Elizabeth Ingulli ◽  
Anna Mondino ◽  
Alexander Khoruts ◽  
Marc K. Jenkins
Keyword(s):  
T Cells ◽  
T Cell ◽  
Lymph Nodes ◽  
T Cell Activation ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Delayed Type Hypersensitivity ◽  
Physical Interaction

Although lymphoid dendritic cells (DC) are thought to play an essential role in T cell activation, the initial physical interaction between antigen-bearing DC and antigen-specific T cells has never been directly observed in vivo under conditions where the specificity of the responding T cells for the relevant antigen could be unambiguously assessed. We used confocal microscopy to track the in vivo location of fluorescent dye-labeled DC and naive TCR transgenic CD4+ T cells specific for an OVA peptide–I-Ad complex after adoptive transfer into syngeneic recipients. DC that were not exposed to the OVA peptide, homed to the paracortical regions of the lymph nodes but did not interact with the OVA peptide-specific T cells. In contrast, the OVA peptide-specific T cells formed large clusters around paracortical DC that were pulsed in vitro with the OVA peptide before injection. Interactions were also observed between paracortical DC of the recipient and OVA peptide-specific T cells after administration of intact OVA. Injection of OVA peptide-pulsed DC caused the specific T cells to produce IL-2 in vivo, proliferate, and differentiate into effector cells capable of causing a delayed-type hypersensitivity reaction. Surprisingly, by 48 h after injection, OVA peptide-pulsed, but not unpulsed DC disappeared from the lymph nodes of mice that contained the transferred TCR transgenic population. These results demonstrate that antigen-bearing DC directly interact with naive antigen-specific T cells within the T cell–rich regions of lymph nodes. This interaction results in T cell activation and disappearance of the DC.

scholarly journals T cell memory. Long-term persistence of virus-specific cytotoxic T cells.

1989 ◽  
Vol 169 (6) ◽  
pp. 1993-2005 ◽  
Author(s):  
B D Jamieson ◽  
R Ahmed
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Population ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Biological Properties ◽  
Antigenic Specificity ◽  
Ctl Response

This study documents that virus-specific CTL can persist indefinitely in vivo. This was accomplished by transferring Thy-1.1 T cells into Thy-1.2 recipient mice to specifically identify the donor T cell population and to characterize its antigenic specificity and function by using a virus-specific CTL assay. Thy-1.1+ T cells from mice previously immunized with lymphocytic choriomeningitis virus (LCMV) were transferred into Thy-1.2 mice persistently infected with LCMV. The transferred LCMV-specific CTL (Thy-1.1+ CD8+) eliminate virus from the chronically infected carriers and persist in the recipient mice in small numbers, comprising only a minor fraction of the total T cells. Upon re-exposure to virus, these long-lived "resting" CD8+ T cells proliferate in vivo to become the predominant cell population. These donor CD8+ T cells can be recovered up to a year post-transfer and still retain antigenic specificity and biological function. They kill LCMV infected H-2-matched cells in vitro and can eliminate virus upon transfer into a second infected host. In addition, these long-lived CD8+ T cells appear not to be dependent on help from CD4+ T cells, since depletion of CD4+ T cells has minimal or no effect on their biological properties (proliferation, CTL response, viral clearance). These donor CTL also exhibit an immunodominance over the host-derived LCMV-specific CTL response. When both host and donor T cells are present, the donor CTL response is dominant over the potential CTL response of the cured carrier host. Taken together, these results suggest that virus-specific CTL can persist for the life span of the host as memory cells.

scholarly journals P025 Identification and validation of predictors for tofacitinib through supervised and unbiased approaches in Inflammatory Bowel Disease

2021 ◽  
Vol 15 (Supplement_1) ◽  
pp. S141-S141
Author(s):  
B Liu ◽  
M Spalinger ◽  
L G Perez ◽  
A Machicote ◽  
N Gagliani ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
Signaling Pathway ◽  
Cd4 T Cells ◽  
Cytokine Production ◽  
Stat Signaling ◽  
Deficient Mice

Abstract Background Inflammatory Bowel Disease (IBD) is characterized by an overwhelming gut inflammation, where CD4+ effector T cells are main mediators of the inflammatory response. Tofacitinib, a small molecular drug recently used in IBD patients, blocks the JAK/STAT signaling pathway necessary for CD4+ effector T-cell activation. However, clinical data show that a percentage of patients do not respond to the treatment. Our main goal is to identify biomarkers predicting the response of patients to tofacitinib. Methods Tofacitinib efficacy was studied in vivo in wild type (WT) and T-cell-specific PTPN2 deficient mice (CD4-Cre;Ptpn2 floxed) in which the JAK/STAT signaling pathway is over activated. WT and PTPN2 deficient mice were gavaged with tofacitinib (50mg/kg, twice daily) or vehicle. Acute DSS-colitis was induced. Colitis development was evaluated by weight loss, colonoscopy and histology. CD4+ T cells were isolated from the colon and analyzed by flow cytometry. To study the effect of tofacitinib on T-cell differentiation, we isolated naïve T cells from mouse spleen and polarized them in vitro to different T-cell subsets with or without tofacitinib. CD4+ T cells differentiation and cytokine production were analyzed by flow cytometry. To evaluate the influence of tofacitinib on human CD4+ T cells, human peripheral blood mononuclear cells (PBMCs) from healthy donors and IBD patients were stimulated in presence of tofacitinib, and analyzed by flow cytometry. Results While no protective effect was found after tofacitinib treatment in WT mice, PTPN2 deficient mice were protected from colitis based on less weight loss, lower endoscopic and histological scores. The expression of pro-inflammatory cytokines such as IL-17 and IFN-γ by colonic CD4+ T cells was also decreased by tofacitinib. Consistent with the in vivo observations, in vitro experiments revealed a strong impact of tofacitinib on CD4+ T-cells cytokine production. In PBMCs from IBD patients, IFN-γ and TNF-α expression was strongly impacted. In contrast, in healthy donors, IL-10 was the most impacted cytokine. Finally, tofacitinib decreased the in vitro differentiation of Th1, Th2, Th17, Th22, Treg and Tr1. Conclusion In the T-cell-specific PTPN2 deficient mice, tofacitinib exerts a protective effect after DSS-induced colitis. In line with the in vivo findings, in vitro experiments show that tofacitinib has a strong impact on pro-inflammatory cytokine production, especially in the IBD patients. Taken together, these data suggest that tofacitinib might be suitable primarily for IBD patients where the JAK/STAT signaling pathway is over activated.

scholarly journals Regulation of layered T cell tolerance mechanisms by the NR4A family is essential to preserve immune homeostasis and suppress autoimmunity

2021 ◽  
Author(s):  
Ryosuke Hiwa ◽  
Hailyn V. Nielsen ◽  
James L. Mueller ◽  
Julie Zikherman
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Inflammatory Disease ◽  
Immune Homeostasis ◽  
T Cell Tolerance ◽  
Cell Tolerance ◽  
Tolerance Mechanisms ◽  
Systemic Inflammatory Disease ◽  
A Cell

ABSTRACTThe NR4A family of orphan nuclear receptors (Nr4a1-3) plays redundant roles upstream of Foxp3 to establish and maintain Treg identity; deletion of multiple family members in the thymus results in Treg deficiency and a severe inflammatory disease. Consequently, it has been challenging to isolate the functions of this family in other immune cells. Here we take advantage of a competitive bone marrow chimera strategy, coupled with conditional genetic tools, to rescue Treg homeostasis and unmask such functions. Unexpectedly, chimeras harboring Nr4a1−/− Nr4a3−/− (DKO) bone marrow develop autoantibodies and a systemic inflammatory disease despite a replete Treg compartment of largely wild-type origin. This disease differs qualitatively from that seen with Treg-deficiency and is B cell-extrinsic. Negative selection of DKO thymocytes is profoundly impaired in a cell-intrinsic manner. Consistent with escape of self-reactive T cells into the periphery, DKO T cells with functional and phenotypic features of anergy accumulate in chimeric mice. Despite this, DKO T cells exhibit enhanced IL-2 production, implying a cell-intrinsic role for the NR4A family in peripheral T cell tolerance. These studies reveal roles for the NR4A family in multiple layered T cell tolerance mechanisms and demonstrate that each is essential to preserve immune homeostasis.

Histone Deacetylase Inhibitors Induce Immuno-Dominant Suppression of Dendritic Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 456-456 ◽  
Author(s):  
Pavan Reddy ◽  
Yoshinobu Maeda ◽  
Raimon Duran-Struuck ◽  
Oleg Krijanovski ◽  
Charles Dinarello ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Mhc Class Ii ◽  
Cd4 T Cells ◽  
Hdac Inhibitors ◽  
Class Ii ◽  
Wild Type ◽  
Tnf Α

Abstract We and others have recently demonstrated that suberoylanilide hydroxamic acid (SAHA), a histone deacetylase (HDAC) inhibitor with anti-neoplastic properties, reduces experimental acute graft-versus-host disease (GVHD). We have now investigated the mechanisms of action of two HDAC inhibitors, SAHA and ITF 2357, on allogeneic immune responses. Bone marrow derived dendritic cells (DCs) were preincubated with the HDAC inhibitors at nanomolar concentrations for 16–18 hours and stimulated with lipopolysaccharide (LPS). Pretreatment of DCs caused a significant reduction in the secretion of TNF-α, IL-12p70 and IL-6 compared to the untreated controls (P< 0.005). Similar effects were seen using human peripheral blood mononuclear cell derived DCs. Pre-treatment of both murine and human DCs also significantly reduced their in vitro stimulation of allogeneic T cells as measured by proliferation and IFN-γ production (P<0.01). We determined the in vivo relevance of these observations utilizing a mouse model where the responses of allogeneic donor bm12 T cells depended on the function of injected host B6 DCs would stimulate. Recipient Class-II −/− B6 (H-2b) received 11 Gy on day -1 and were injected with 4–5 x 106 wild type B6 DCs treated with SAHA or with media on days -1 and 0 and then transplanted with 2 x 106 T cells and 5 x 106 TCDBM cells from either syngeneic B6 or allogeneic bm12 donors. SAHA treatment of DCs significantly reduced expansion of allogeneic donor CD4+ T cells on day +7 after BMT compared to controls (P<0.05). SAHA treatment induced a similarly significant reduction in the expansion of CD8+ cells in Class I disparate [bm1→β2M−/−] model. In vitro, SAHA treatment significantly suppressed the expression of CD40 and CD80 but did not alter MHC class II expression. Surprisingly, when mixed with normal DCs at 1:1 ratio, SAHA treated DCs dominantly suppressed allogeneic T cell responses. The regulation of T cell proliferation was not reversible by addition of IL-12, TNF-α, IL-18, anti-IL-10 or anti-TGFβ, either alone or in combination. Suppression of allogeneic responses was contact dependent in trans-well experiments. To address whether the regulation of SAHA treated DCs required contact with T cells, we devised a three cell experiment where SAHA treated DCs lacked the capacity to present antigens to T cells. DCs from B6 MHC Class II deficient (H-2b) were treated with SAHA and co-cultured with wild type B6 (H-2b) DCs along with purified allogeneic BALB/c (H-2d) CD4+ T cells in an MLR. Allogeneic CD4+ T cells proliferated well, demonstrating the regulation to be dependent on contact between SAHA treated DCs and T cells. To address the in vivo relevance of this suppression, we utilized a well characterized [BALB/c →B6] mouse model of acute GVHD. Recipient B6 animals received 11Gy on day -1 and were injected with of 5 million host type SAHA treated or control DCs on days −1, 0, and +2. Mice were transplanted on day 0 with 2 x 106 T cells and 5 x 106 BM from either syngeneic B6 or allogeneic BALB/c donors. Injection of SAHA treated DCs resulted in significantly better survival (60% vs. 10%, P < 0.01) and significantly reduced serum levels of TNF-α, donor T cell expansion and histopathology of GVHD on day +7 after BMT compared to the controls. We conclue that HDAC inhibitors are novel immunomodulators that regulate DC function and might represent a novel strategy to prevent GVHD.

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