PRDM1 Is Sufficient for Inducing Human Primary T Cell Hyporesponsiveness and Implicates Low Gvhd Occurrence after Allo-HSCT

Abstract T cell hyporesponsiveness is crucial for functional immune system and prevent the damage induced by alloreactive T cells in autoimmune pathology and transplantation. As one of the most important regulators during B cell development, PRDM1 (also known as BLIMP-1) has been demonstrated its essential role for maintaining T cell hyporesponsiveness and homeostasis, evidenced by Prdm1 deficient mice accumulating activated T cells and developing multiorgan inflammatory disease. However, the mechanism of PRDM1 regulating T cell hyporesponsiveness is still ambiguous. In this study, we took advantage of multiomics technologies and systemic report the central role of PRDM1 in inducing human primary T cell hyporesponsiveness. Firstly, we overexpressed PRDM1 in human primary T cells and found increased ratio of CD4 +CD25 +FOXP3 + Treg cell subsets and increased IL-4 secretion. In parallel, inhibited PRDM1 expression level in human T cells decreased ratio of Treg cells and secretion of IL-4. Meanwhile, transcriptome analyses revealed that overexpressed PRDM1 enriched negative regulation of cell proliferation signaling pathway and resulted in a global reduction in IL-2 and inflammatory response signaling pathways. Furthermore, overexpressed PRDM1 in primary T cells upregulated several negative regulators of T cell function like EOMES, KLF2, LILRB1, KLRB1 and CD244, indicating a pioneer role of PRDM1 in inducing T cell hyporesponsive. To further investigate the regulation role of T cell hyporesponsiveness of PRDM1, we performed CUT&Tag and ATAC-seq in PRDM1 overexpressed primary T cells. CUT&Tag analysis showed PRDM1 could directly upregulated T cell hyporesponsiveness related gene such as KLF2, CD244 and KLRD1. Importantly, we observed consistent changes of IL-2, central regulator of T cell activation, in PRDM1 overexpressed T cell from ATAC-seq, CUT&Tag and RNA-seq data. We found PRDM1 could binding to IL-2 locus and decreased the chromatin accessibility of IL-2, consequently downregulated the expression level of IL-2 in human primary T cells. Moreover, altered open chromatin regions (OCRs) in PRDM1 overexpressed T cells enriched the similar transcription factors (TFs) with PRDM1 binding sites, indicating PRDM1 might be a pioneer TF in T cell hyporesponsiveness. These results demonstrated PRDM1 is sufficient for inducing T cell hyporesponsiveness in human primary T cells. To further validate the coexpression relationship between PRDM1 and Treg cell central TF FOXP3, we upregulated PRDM1 expression level on Jurkat T cells lines. The results also showed that elevated FOXP3 both in mRNA and protein level accompanied with upregulated PRDM1 expression level. To analyze the mechanism of PRDM1 regulating FOXP3 expression level, CUT&Tag data analyses showed that PRDM1 might upregulated FOXP3 by directly binding to the enhancer region of upstream of FOXP3 locus. Meanwhile, PRDM1 indirectly upregulated FOXP3 by upregulated KLF2, evidenced by inhibiting KLF2 in PRDM1 overexpressed primary T cells downregulated FOXP3 expression level. To further investigate the clinical implication of PRDM1 inducing T cell hyporesponsiveness, we detected the relationship of PRDM1 expression level and acute graft-versus-host disease(aGVHD) occurrence after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Our results showed that patients with aGVHD (n=7) exhibited lower PRDM1 expression level than those without aGVHD in the same period after HSCT (n=7). Furthermore, we detected the expression level of PRDM1 in CD4 + T cell and CD8 + T cells from bone marrow allografts (BM) or peripheral blood allografts (PB) and followed up the occurrence of GVHD after HSCT for 2 years (n=18). There are low expression levels of PRDM1 in CD4 + T cells both from BM or PB grafts corelated with aGVHD occurrence in patients after allo-HSCT compared with those without aGVHD occurrence. In conclusion, our study provides the global regulatory model of PRDM1 in human primary T cell. We introduced PRDM1 as a sufficient regulator in T cell hyporesponsiveness induction, which is altering the chromatin accessibility and directly upregulated T cell inhibitory signals and downregulated T cell activated signals. The negative relationship between PRDM1 expression level with GVHD occurrence indicated it might be a potential biomarker for indicating HSCT prognosis. Disclosures No relevant conflicts of interest to declare.

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A Hypomorphic Cbfb Allele Reveals a Critical Dosage-Sensitive Function of Core Binding Factors at the Earliest Stages of T Cell Development.

Blood ◽

10.1182/blood.v106.11.124.124 ◽

2005 ◽

Vol 106 (11) ◽

pp. 124-124

Author(s):

Ivan Maillard ◽

Laleh Talebian ◽

Zhe Li ◽

Yalin Guo ◽

Daisuke Sugiyama ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Dna Binding ◽

B Cell ◽

Bone Marrow Cells ◽

T Cell Development ◽

Cell Development ◽

B Cell Development ◽

Hematopoietic Stem

Abstract The family of core binding factors includes the DNA-binding subunits Runx1-3 and the common non-DNA binding partner CBFβ. Runx1 and CBFβ are essential for the emergence of hematopoietic stem cells during fetal development, but not for stem cell maintenance during later ontogeny. Runx1 is also required for megakaryocyte differentiation, B cell development, and for the DN2 to DN3 transition in thymocyte development. Runx2/CBFβ are critical for normal osteogenesis, and Runx3 for CD4 silencing in CD8+ T cells, but their contribution to other steps of hematopoietic development is unknown. To examine the collective role of core binding factors in hematopoiesis, we generated a hypomorphic Cbfb allele (Cbfbrss). CBFβ protein levels were reduced by approximately 2–3 fold in fetuses homozygous for the Cbfbrss allele (Cbfbrss/rss), and 3–4 fold in fetuses carrying one hypomorphic and one knockout allele (Cbfbrss/−). Cbfbrss/rss and Cbfbrss/− fetuses had normal erythroid and B cell development, and relatively mild abnormalities in megakaryocyte and granulocyte differentiation. In contrast, T cell development was very sensitive to an incremental reduction of CBFβ levels: mature thymocytes were decreased in Cbfbrss/rss fetuses, and virtually absent in Cbfbrss/−fetuses. We next assessed the development of Cbfbrss/rss and Cbfbrss/− fetal liver progenitors after transplantation to irradiated adult recipients, in competition with wild-type (wt) bone marrow cells. Wt, Cbfbrss/rss and Cbfbrss/− fetal progenitors replenished the erythroid, myeloid and B cell compartments equally well. The overall development of Cbfbrss/rss T cells was preserved, although CD4 expression was derepressed in double negative thymocytes. In Cbfbrss/− chimeras, mature thymocytes were entirely derived from competitor cells. Furthermore, the developmental block in Cbfbrss/− progenitors was present at the earliest stages of T cell development within the DN1 (ETP) and DN2 subsets. Our data define a critical CBFβ threshold for normal T cell development, and they situate an essential role of core binding factors during the earliest stages of T cell development. In addition, early thymopoiesis appeared more severely affected by reduced CBFβ dosage than by the lack of Runx1 (Ichikawa et al., Nat Med 2004; Growney et al., Blood 2005), suggesting that Runx2/3 may contribute to core binding factor activity in the T cell lineage.

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Generation of CMV Specific T Cells From CMV Seronegative Donors by T Cell Receptor RNA Transfer.

Blood ◽

10.1182/blood.v114.22.3577.3577 ◽

2009 ◽

Vol 114 (22) ◽

pp. 3577-3577

Author(s):

Simone Thomas ◽

Sebastian Klobuch ◽

Mirjam H.M. Heemskerk ◽

Diana Stolle ◽

Katrin Besold ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Cd8 T Cells ◽

Human Fibroblasts ◽

Expression Level ◽

Specific Lysis ◽

Hematopoietic Stem ◽

Peptide Epitope ◽

Retroviral Transduction ◽

Ifn Γ

Abstract Abstract 3577 Poster Board III-514 Reactivation of latent human cytomegalovirus (CMV) infection is a frequent complication in CMV seropositive patients after allogeneic hematopoietic stem cell transplantation (HSCT). Although antiviral drug therapy is successfully used to reduce the risk of CMV disease, long-term virus control requires the re-establishment of protective antiviral T cell immunity in the host. The latter is challenging, particularly if the donor is CMV seronegative and thus, no CMV specific T cells are being transferred from donor to recipient during HSCT. Grafting nonreactive T cells of CMV seronegative donors by virus-antigen specific T cell receptors (TCR) may be an efficient means to transfer CMV specific T cell function into allogeneic HSCT recipients. In this study, we intended to reprogram T cells of CMV seronegative donors with human TCR recognizing the immunodominant HLA-A*0201 binding peptide epitope NLVPMVATV (495-503) derived from the CMV pp65 protein. A common approach for TCR gene transfer into T cells is retroviral transduction bearing the risk of insertional mutagenesis which hampers clinical translation. In addition, heterologous recombination between introduced and naturally expressed TCR chains might lead to the induction of harmful autoimmunity. Herein we used in vitro transcribed RNA encoding the CMV pp65/HLA-A*0201-specific TCR for electroporation of anti-CD3 stimulated T cells in peripheral blood mononuclear cells (PBMC) of CMV seronegative donors. This procedure resulted in transient surface expression of the introduced TCR chains up to 5 days as shown by flow cytometry. Maximum expression level was observed at 4 to 24 h after electroporation, with up to 70% of total CD8+ and CD4+ T cells staining positive for the vβ13.1 subfamily domain of the TCRβ chain. After introduction of TCR RNA, the intensity of CMV pp65/HLA-A*0201 tetramer staining was 60% and 50% of total CD8+ and CD4+ cells, respectively. In IFN-γ ELISPOT and 51Chromium-release assays, TCR RNA transfected T cells recognized HLA-A*0201 expressing T2 cells pulsed with titrated amounts of CMV pp65 (495-503) peptide. Minimal peptide concentration triggering specific lysis was 0.1 nM to 1 nM at a CD8+ to target (CD8+:T) ratio of 2:1. The EC50 value (0.2 nM) was in the same range of avidity compared to that of a retrovirally transduced counterpart construct of this TCR. Most importantly, TCR recipient CD8+ T cells gained the ability to lyse HLA-A*0201 positive human fibroblasts upon infection with CMV. Specific lysis between 20% and 100% was observed at a CD8+:T ratio of 1:1 or higher. We next sorted CD8+ T cells from PBMC of CMV seronegative donors into naïve and memory cells according to expression of the differentiation markers CD45RA and CD45RO. Although 90% of naïve CD8+ T cells stained positive for the CMV pp65/HLA-A*0201 tetramer after electroporation of TCR RNA, they mediated only marginal lysis toward CMV infected fibroblasts. In contrast, TCR RNA transfected memory CD8+ T cells showed strong lysis against CMV infected fibroblasts at a CD8+:T ratio of 0.7:1 or higher. Specific lysis was detected for at least 3 days after electroporation. In summary, our data demonstrate that nonreactive human T cells can be successfully redirected with CMV pp65 TCR RNA. The expression level of the introduced TCR is sufficient to trigger IFN-γ production and cytolytic activity toward CMV infected human fibroblasts. Electroporation of TCR RNA is comparably easy and eliminates the risk of retroviral transduction. We therefore believe that CMV pp65 TCR RNA has the potential to be further developed as a therapeutic “off-the-shelf” reagent for patients who undergo drug-resistant CMV reactivation after HSCT. Disclosures: No relevant conflicts of interest to declare.

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TCR-dependent differentiation of thymic Foxp3+ cells is limited to small clonal sizes

Journal of Experimental Medicine ◽

10.1084/jem.20091033 ◽

2009 ◽

Vol 206 (10) ◽

pp. 2121-2130 ◽

Cited By ~ 102

Author(s):

Monica W.L. Leung ◽

Shiqian Shen ◽

Juan J. Lafaille

Keyword(s):

T Cells ◽

T Cell ◽

T Cell Receptors ◽

Foxp3 Expression ◽

Bone Marrow Chimera ◽

High Affinity ◽

Intrathymic Injection ◽

Deficient Mice ◽

Selection Of

Numerous studies have highlighted the importance of high-affinity interactions between T cell receptors (TCRs) and their ligands in the selection of Foxp3+ regulatory T cells (T reg cells). To determine the role of the TCR in directing T cells into the Foxp3+ lineage, we generated transgenic (Tg) mice expressing TCRs from Foxp3+ cells. Initial analyses of the TCR Tg mice crossed with RAG-deficient mice showed that the percentage of Foxp3+ cells was very low. However, intrathymic injection and bone marrow chimera experiments showed a saturable increase of the Foxp3+ population when T reg TCR Tg cells were present in low numbers. Furthermore, when analyzing whole thymi of T reg TCR Tg RAG-deficient mice, we found significantly more Foxp3+ cells than in conventional T cell TCR Tg mice. Our results indicate that although the TCR has an instructive role in determining Foxp3 expression, selection of Foxp3+ individual clones in the thymus is limited by a very small niche.

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Role of regulatory T-cells in autoimmunity

Clinical Science ◽

10.1042/cs20080200 ◽

2009 ◽

Vol 116 (8) ◽

pp. 639-649 ◽

Cited By ~ 20

Author(s):

Richard J. Mellanby ◽

David C. Thomas ◽

Jonathan Lamb

Keyword(s):

T Cells ◽

T Cell ◽

Regulatory T Cells ◽

Treg Cell ◽

Cell Function ◽

Cell Subset ◽

Treg Cells ◽

Self Tolerance

There has been considerable historical interest in the concept of a specialist T-cell subset which suppresses over-zealous or inappropriate T-cell responses. However, it was not until the discovery that CD4+CD25+ T-cells had suppressive capabilities both in vitro and in vivo that this concept regained credibility and developed into one of the most active research areas in immunology today. The notion that in healthy individuals there is a subset of Treg-cells (regulatory T-cells) involved in ‘policing’ the immune system has led to the intensive exploration of the role of this subset in disease resulting in a number of studies concluding that a quantitative or qualitative decline in Treg-cells is an important part of the breakdown in self-tolerance leading to the development of autoimmune diseases. Although Treg-cells have subsequently been widely postulated to represent a potential immunotherapy option for patients with autoimmune disease, several studies of autoimmune disorders have demonstrated high numbers of Treg-cells in inflamed tissue. The present review highlights the need to consider a range of other factors which may be impairing Treg-cell function when considering the mechanisms involved in the breakdown of self-tolerance rather than focussing on intrinsic Treg-cell factors.

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Memory/effector (CD45RBlo) CD4 T cells are controlled directly by IL-10 and cause IL-22–dependent intestinal pathology

Journal of Experimental Medicine ◽

10.1084/jem.20102149 ◽

2011 ◽

Vol 208 (5) ◽

pp. 1027-1040 ◽

Cited By ~ 110

Author(s):

Masahito Kamanaka ◽

Samuel Huber ◽

Lauren A. Zenewicz ◽

Nicola Gagliani ◽

Chozhavendan Rathinam ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Treg Cell ◽

Cd4 T Cells ◽

Messenger Rna ◽

Dominant Negative ◽

T Cell Subsets ◽

Intestinal Pathology ◽

Inflammatory Bowel

The role of direct IL-10 signaling in different T cell subsets is not well understood. To address this, we generated transgenic mice expressing a dominant-negative IL-10 receptor specifically in T cells (CD4dnIL-10Rα). We found that Foxp3-depleted CD45RBlo (regulatory T cell [Treg cell]–depleted CD45RBlo) but not CD45RBhi CD4+ T cells are controlled directly by IL-10 upon transfer into Rag1 knockout (KO) mice. Furthermore, the colitis induced by transfer of Treg cell–depleted CD45RBlo CD4+ T cells into Rag1 KO mice was characterized by reduced Th1 and increased Th17 cytokine messenger RNA levels in the colon as compared with the colitis induced by transfer of CD45RBhi T cells. In contrast to the CD45RBhi transfer colitis model, in which IL-22 is protective, we found that T cell–derived IL-22 was pathogenic upon transfer of Treg cell–depleted CD45RBlo T cells into Rag1 KO mice. Our results highlight characteristic differences between colitis induced by naive (CD45RBhi) and memory/effector (Treg cell–depleted CD45RBlo) cells and different ways that IL-22 impacts inflammatory bowel disease.

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Stability of Foxp3 Expression Is a Critical Factor In the Ability of Regulatory T Cells to Mitigate Graft Versus Host Disease

Blood ◽

10.1182/blood.v116.21.731.731 ◽

2010 ◽

Vol 116 (21) ◽

pp. 731-731

Author(s):

Amy Beres ◽

Richard Komorowski ◽

William R. Drobyski

Keyword(s):

T Cells ◽

T Cell ◽

Regulatory T Cells ◽

Critical Factor ◽

Foxp3 Expression ◽

Spleen Cells ◽

Graft Versus Host

Abstract Abstract 731 Graft versus host disease (GVHD) is a proinflammatory T cell-mediated syndrome that is the major complication of allogeneic bone marrow transplantation (BMT). During the course of GVHD, there is a progressive loss of regulatory T cells (Tregs), leading to an imbalance between the effector and regulatory arms of the immune system. Tregs have been subdivided into two distinct subsets, termed natural and induced, which have overlapping yet unique characteristics. While the role of natural regulatory T cells (nTregs) in GVHD biology has been extensively examined, the role of induced regulatory T cells (iTregs) remains largely unknown. An attractive aspect of the latter cell population is that they can be differentiated in vitro from conventional T cells and expanded in large numbers making them a potential source for regulatory T cell therapy in vivo. To determine whether in vitro-expanded iTregs were able to suppress alloreactive donor T cell responses and to compare the efficacy of these cells relative to nTregs, studies were performed using an MHC-incompatible murine BMT model (B6[H−2b]−Balb/c[H−2d]). In initial studies, purified CD4+ Foxp3EGFP– T cells obtained from B6 Foxp3EGFP reporter mice were cultured with anti-CD3 and anti-CD28 antibodies in the presence of IL-2 and TGF-b. After three days in culture, approximately 60–70% of cells were Foxp3+, expressed GITR, CD25, and CD103, and were equally suppressive to nTregs in mixed lymphocyte cultures. To determine if iTregs were suppressive in vivo, lethally irradiated Balb/c mice were transplanted with either B6 BM alone, B6 BM and spleen cells, or B6 BM/spleen cells and in vitro-expanded iTregs. In contrast to in vitro results, adoptive transfer of iTregs failed to protect mice from lethal GVHD even when administered at high Treg: effector T cell ratios (5:1) and were much less effective than equivalent doses of nTregs at abrogating GVHD pathology. iTregs also had no additive effect when co-administered with nTregs. Notably, we observed that whereas transferred nTregs persisted for up to 60 days in transplanted animals, iTregs were undetectable after only 14 days in liver, lung, colon and spleen, indicating that reduced in vivo survival was a potential explanation for the lack of protection. Further examination, however, revealed that the inability to detect iTregs was primarily attributable to the loss of Foxp3 expression and the subsequent in vivo reversion of these cells to a proinflammatory phenotype characterized by the secretion of interferon-gamma. In prior studies (Chen et al, Blood, 2009), we demonstrated that blockade of IL-6 signaling augmented reconstitution of nTregs and reduced overall GVHD severity. To determine whether inhibition of IL-6 could stabilize Foxp3 expression and prevent phenotypic reversion of iTregs, lethally irradiated Balb/c recipients were transplanted with B6 BM and spleen cells along with in vitro-differentiated iTregs and then treated with either isotype control or anti-IL-6R-specific antibody. Analysis of cells obtained from spleen, liver, lung and colon revealed that blockade of IL-6 signaling did not prevent loss of Foxp3 expression or reversion of iTregs to a Th1 cytokine phenotype. While Tregs can be converted from conventional T cells in vitro, they can also be generated in vivo during inflammatory syndromes. We therefore examined whether in vivo induction of iTregs occurred during GVHD and the extent to which blockade of IL-6 signaling affected iTreg expansion and overall GVHD protection. To address this question, lethally irradiated Balb/c mice were transplanted with B6 Rag-1 BM cells and purified CD4+ Foxp3EGFP– T cells, and then treated with either anti-IL-6R or control antibody. We observed that in vivo conversion of Tregs was negligible in control animals (<1%), but that administration of anti-IL-6R antibody significantly increased the relative and absolute number of iTregs in GVHD target tissues with a commensurate reduction in overall pathological damage. Thus, blockade of IL-6 signaling was able to enhance reconstitution of iTregs in vivo, but had no discernible affect on adoptively transferred iTregs. In summary, these studies demonstrate that the stability of Foxp3 expression is a critical factor in the maintenance of transplantation tolerance and that instability of expression limits the utility of adoptively transferred iTregs as a source of cellular therapy for the abrogation of GVHD. Disclosures: No relevant conflicts of interest to declare.

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Insulin-Like Growth Factor I Is a Positive Regulator of Thymic Function.

Blood ◽

10.1182/blood.v104.11.732.732 ◽

2004 ◽

Vol 104 (11) ◽

pp. 732-732

Author(s):

Yu-Waye Chu ◽

Shoshana Yakar ◽

Derek LeRoith ◽

Ronald Gress

Keyword(s):

T Cells ◽

Stem Cell ◽

T Cell ◽

Growth Factor ◽

Hematopoietic Stem Cell ◽

Brdu Incorporation ◽

Cell Populations ◽

Hematopoietic Stem ◽

Thymic Function

Abstract The thymus is important in the recovery and maintenance of T-cell populations following hematopoietic stem cell transplantation. Neuroendocrine growth factors have been implicated in the regulation of thymocyte development and overall thymic function through direct effects on developing thymocytes and/or modulation of supporting thymic stroma. Insulin-like growth factor 1 (IGF-1) is a ubiquitously expressed mediator of cellular anabolic function that has been shown to affect the growth of primary and secondary immune organs. We present evidence supporting the role of IGF-1 as a regulator of thymic function in vivo. Recombinant human IGF-1 was administered into healthy young adult mice by continuous infusion for up to four weeks at a dose of 100 ug/day (5 mg/kg/day). During this period, thymic function was measured by multiple parameters including thymocyte number, thymocyte subset composition, thymic T-cell rearrangement circle (TREC) content, proliferation and developmental kinetics as measured by BrdU incorporation, peripheral naïve T-cell subsets, and peripheral TREC content. Two-week infusions of IGF-1 resulted in significant increases in thymic size, cellularity, and thymic TREC numbers. Within the earliest lineage-negative thymocyte subpopulations defined by CD44 and CD25 co-expression patterns, significant increases in the DN2 through DN4 subpopulations were observed. CD4+CD8+, CD4+CD8-, and CD4-CD8+ thymic populations were also uniformly increased with IGF-1 treatment. BrdU incorporation was increased in all thymocyte subsets with IGF-1 administration, suggesting a stimulatory effect of IGF-1 on thymocyte proliferation and developmental kinetics. In peripheral T-cell populations, significant increases in the number of CD4+ and CD8+ naïve cells were observed after four weeks of IGF-1 treatment. The expansion of peripheral naïve cell populations occurred in the absence of increased cell division as measured by Ki67 expression. On the other hand, significant increases in total peripheral TREC numbers were observed at this time, indicating an increase in the number of recent thymic emigrants with IGF-1 administration. IGF-1 receptor (IGF-1R) is expressed on both thymocytes and peripheral T-cells. Mice constitutively homozygous for null mutations of IGF-1R die at birth. Consequently, to assess the role of IGF-1R signaling on thymic function and T-cell homeostasis, mice lacking IGF-1R specifically on thymocytes and T-cells were generated by cre recombinase mediated deletion of the IGF-1R high-affinity binding site (LCK-Cre/lox-IGF1R). Compared to cre-negative littermates, LCK-Cre/lox-IGF1R mice exhibited a 50% decrease in thymocyte cellularity that is due almost entirely to decreases in the CD4+CD8+ double-positive thymocyte population and in the lineage-negative DN4 subpopulation. The size of the CD4+CD8- and CD4-CD8+ thymocyte populations were not affected. Thymic TREC numbers were also decreased by 50%. Diminished thymic output in LCK-Cre/lox-IGF1R mice was manifested by a decrease in the size of splenic naïve CD4+ and CD8+ populations and splenic TREC numbers. These results demonstrate that: 1) administration of IGF-1 enhances thymic function through the expansion of thymocyte subpopulations and thymic output; and 2) IGF-1R signaling is important in the maintenance of thymocyte and peripheral T-cell populations. These findings support the concept of IGF-1 as a potentially useful agent in facilitating T-cell reconstitution following hematopoietic stem cell transplant.

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Dysfunctional T Regulatory Cells in Myeloma: Molecular Mechanisms of Dysregulation.

Blood ◽

10.1182/blood.v106.11.3462.3462 ◽

2005 ◽

Vol 106 (11) ◽

pp. 3462-3462

Author(s):

Rao H. Prabhala ◽

Paola Neri ◽

Pierfrancesco Tassone ◽

Jooeun E. Bae ◽

Masood A. Shammas ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Treg Cell ◽

Cell Function ◽

Cell Activity ◽

T Cell Responses ◽

Treg Cells ◽

Cell Responses ◽

Treg Cell Function

Abstract Multiple myeloma (MM) is characterized by production of monoclonal immunoglobulin, associated with suppressed uninvolved immunoglobulins and dysfunctional T cell responses. The biological basis of this dysfunction remains ill defined. Since T regulatory (Treg) cells play an important role in suppressing normal immune responses, we have here evaluated the potential role of Treg cells in immune dysfunction in MM. We observed a significant increase in CD4+CD25+ T cells in individuals with monoclonal gammopathy of undetermined significance (MGUS) and patients with MM compared to normal donors (25% and 26% versus 14%, respectively); however, Treg cells as measured by Foxp3 expression are significantly decreased in both MGUS (1.6±0.5%, p<0.01) and MM (1.6±0.5%, p<0.01) compared to normal donors (6.0±0.8%). Additionally, these Treg cells also do not function normally. Treg cells from patients with MM and MGUS even when added in higher proportions are unable to suppress anti-CD3-mediated T cell proliferation. This decreased number and function of Treg cells in MGUS and in MM may account, at least in part, for the non-specific increase in CD4+CD25+ T cells, thereby contributing to dysfunctional T cell responses. We have further analyzed the molecular basis for the Treg cell dysfunction in myeloma. Based on the preliminary results suggesting a role of IL-6 in Treg cell function and since both serum IL-6 and soluble IL-6 receptor levels are significantly elevated in MGUS and MM, we evaluated the role of IL-6 and its soluble receptor on Treg cell function. We observed that the addition of IL-6 and/or sIL-6 receptor to the culture leads to loss of Treg cell activity in normal donor cells similar to one observed in myeloma patients; and conversely, when Treg cells from MM patients are treated with the anti-IL-6 antibody or IL-6 receptor super antagonist, sant 7, the suppressive activity of Treg cells is restored. Additionally, we have preliminary evidence of expansion of Foxp3+ cell numbers in PBMC from MM patients following in vitro treatment with anti-IL-6 antibody. This data suggests a role of IL-6 and bone marrow microenvironment in dysfunctional Treg cells in MM and that inhibition of IL-6 signaling results in beneficial effects on T cell activity by increasing Treg cell activity. A blockade of IL-6 signaling thus emerges as a potential approach to establish immune homeostasis to improve immune function in MM.

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Notch Is a Novel Critical Signaling Pathway Regulating Responses of T Cell and Antigen Presenting Cells in Multiple Murine aGVHD Models

Blood ◽

10.1182/blood.v126.23.5418.5418 ◽

2015 ◽

Vol 126 (23) ◽

pp. 5418-5418

Author(s):

Xiaodan Luo ◽

Pengfei Qin ◽

Chunyan Wang ◽

Zhenqian Huang ◽

Huo Tan

Keyword(s):

T Cells ◽

Body Weight ◽

T Cell ◽

Signaling Pathway ◽

Antigen Presenting Cells ◽

Hematopoietic Stem ◽

Secretase Inhibitor ◽

Donor T Cells ◽

Antigen Presenting

Abstract Introduction: Acute graft-versus-host disease (aGVHD) is a potentially life-threatening complication mediated by both host-derived antigen presenting cells (APCs) and donor T cells after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Despite prophylaxis and treatments, aGVHD stell occurs in many allo-HSCT patients. The role of Notch1 signal inhibition becomes more and more important in aGVHD study. This study is to investigate the role of Notch1 inhibition by γ-secretase inhibitor DAPT in murine aGVHD model. Methods: We established a C57BL/6 BALB/c murine aGVHD model. γ-secretase inhibitor-DAPT is used to inhibit Notch1 signal in vivo and in vitro before transplantation. The degree of clinical and histopathologic GVHD is assessed by aGVHD scores and body weight. The functions of host-derived APCs and donor T cells are analyzed by flow cytometry, ELISA and PCR. Results: All mice survived at least 14 days after transplantation and all of them developed aGVHD (n=20). The expression of Hes-1, as one of the target genes of Notch1 signal pathway, decreased significantly after DAPT inhibition. Body weight of mice in control groups decreased significantly compared to mice with Notch1 inhibition by DAPT after transplantation. Notch1 inhibited recipients produced markedly decreased amounts of the pro-inflammatory cytokines IFN-γ. The expressions of CD4 and Foxp3 increased while CD11c, CD80 and CD86 decreased after Notch1 inhibition. Conclusions: These results indicate that Notch is a novel critical signaling pathway regulating responses of T cell and antigen presenting cells in multiple murine aGVHD models. Notch signaling inhibition appears to limit the harmful effects of aGVHD. Disclosures No relevant conflicts of interest to declare.

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Human Wnt/β-Catenin Regulates Alloimmune Signaling during Allogeneic Transplantation

Cancers ◽

10.3390/cancers13153798 ◽

2021 ◽

Vol 13 (15) ◽

pp. 3798

Author(s):

Mahinbanu Mammadli ◽

Rebecca Harris ◽

Sara Mahmudlu ◽

Anjali Verma ◽

Adriana May ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Cell Biology ◽

Allogeneic Transplantation ◽

Hematopoietic Stem ◽

Inflammatory Cytokine Production ◽

Formidable Challenge ◽

T Cell Biology ◽

Donor T Cells

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is one of the most widely applied forms of adoptive immunotherapy for the treatment of hematological malignancies. Detrimental graft-versus-host disease (GVHD), but also beneficial graft-versus-leukemia (GVL) effects occurring after allo-HSCT are largely mediated by alloantigen-reactive donor T cells in the graft. Separating GVHD from GVL effects is a formidable challenge, and a greater understanding of donor T cell biology is required to accomplish the uncoupling of GVHD from GVL. Here, we evaluated the role of β-catenin in this process. Using a unique mouse model of transgenic overexpression of human β-catenin (Cat-Tg) in an allo-HSCT model, we show here that T cells from Cat-Tg mice did not cause GVHD, and surprisingly, Cat-Tg T cells maintained the GVL effect. Donor T cells from Cat-Tg mice exhibited significantly lower inflammatory cytokine production and reduced donor T cell proliferation, while upregulating cytotoxic mediators that resulted in enhanced cytotoxicity. RNA sequencing revealed changes in the expression of 1169 genes for CD4, and 1006 genes for CD8+ T cells involved in essential aspects of immune response and GVHD pathophysiology. Altogether, our data suggest that β-catenin is a druggable target for developing therapeutic strategies to reduce GVHD while preserving the beneficial GVL effects following allo-HSCT treatment.

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