Granzyme B Produced by Human Plasmacytoid Dendritic Cells Suppresses T Cell Expansion.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2674-2674
Author(s):  
Dorit Fabricius ◽  
Angelika Vollmer ◽  
Sue Blackwell ◽  
Julia Maier ◽  
Kai Sontheimer ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Conflicts Of Interest ◽  
Granzyme B ◽  
Cd40 Ligand ◽  
Plasmacytoid Dendritic Cells ◽  
Transcriptional Level ◽  
Independent Manner ◽  
Complex Interactions

Abstract Abstract 2674 Poster Board II-650 Human plasmacytoid dendritic cells (pDC) play a central role in regulating adaptive T cell responses in the course of neoplastic, viral and autoimmune disorders. In several of these diseases, elevated extracellular levels of the serine protease granzyme B (GrB) are observed. We found that human pDC can be an abundant source of GrB based on FACS analysis, ELISpot, ELISA, Sensizyme, Western immunoblotting, RT-PCR, and fluorescence microscopy. GrB is actively secreted by pDCs and reaches maximal levels up to two logs higher than those produced by classical GrB producers such as CTL or NK cells. However, pDC GrB production is not accompanied by perforin secretion. Spinning disk confocal microscopy revealed that GrB+ pDC bind to and transfer active GrB to T cells. Importantly, this GrB transfer induces a suppression of T cell proliferation in a GrB-dependent, perforin-independent manner, a process reminiscent of regulatory T cells. GrB expression in pDC is regulated on a transcriptional level by JAK1, STAT3 and STAT5. IL-3 and IL-10 enhance GrB production by pDCs while GrB production is inhibited by toll-like-receptor agonists and CD40 ligand. These findings suggest that GrB production by pDCs is involved in the complex interactions between pDC and T cells and that GrB-secreting pDC may play a regulatory role related to anti-tumor immunity, anti-viral immune responses, and autoimmune processes. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Granzyme B produced by human plasmacytoid dendritic cells suppresses T-cell expansion

Blood ◽  
2010 ◽  
Vol 115 (6) ◽  
pp. 1156-1165 ◽  
Author(s):  
Bernd Jahrsdörfer ◽  
Angelika Vollmer ◽  
Sue E. Blackwell ◽  
Julia Maier ◽  
Kai Sontheimer ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Granzyme B ◽  
Cd40 Ligand ◽  
Plasmacytoid Dendritic Cells ◽  
Janus Kinase ◽  
Transcriptional Level ◽  
Activated T Cells ◽  
Independent Manner

Abstract Human plasmacytoid dendritic cells (pDCs) are crucially involved in the modulation of adaptive T-cell responses in the course of neoplastic, viral, and autoimmune disorders. In several of these diseases elevated extracellular levels of the serine protease granzyme B (GrB) are observed. Here we demonstrate that human pDCs can be an abundant source of GrB and that such GrB+ pDCs potently suppress T-cell proliferation in a GrB-dependent, perforin-independent manner, a process reminiscent of regulatory T cells. Moreover, we show that GrB expression is strictly regulated on a transcriptional level involving Janus kinase 1 (JAK1), signal transducer and activator of transcription 3 (STAT3), and STAT5 and that interleukin-3 (IL-3), a cytokine secreted by activated T cells, plays a central role for GrB induction. Moreover, we find that the immunosuppressive cytokine IL-10 enhances, while Toll-like receptor agonists and CD40 ligand strongly inhibit, GrB secretion by pDCs. GrB-secreting pDCs may play a regulatory role for immune evasion of tumors, antiviral immune responses, and autoimmune processes. Our results provide novel information about the complex network of pDC–T-cell interactions and may contribute to an improvement of prophylactic and therapeutic vaccinations.

scholarly journals Generation of Human CD8 T Regulatory Cells by CD40 Ligand–activated Plasmacytoid Dendritic Cells

2002 ◽  
Vol 195 (6) ◽  
pp. 695-704 ◽  
Author(s):  
Michel Gilliet ◽  
Yong-Jun Liu
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Transforming Growth Factor ◽  
Cd8 T Cell ◽  
Cd40 Ligand ◽  
Plasmacytoid Dendritic Cells ◽  
Target Cells ◽  
Ifn Γ

Although CD8 T cell–mediated immunosuppression has been a well-known phenomenon during the last three decades, the nature of primary CD8 T suppressor cells and the mechanism underlying their generation remain enigmatic. We demonstrated that naive CD8 T cells primed with allogeneic CD40 ligand–activated plasmacytoid dendritic cells (DC)2 differentiated into CD8 T cells that displayed poor secondary proliferative and cytolytic responses. By contrast, naive CD8 T cells primed with allogeneic CD40 ligand–activated monocyte-derived DCs (DC1) differentiated into CD8 T cells, which proliferated to secondary stimulation and killed allogeneic target cells. Unlike DC1-primed CD8 T cells that produced large amounts of interferon (IFN)-γ upon restimulation, DC2-primed CD8 T cells produced significant amounts of interleukin (IL)-10, low IFN-γ, and no IL-4, IL-5, nor transforming growth factor (TGF)-β. The addition of anti–IL-10–neutralizing monoclonal antibodies during DC2 and CD8 T cell coculture, completely blocked the generation of IL-10–producing anergic CD8 T cells. IL-10–producing CD8 T cells strongly inhibit the allospecific proliferation of naive CD8 T cells to monocytes, and mature and immature DCs. This inhibition was mediated by IL-10, but not by TGF-β. IL-10–producing CD8 T cells could inhibit the bystander proliferation of naive CD8 T cells, provided that they were restimulated nearby to produce IL-10. IL-10–producing CD8 T cells could not inhibit the proliferation of DC1-preactivated effector T cells. This study demonstrates that IL-10–producing CD8 T cells are regulatory T cells, which provides a cellular basis for the phenomenon of CD8 T cell–mediated immunosuppression and suggests a role for plasmacytoid DC2 in immunological tolerance.

Regulation of Plasmacytoid Dendritic Cells by Commonly Used Anti-Viral Vaccines,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3221-3221
Author(s):  
Bernd Jahrsdörfer ◽  
Benedikt Nussbaum ◽  
Birgit Mandel ◽  
Daniel Busch ◽  
Verena Panitz ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Conflicts Of Interest ◽  
T Cell Responses ◽  
Plasmacytoid Dendritic Cells ◽  
T Cell Proliferation ◽  
Cell Responses ◽  
Viral Vaccines

Abstract Abstract 3221 Human plasmacytoid dendritic cells (pDC) play a central role in regulating adaptive T cell responses in the course of neoplastic, viral and autoimmune disorders. Recently, we demonstrated that apart from their pro-inflammatory effects, which are mainly mediated by secretion of interferon-alpha (IFN-a), pDC can also exhibit potent anti-inflammatory functions borne by active secretion of granzyme B (GrB), a serine protease classically known from CTL, NK cells and regulatory T cells. Here, we hypothesized, that commonly used anti-viral vaccines may affect pDC on several levels including their immunophenotype as well as their capacity to secrete either IFN-a or GrB. Using various methods including FACS, ELISpot, ELISA, spinning-disk confocal microscopy and RT-PCR, we could demonstrate that various anti-viral vaccines including vaccines against TBEV, yellow fever, polio, measles, rotavirus, varicella and hepatitis B were able to affect pDC by modulating expression of a series of surface molecules involved in cell adhesion, antigen-presentation and co-stimulation. In addition, major differences between the vaccines were found in terms of their effects on secretion of IFN-a and GrB. Interestingly, while only one vaccine, FSME Immun (TBEV) induced substantial IFN-a responses in pDC, all others tested did not. In contrast, virtually all vaccines tested induced more or less strong suppression of GrB secretion by maturing pDC. Of note, pDC that secreted high amounts of GrB induced by far lower allogeneic T cell proliferation as compared to pDC that secreted little or no GrB. Moreover, suppression of pDC-derived GrB by a substrate-specific GrB inhibitor resulted in significant enhancement of T cell proliferation in co-cultures of GrB-secreting pDC with allogeneic T cells. Our data demonstrate 1) that anti-viral vaccines may have distinct effects on both the immunophenotype and the secretory potential of pDC, and 2) that GrB is an important novel variable affecting the capacity of pDC to either trigger or dampen adaptive T cell responses. Our results may have implications for further study of the role pDC play in the regulation of adaptive immune responses, and for the potential application of this knowledge in the development of novel adjuvants admixed to anti-viral vaccines. Disclosures: No relevant conflicts of interest to declare.

Activated T cells enhance interferon-α production by plasmacytoid dendritic cells stimulated with RNA-containing immune complexes

2015 ◽  
Vol 75 (9) ◽  
pp. 1728-1734 ◽  
Author(s):  
Dag Leonard ◽  
Maija-Leena Eloranta ◽  
Niklas Hagberg ◽  
Olof Berggren ◽  
Karolina Tandre ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Immune Complexes ◽  
Serum Levels ◽  
Plasmacytoid Dendritic Cells ◽  
Interferon Α ◽  
Type I ◽  
Activated T Cells ◽  
Gm Csf

ObjectivesPatients with systemic lupus erythematosus (SLE) have an ongoing interferon-α (IFN-α) production by plasmacytoid dendritic cells (pDCs). We investigated whether T cells can promote IFN-α production by pDCs.MethodsHuman pDCs were stimulated with immune complexes (ICs) containing U1 small nuclear ribonucleic proteins particles and SLE-IgG (RNA-IC) in the presence of T cells or T cell supernatants. T cells were activated by anti-CD3/CD28 antibodies or in a mixed leucocyte reaction. IFN-α and other cytokines were determined in culture supernatants or patient sera with immunoassays. The effect of interleukin (IL) 3 and granulocyte-macrophage-colony-stimulating factor (GM-CSF) on pDCs was examined by the use of antibodies, and the expression of CD80/CD86 was determined using flow cytometry.ResultsActivated T cells and supernatants from activated T cells increased IFN-α production by >20-fold. The stimulatory effect of T cell supernatants was reduced after depletion of GM-CSF (81%) or by blocking the GM-CSF receptor (55%–81%). Supernatant from activated T cells, furthermore, increased the frequency of CD80 and CD86 expressing pDCs stimulated with RNA-IC from 6% to 35% (p<0.05) and from 10% to 26% (p<0.01), respectively. Activated SLE T cells enhanced IFN-α production to the same extent as T cells from healthy individuals and a subset of patients with SLE had increased serum levels of GM-CSF.ConclusionsActivated T cells enhance IFN-α production by RNA-IC stimulated pDCs via GM-CSF and induce pDC maturation. Given the increased serum levels of GM-CSF in a subset of patients with SLE, these findings suggest that activated T cells may upregulate type I IFN production in SLE.

scholarly journals Transcription Factor Gene Fluorescent Reporters Track Lineage Fate in Lymphoid Commitment

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. SCI-3-SCI-3
Author(s):  
Ellen Rothenberg
Keyword(s):  
T Cells ◽  
Transcription Factor ◽  
T Cell ◽  
Conflicts Of Interest ◽  
Epigenetic Modification ◽  
Single Cells ◽  
Transcriptional Level ◽  
Regulatory State ◽  
Cell Commitment

Abstract The transition from multipotency to lineage commitment can be followed with particular clarity for T cell precursors. In this lineage, the role of environmental signals can be clearly separated from the role of intrinsic fate programming in individual cells and the cells' developmental responses to changing conditions and can be tracked in real time. T cell precursors are still multipotent when they first enter the thymus, and if they are removed from the thymic microenvironment at this stage they can give rise to non-T cells including dendritic cells and myeloid cells. For multiple cell divisions, they preserve this multipotency and are only kept in line to become T cells conditionally, by Notch signaling from the thymic stroma. Then at a specific point of no return, the cells become unable to give rise to anything except T cells regardless of environment, and this is the point of commitment. Commitment is clearly the readout of a change in internal transcriptional regulatory state. To determine how this is controlled, we and others have charted transcription factor expression changes across this interval and changes in chromatin modification and DNA accessibility that accompany the transition, and we have been able to use functional perturbation tests to narrow down the key regulators that catalyze and enforce this transition. A particularly important commitment factor is encoded by the Bcl11b gene, which is released from previously repressed chromatin and sharply activated at the transcriptional level just as the cells become committed. The Bcl11b gene product is required in all alpha beta and most gamma delta T cells to enable the commitment process to occur. These properties make it highly illuminating as an indicator of the regulatory state in individual differentiating T-cell precursors. We have generated a series of knock-in Bcl11b fluorescent reporter alleles to probe the correlation of Bcl11b expression with changes in specific target genes, to determine the transcription factor requirements for Bcl11b gene activation in the gene regulatory network controlling commitment in single cells, and to measure the role of epigenetic modification of the Bcl11b locus on the kinetics of transition from uncommitted to committed states. These results and their implications will be presented. Importantly, the use of live-cell reporters reveals a level of all-or-none, stochastic regulation in the responses of individual cells to combinatorial transcription factor action at this developmental watershed1. Reference: 1. Kueh HY, Yui MA, Ng KKH, et al. Asynchronous combinatorial action of four regulatory factors activates Bcl11b for T cell commitment. Nature Immunology. 2016.17, 956-965. Disclosures No relevant conflicts of interest to declare.

scholarly journals Effector T Cells Boost Regulatory T Cell Expansion by IL-2, TNF, OX40, and Plasmacytoid Dendritic Cells Depending on the Immune Context

2014 ◽  
Vol 194 (3) ◽  
pp. 999-1010 ◽  
Author(s):  
Audrey Baeyens ◽  
David Saadoun ◽  
Fabienne Billiard ◽  
Angéline Rouers ◽  
Sylvie Grégoire ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Regulatory T Cell ◽  
Cell Expansion ◽  
Plasmacytoid Dendritic Cells ◽  
Effector T Cells ◽  
T Cell Expansion

Failure of Plasmacytoid Dendritic Cells to Drive Functional Regulatory T Cell Differentiation In Human Disease

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1746-1746
Author(s):  
Derek NJ Hart ◽  
Xinsheng Ju ◽  
Yitian Ding ◽  
Maryam Azlan ◽  
Georgina Clark
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Cd4 T Cells ◽  
Conflicts Of Interest ◽  
Topical Therapy ◽  
Type I ◽  
Indoleamine 2 ◽  
Disease States

Abstract Abstract 1746 Dendritic cells (DC) initiate and direct immune responses. Plasmacytoid DC (pDC) produce type I interferon (IFN) in response to viral and/or bacterial challenge and have been implicated in the pathogenesis of a number of chronic inflammatory disesases. pDC also induce the generation of regulatory T cells (Treg) from naïve CD4+ T cells. Treg control the balance of T cell responses and maintain immune homeostasis so the primary contribution of pDC to Treg induction in disease states is an important question. Psoriasis is a common chronic inflammatory skin disease associated with over expression of type I IFN–inducible genes in patients and this response apparently overwhelms any pDC regulatory contribution. The numbers of peripheral blood Treg cells and pDC are decreased in psoriatic patients but why pDC fail to generate a significant counterbalancing Treg response in psoriasis is unknown. We compared purified blood pDC from normal donors with those from psoriasis donors, who were well and either untreated or only on topical therapy at the time of donation, for their ability to induce Treg. Freshly purified pDC from both normal and psoriasis donors lacked CD80 and CD83, but expressed similar amounts of HLA-DR and CD86. pDC from healthy psoriasis donors, when stimulated with TLR9 ligands, secreted high amounts of IL-6, expressed little surface inducible costimulator-ligand (ICOS-L, CD275) and exhibited low indoleamine 2, 3–dioxygenase (IDO) enzymatic activity compared to normal controls. To assess the capacity of pDC from psoriasis patients to induce the differentiation of naïve CD4+ T cells, we performed sequential co-culture experiments. In these, TLR-9 stimulated pDC from psoriasis patients failed to induce the differentiation of naïve CD4+ T cells into IL-10 secreting, functional regulatory T cells. In addition, the CD4+CD25+ T cells resulting from co-culture with psoriasis pDC were less effective in suppressing an allogeneic MLR. In contrast, the pDC from psoriasis patients, unlike those from normal donors, induced T cells capable of secreting IL-22. Further investigation as to why psoriatic pDC failed to induce Treg showed that the differentiated CD4+CD25+ (CD127-) T cells from these co-cultures expressed dramatically less Foxp3 following priming with psoriatic pDC. Exogenous kynurenine, to replace IDO restored the ability of psoriatic pDC to induce Treg. In conclusion, our data demonstrate that aberrant pDC produce dysfunctional Treg development in psoriasis and highlights the contributions of IL-6/IDO/CD275/IL-22 to the disease pathogenesis. This insight is now driving studies in other disease states, notably graft versus host disease, where our focus is on manipulating DC for therapeutic benefit. Disclosures: No relevant conflicts of interest to declare.

Maturation of Monocyte-Derived Dendritic Cells Induced by Dual Targeting of Endocytosis Receptor and CD40

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4293-4293
Author(s):  
Matthias Staudinger ◽  
Inga Rebekka Catenhusen ◽  
Matthias Peipp ◽  
Christian Kellner ◽  
Roland Repp ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Conflicts Of Interest ◽  
Specific Binding ◽  
Surface Expression ◽  
Tumor Vaccination ◽  
Dual Targeting

Abstract Abstract 4293 Dendritic cells (DC) are professional antigen presenting cells, which can induce and regulate adaptive immune responses. Hence, DC are attractive candidates for anti-tumor vaccination strategies. The aim of this study was to create recombinant bispecific Fab to target and activate DC, by using derivates of monovalent agonistic CD40 scFv and endocytosis receptors FcγRI (CD64), FcαR (CD89), DEC205 (CD205), as well as DC-SIGN (CD209), respectively. The recombinant bispecific molecules were expressed in the eukaryotic cell line Hek 293T, enriched to homogeneity by affinity chromatography and analyzed for specific binding to the targeted receptors. Functional analysis of these molecules indicated, that concurrent engagement of CD40 and endocytosis receptors expressed by immature IL-4/GM-CSF cultured monocytes-derived DC induced maturation of these DC. This was suggested by changes of DC phenotype, especially enhanced surface expression of co-stimulatory molecules like CD80 and CD86, and gain of CD83 expression as measured by flow cytometry. To particularly analyze the T cell activation properties of these DC, MACS enriched T cells of autologous donors were co-cultivated with antigen-loaded and subsequently maturated DC for seven days. After restimulation of T cells, flow cytometric detection of intracellular IFN-γ levels and surface expression of CD69 used as markers for T cell activation implies, that these DC exhibit elevated properties to activate antigen specific T cells. In contrast, a bispecific [CD64scFv × CD40scFv] Fab used as control failed to induce maturation of DC lacking CD64 expression, which may be due to abrogation of bivalent binding of this molecule and therefore CD40 cross-linking. Moreover, dual targeting with these novel immunoconstructs when linked to peptides of a tumor-associated antigen may allow direct antigen delivery to DC in combination with DC activation, and thus anti-tumor vaccination in vivo. Disclosures: No relevant conflicts of interest to declare.

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