TY - JOUR A1 - Badr, Mohammad A1 - McFleder, Rhonda L. A1 - Wu, Jingjing A1 - Knorr, Susanne A1 - Koprich, James B. A1 - Hünig, Thomas A1 - Brotchie, Jonathan M. A1 - Volkmann, Jens A1 - Lutz, Manfred B. A1 - Ip, Chi Wang T1 - Expansion of regulatory T cells by CD28 superagonistic antibodies attenuates neurodegeneration in A53T-α-synuclein Parkinson’s disease mice JF - Journal of Neuroinflammation N2 - Background Regulatory CD4\(^+\)CD25\(^+\)FoxP3\(^+\) T cells (Treg) are a subgroup of T lymphocytes involved in maintaining immune balance. Disturbance of Treg number and impaired suppressive function of Treg correlate with Parkinson’s disease severity. Superagonistic anti-CD28 monoclonal antibodies (CD28SA) activate Treg and cause their expansion to create an anti-inflammatory environment. Methods Using the AAV1/2-A53T-α-synuclein Parkinson’s disease mouse model that overexpresses the pathogenic human A53T-α-synuclein (hαSyn) variant in dopaminergic neurons of the substantia nigra, we assessed the neuroprotective and disease-modifying efficacy of a single intraperitoneal dose of CD28SA given at an early disease stage. Results CD28SA led to Treg expansion 3 days after delivery in hαSyn Parkinson’s disease mice. At this timepoint, an early pro-inflammation was observed in vehicle-treated hαSyn Parkinson’s disease mice with elevated percentages of CD8\(^+\)CD69\(^+\) T cells in brain and increased levels of interleukin-2 (IL-2) in the cervical lymph nodes and spleen. These immune responses were suppressed in CD28SA-treated hαSyn Parkinson’s disease mice. Early treatment with CD28SA attenuated dopaminergic neurodegeneration in the SN of hαSyn Parkinson’s disease mice accompanied with reduced brain numbers of activated CD4\(^+\), CD8\(^+\) T cells and CD11b\(^+\) microglia observed at the late disease-stage 10 weeks after AAV injection. In contrast, a later treatment 4 weeks after AAV delivery failed to reduce dopaminergic neurodegeneration. Conclusions Our data indicate that immune modulation by Treg expansion at a timepoint of overt inflammation is effective for treatment of hαSyn Parkinson’s disease mice and suggest that the concept of early immune therapy could pose a disease-modifying option for Parkinson’s disease patients. KW - Parkinson’s disease KW - neuroinflammation KW - T cells KW - regulatory T cells KW - neuroprotection Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-300580 VL - 19 ER - TY - JOUR A1 - Chopra, Martin A1 - Lang, Isabell A1 - Salzmann, Steffen A1 - Pachel, Christina A1 - Kraus, Sabrina A1 - Bäuerlein, Carina A. A1 - Brede, Christian A1 - Jordán Garrote, Ana-Laura A1 - Mattenheimer, Katharina A1 - Ritz, Miriam A1 - Schwinn, Stefanie A1 - Graf, Carolin A1 - Schäfer, Viktoria A1 - Frantz, Stefan A1 - Einsele, Hermann A1 - Wajant, Harald A1 - Beilhack, Andreas T1 - Tumor Necrosis Factor Induces Tumor Promoting and Anti-Tumoral Effects on Pancreatic Cancer via TNFR1 JF - PLoS ONE N2 - Multiple activities are ascribed to the cytokine tumor necrosis factor (TNF) in health and disease. In particular, TNF was shown to affect carcinogenesis in multiple ways. This cytokine acts via the activation of two cell surface receptors, TNFR1, which is associated with inflammation, and TNFR2, which was shown to cause anti-inflammatory signaling. We assessed the effects of TNF and its two receptors on the progression of pancreatic cancer by in vivo bioluminescence imaging in a syngeneic orthotopic tumor mouse model with Panc02 cells. Mice deficient for TNFR1 were unable to spontaneously reject Panc02 tumors and furthermore displayed enhanced tumor progression. In contrast, a fraction of wild type (37.5%), TNF deficient (12.5%), and TNFR2 deficient mice (22.2%) were able to fully reject the tumor within two weeks. Pancreatic tumors in TNFR1 deficient mice displayed increased vascular density, enhanced infiltration of CD4+ T cells and CD4+ forkhead box P3 (FoxP3)+ regulatory T cells (Treg) but reduced numbers of CD8+ T cells. These alterations were further accompanied by transcriptional upregulation of IL4. Thus, TNF and TNFR1 are required in pancreatic ductal carcinoma to ensure optimal CD8+ T cell-mediated immunosurveillance and tumor rejection. Exogenous systemic administration of human TNF, however, which only interacts with murine TNFR1, accelerated tumor progression. This suggests that TNFR1 has basically the capability in the Panc02 model to trigger pro-and anti-tumoral effects but the spatiotemporal availability of TNF seems to determine finally the overall outcome. KW - Bioluminescence KW - cancer treatment KW - cell staining KW - cytokines KW - immune cells KW - metastasis KW - regulatory T cells KW - T cells Y1 - 2013 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-97246 ER - TY - JOUR A1 - Cochain, Clement A1 - Chaudhari, Sweena M. A1 - Koch, Miriam A1 - Wiendl, Heinz A1 - Eckstein, Hans-Henning A1 - Zernecke, Alma T1 - Programmed Cell Death-1 Deficiency Exacerbates T Cell Activation and Atherogenesis despite Expansion of Regulatory T Cells in Atherosclerosis-Prone Mice JF - PLoS ONE N2 - T cell activation represents a double-edged sword in atherogenesis, as it promotes both pro-inflammatory T cell activation and atheroprotective Foxp3(+) regulatory T cell (Treg) responses. Here, we investigated the role of the co-inhibitory receptor programmed cell death-1 (PD-1) in T cell activation and CD4(+) T cell polarization towards pro-atherogenic or atheroprotective responses in mice. Mice deficient for both low density lipoprotein receptor and PD-1 (Ldlr(-/-)Pd1(-/-)) displayed striking increases in systemic CD4(+) and CD8(+) T cell activation after 9 weeks of high fat diet feeding, associated with an expansion of both pro-atherogenic IFNγ-secreting T helper 1 cells and atheroprotective Foxp3+ Tregs. Importantly, PD-1 deficiency did not affect Treg suppressive function in vitro. Notably, PD-1 deficiency exacerbated atherosclerotic lesion growth and entailed a massive infiltration of T cells in atherosclerotic lesions. In addition, aggravated hypercholesterolemia was observed in Ldlr(-/-)Pd1(-/-) mice. In conclusion, we here demonstrate that although disruption of PD-1 signaling enhances both pro- and anti-atherogenic T cell responses in Ldlr(-/-) mice, pro-inflammatory T cell activation prevails and enhances dyslipidemia, vascular inflammation and atherosclerosis. KW - nutritional deficiencies KW - atherosclerosis KW - spleen KW - aorta KW - diet KW - cytotoxic T cells KW - regulatory T cells KW - T cells Y1 - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-119823 SN - 1932-6203 VL - 9 IS - 4 ER - TY - JOUR A1 - Döhler, Anja A1 - Schneider, Theresa A1 - Eckert, Ina A1 - Ribechini, Eliana A1 - Andreas, Nico A1 - Riemann, Marc A1 - Reizis, Boris A1 - Weih, Falk A1 - Lutz, Manfred B. T1 - RelB\(^{+}\) Steady-State Migratory Dendritic Cells Control the Peripheral Pool of the Natural Foxp3\(^{+}\) Regulatory T Cells JF - Frontiers in Immunology N2 - Thymus-derived natural Foxp3\(^{+}\) CD4\(^{+}\) regulatory T cells (nTregs) play a key role in maintaining immune tolerance and preventing autoimmune disease. Several studies indicate that dendritic cells (DCs) are critically involved in the maintenance and proliferation of nTregs. However, the mechanisms how DCs manage to keep the peripheral pool at constant levels remain poorly understood. Here, we describe that the NF-κB/Rel family transcription factor RelB controls the frequencies of steady-state migratory DCs (ssmDCs) in peripheral lymph nodes and their numbers control peripheral nTreg homeostasis. DC-specific RelB depletion was investigated in CD11c-Cre × RelB\(^{fl/fl}\) mice (RelB\(^{DCko}\)), which showed normal frequencies of resident DCs in lymph nodes and spleen while the subsets of CD103\(^{-}\) Langerin\(^{-}\) dermal DCs (dDCs) and Langerhans cells but not CD103\(^{+}\) Langerin\(^{+}\) dDC of the ssmDCs in skin-draining lymph nodes were increased. Enhanced frequencies and proliferation rates were also observed for nTregs and a small population of CD4\(^{+}\) CD44\(^{high}\) CD25\(^{low}\) memory-like T cells (Tml). Interestingly, only the Tml but not DCs showed an increase in IL-2-producing capacity in lymph nodes of RelB\(^{DCko}\) mice. Blocking of IL-2 in vivo reduced the frequency of nTregs but increased the Tml frequencies, followed by a recovery of nTregs. Taken together, by employing RelB\(^{DCko}\) mice with increased frequencies of ssmDCs our data indicate a critical role for specific ssmDC subsets for the peripheral nTreg and IL-2\(^{+}\) Tml frequencies during homeostasis. KW - lymph nodes KW - dendritic cells KW - RelB KW - regulatory T cells KW - IL-2 Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-158121 VL - 8 IS - 726 ER - TY - JOUR A1 - Heuser, Christoph A1 - Gotot, Janine A1 - Piotrowski, Eveline Christina A1 - Philipp, Marie-Sophie A1 - Courrèges, Christina Johanna Felicia A1 - Otte, Martin Sylvester A1 - Guo, Linlin A1 - Schmid-Burgk, Jonathan Leo A1 - Hornung, Veit A1 - Heine, Annkristin A1 - Knolle, Percy Alexander A1 - Garbi, Natalio A1 - Serfling, Edgar A1 - Evaristo, César A1 - Thaiss, Friedrich A1 - Kurts, Christian T1 - Prolonged IKK\(\beta\) Inhibition Improves Ongoing CTL Antitumor Responses by Incapacitating Regulatory T Cells JF - Cell Reports N2 - Regulatory T cells (Tregs) prevent autoimmunity but limit antitumor immunity. The canonical NF-\(\kappa\)B signaling pathway both activates immunity and promotes thymic Treg development. Here, we report that mature Tregs continue to require NF-\(\kappa\)B signaling through I\(\kappa\)B-kinase \(\beta\) (IKK\(\beta\)) after thymic egress. Mice lacking IKK\(\beta\) in mature Tregs developed scurfy-like immunopathology due to death of peripheral FoxP3\(^+\) Tregs. Also, pharmacological IKK\(\beta\) inhibition reduced Treg numbers in the circulation by ~50% and downregulated FoxP3 and CD25 expression and STAT5 phosphorylation. In contrast, activated cytotoxic T lymphocytes (CTLs) were resistant to IKK\(\beta\) inhibition because other pathways, in particular nuclear factor of activated T cells (NFATc1) signaling, sustained their survival and expansion. In a melanoma mouse model, IKK\(\beta\) inhibition after CTL cross-priming improved the antitumor response and delayed tumor growth. In conclusion, prolonged IKK\(\beta\) inhibition decimates circulating Tregs and improves CTL responses when commenced after tumor vaccination, indicating that IKK\(\beta\) represents a druggable checkpoint. KW - medicine KW - regulatory T cells KW - NF-\(\kappa\)B pathway KW - tumor vaccination KW - checkpoint inhibition KW - cytotoxic T cells KW - cross-priming KW - apoptosis KW - tumor immunology KW - melanoma Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-173643 VL - 21 IS - 3 ER - TY - JOUR A1 - Jost, Priska A1 - Klein, Franziska A1 - Brand, Benjamin A1 - Wahl, Vanessa A1 - Wyatt, Amanda A1 - Yildiz, Daniela A1 - Boehm, Ulrich A1 - Niemeyer, Barbara A. A1 - Vaeth, Martin A1 - Alansary, Dalia T1 - Acute downregulation but not genetic ablation of murine MCU impairs suppressive capacity of regulatory CD4 T cells JF - International Journal of Molecular Sciences N2 - By virtue of mitochondrial control of energy production, reactive oxygen species (ROS) generation, and maintenance of Ca\(^{2+}\) homeostasis, mitochondria play an essential role in modulating T cell function. The mitochondrial Ca\(^{2+}\) uniporter (MCU) is the pore-forming unit in the main protein complex mediating mitochondrial Ca\(^{2+}\) uptake. Recently, MCU has been shown to modulate Ca\(^{2+}\) signals at subcellular organellar interfaces, thus fine-tuning NFAT translocation and T cell activation. The mechanisms underlying this modulation and whether MCU has additional T cell subpopulation-specific effects remain elusive. However, mice with germline or tissue-specific ablation of Mcu did not show impaired T cell responses in vitro or in vivo, indicating that ‘chronic’ loss of MCU can be functionally compensated in lymphocytes. The current work aimed to specifically investigate whether and how MCU influences the suppressive potential of regulatory CD4 T cells (Treg). We show that, in contrast to genetic ablation, acute siRNA-mediated downregulation of Mcu in murine Tregs results in a significant reduction both in mitochondrial Ca\(^{2+}\) uptake and in the suppressive capacity of Tregs, while the ratios of Treg subpopulations and the expression of hallmark transcription factors were not affected. These findings suggest that permanent genetic inactivation of MCU may result in compensatory adaptive mechanisms, masking the effects on the suppressive capacity of Tregs. KW - mitochondrial calcium uniporter KW - regulatory T cells KW - suppressive capacity Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-313621 SN - 1422-0067 VL - 24 IS - 9 ER - TY - JOUR A1 - Kraft, Peter A1 - Drechsler, Christiane A1 - Schuhmann, Michael K. A1 - Gunreben, Ignaz A1 - Kleinschnitz, Christoph T1 - Characterization of Peripheral Immune Cell Subsets in Patients with Acute and Chronic Cerebrovascular Disease: A Case-Control Study JF - International Journal of Molecular Science N2 - Immune cells (IC) play a crucial role in murine stroke pathophysiology. However, data are limited on the role of these cells in ischemic stroke in humans. We therefore aimed to characterize and compare peripheral IC subsets in patients with acute ischemic stroke/transient ischemic attack (AIS/TIA), chronic cerebrovascular disease (CCD) and healthy volunteers (HV). We conducted a case-control study of patients with AIS/TIA (n = 116) or CCD (n = 117), and HV (n = 104) who were enrolled at the University Hospital Würzburg from 2010 to 2013. We determined the expression and quantity of IC subsets in the three study groups and performed correlation analyses with demographic and clinical parameters. The quantity of several IC subsets differed between the AIS/TIA, CCD, and HV groups. Several clinical and demographic variables independently predicted the quantity of IC subsets in patients with AIS/TIA. No significant changes in the quantity of IC subsets occurred within the first three days after AIS/TIA. Overall, these findings strengthen the evidence for a pathophysiologic role of IC in human ischemic stroke and the potential use of IC-based biomarkers for the prediction of stroke risk. A comprehensive description of IC kinetics is crucial to enable the design of targeted treatment strategies. KW - chronic cerebrovascular disease KW - lymphocytes KW - leukocytes KW - immune cells KW - biomarker KW - monocytes KW - regulatory T cells KW - ischemic stroke KW - thromboinflammation Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-126319 VL - 16 IS - 10 ER - TY - JOUR A1 - Langenhorst, Daniela A1 - Tabares, Paula A1 - Gulde, Tobias A1 - Becklund, Bryan R. A1 - Berr, Susanne A1 - Surh, Charles D. A1 - Beyersdorf, Niklas A1 - Hünig, Thomas T1 - Self-recognition sensitizes mouse and human regulatory T cells to low-dose CD28 superagonist stimulation JF - Frontiers in Immunology N2 - In rodents, low doses of CD28-specific superagonistic monoclonal antibodies (CD28 superagonists, CD28SA) selectively activate regulatory T cells (Treg). This observation has recently been extended to humans, suggesting an option for the treatment of autoimmune and inflammatory diseases. However, a mechanistic explanation for this phenomenon is still lacking. Given that CD28SA amplify T cell receptor (TCR) signals, we tested the hypothesis that the weak tonic TCR signals received by conventional CD4\(^{+}\) T cells (Tconv) in the absence of cognate antigen require more CD28 signaling input for full activation than the stronger TCR signals received by self-reactive Treg. We report that in vitro, the response of mouse Treg and Tconv to CD28SA strongly depends on MHC class II expression by antigen-presenting cells. To separate the effect of tonic TCR signals from self-peptide recognition, we compared the response of wild-type Treg and Tconv to low and high CD28SA doses upon transfer into wild-type or H-2M knockout mice, which lack a self-peptide repertoire. We found that the superior response of Treg to low CD28SA doses was lost in the absence of self-peptide presentation. We also tested if potentially pathogenic autoreactive Tconv would benefit from self-recognition-induced sensitivity to CD28SA stimulation by transferring TCR transgenic OVA-specific Tconv into OVA-expressing mice and found that low-dose CD28SA application inhibited, rather than supported, their expansion, presumably due to the massive concomitant activation of Treg. Finally, we report that also in the in vitro response of human peripheral blood mononuclear cells to CD28SA, HLA II blockade interferes with the expansion of Treg by low-dose CD28SA stimulation. These results provide a rational basis for the further development of low-dose CD28SA therapy for the improvement of Treg activity. KW - D665 KW - regulatory T cells KW - self-reactivity KW - autoimmunity KW - CD28 superagonists KW - TGN1412 KW - TAB08 Y1 - 2018 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-159387 VL - 8 IS - 1985 ER - TY - THES A1 - Matuschek, Anja T1 - Characterization of tolerogenic rat bone marrow-derived dendritic cells and regulatory T cells T1 - Charakterisierung tolerogener dendritischer Knochenmarkszellen und regulatorischer T-Lymphozyten aus der Ratte N2 - Tolerogene Dendritische Zellen (DZ) und regulatorische T-Lymphozyten (Treg) verfügen über die Fähigkeit, destruktive Immunantworten zu verhindern. Die Hoffnung besteht, solche Zellen in naher Zukunft für therapeutische Zwecke einzusetzen, um z. B. Immunantworten nach Transplantation, aber auch bei Autoimmunität und Allergie antigenspezifisch zu supprimieren. Zum jetzigen Zeitpunkt ist die Generierung solcher Zellen aufwendig und noch nicht für die klinische Routine geeignet. Zudem sind die Mechanismen noch wenig verstanden, wie diese Zellen eine gewünschte Immunhemmung in vivo auszulösen und wie der möglichen Gefahr einer zu starken Immunhemmung zu begegnen ist. Das Kleinnagermodell Ratte ist für die biomedizinische Forschung noch immer von großer Bedeutung, umso überraschender ist es, dass insbesondere tolerogene DZ und Treg in diesem Modell bisher nur unzureichend untersucht wurden. Das Ziel der Arbeit war deshalb, diese Immunzellen umfassend zu charakterisieren und ihre Funktion auf das Immunsystem zu untersuchen. Tolerogene DZ wurden mit GM-CSF und IL-4 aus Knochenmarkvorläuferzellen generiert (= IL-4 DC). Der Anteil an natürlich vorkommenden Treg mit einem Phänotyp CD4posCD25posFoxp3pos umfasst ca. 5-8% der peripheren naiven CD4pos TLymphozyten. Die Charakterisierung der IL-4 DC zeigte im Vergleich zu reifen DZ der Milz eine bis zu 26-fach geringere Expression von Oberflächenmolekülen wie MHC-Klasse II Molekül, CD80, CD86, ICAM-1 und CD25. Diese geringe Expression änderte sich auch nicht, wenn die Zellen verschiedensten Reifungssignalen wie das Replattieren,LPS, TNF-α und CD40L ausgesetzt wurden. IL-4 DC verfügen somit über einen robusten und gegenüber Reifungssignalen überaus resistenten Phänotyp. IL-4 DC nehmen Antigene durch Endozytose auf und sind unfähig, sowohl naive TLymphozyten zu aktivieren, als auch antigenspezifische T-Lymphozyten zu restimulieren. Zudem sind sie in der Lage, die Aktivierung naiver T-Lymphozyten und die Restimulierung antigenspezifischer T-Lymphozyten durch reife Milz-DZ zu bzw. zu verzögern. Dabei verringerte sich die Proliferation der TLymphozyten um bis zu 95%. Diese Beeinflussung der Proliferation ist nach Zugabe der IL-4 DC bereits innerhalb von 24 Stunden zu messen. Die verringerte Aktivierung geht zu dem mit einer verringerten Zytokinausschüttung (IL-2 um 49% und IFN-γ um 92%) einher. Die inhibitorischen Eigenschaften der IL-4 DC scheinen aber nicht ausschließlich auf der verringerten Expression kostimulatorischer Moleküle zu beruhen. Der Nachweis der beiden inhibitorischen Oberflächenmoleküle PD-L1 und PD-L2 auf IL-4 DC lässt ebenfalls eine Bedeutung dieser Moleküle bei der Vermittlung inhibierender Signale vermuten. Auch die suppressive Wirkung löslicher Faktoren wurde in der vorliegenden Arbeit gezeigt. Überstände einer 24-stündigen Kultur mit einer Million IL-4 DC hemmten die Aktivierung naiver T-Lymphozyten durch reife Milz-DZ um etwa 90%. Für diese Immunhemmung scheint das in diesen Überständen nachgewiesene Zytokin TGF-β (bis 300 pg/ml) verantwortlich zu sein. Im Vergleich dazu wiesen Überstände reifer Milz-DZ, die nicht die Aktivierung von T-Lymphozyten hemmten, eine TGF-β Konzentrationen von bis 100 pg/ml auf. Im Gegensatz dazu scheint zelltoxisches Stickstoffmonoxid nur eine geringe Rolle bei der Inhibierung der T-Zellproliferation zu spielen. Die Zugabe des NO Synthase-Inhibitors NMMA verringerte zwar den Anteil an NO um ca. 50%, doch führte dies nicht zu einer Steigerung der Proliferation von T-Lymphozyten. IL-4 DC sind zwar nicht in der Lage, T-Lymphozyten zur Proliferation zu bringen, doch bedeutet dies nicht, dass keinerlei Veränderungen auf molekularer Ebene festzustellen wären. So sind T-Lymphozyten nach ihrer Inkubation mit IL-4 DC nicht in der Lage, in Gegenwart von reifen Milz-DZ zu proliferieren. Dieser anergische Zustand wurde nach Zugabe von IL-2 aufgehoben. Zudem können diese TLymphozyten nach ihrer Inkubation mit IL-4 DC die Aktivierung naïver TLymphozyten hemmen. Naïve und aktivierte T-Lymphozyten können dies nicht. Diese Beobachtung, die auf eine Induktion von Treg schließen lässt, wurde genauer untersucht. In der Tat zeigten durchflusszytometrische Analysen eine 1,6-fach verstärkte Expansion von CD4posCD25posFoxp3pos T-Lymphozyten aus natürlich vorkommenden Treg in Gegenwart von IL-4 DC. Dabei erfolgte die Expansion von CD4posCD25posFoxp3pos T-Lymphozyten unabhängig vom Reifegrad der DZ. So waren auch reife Milz-DZ dazu in der Lage, die Zahl der natürlich vorkommenden Treg zu erhöhen. Doch wiesen diese mit Milz-DZ inkubierten Treg einen verminderten inhibitorischen Effekt auf. Im Gegensatz dazu waren die mit IL-4 DC inkubierten Treg in der Lage die Aktivierung naiver T-Lymphozyten zu hemmen. In dieser Arbeit wurde gezeigt, dass sich das regulatorische Potential von DZ nicht ausschließlich vom Phänotyp bzw. ihrem Reifegrad ableiten lässt, sondern dass hierzu auch ihre funktionellen Eigenschaften zu untersuchen sind. Die Induktion von Treg mit suppressiven Eigenschaften durch in vitro generierte tolerogene IL-4 DC könnte ein wichtiger Mechanismus zur Aufrechterhaltung der peripheren Toleranz darstellen. Vor einer klinischen Umsetzung sind aber noch weitergehende Untersuchungen notwendig, um das Zusammenspiel zwischen tolerogenen DZ und Treg zu verstehen, aber auch um die Auswirkungen eines Transfers großer Mengen regulatorischer Zellen auf das Immunsystem des Empfängers zu untersuchen. N2 - Tolerogenic dendritic cells (DC) and regulatory T (Treg) cells are able to prevent destructive immune responses. There is reason to hope that it may soon be possible to use DC and Treg cells to suppress immune responses antigen-specific, not only after transplantation, but also in the case of autoimmunity and allergy. At the moment, the generation of such cell types is very time-consuming and not suitable for clinical routine. In addition, it is not yet fully understood how these cells elicit a desired protective immune response in vivo and how the risks of an excessive immune suppression can be managed. The rat is one of the most important animal models in biomedical research. It is therefore surprising that tolerogenic DC and Treg cells in particular have not been more thoroughly investigated in this model. Thus, the aim of the present study was to systematically characterize these immune cells and investigate their impact on the immune system. Tolerogenic DC were generated from bone marrow precursors cultured with GM-CSF and IL-4 (= IL-4 DC). The proportion of naturally occurring Treg cells with a CD4posCD25posFoxp3pos phenotype comprises approximately 5-8% of the peripheral CD4pos T cells. The characterization of IL-4 DC revealed an up to 26-fold reduced expression of surface molecules such as MHC class II molecules, CD80, CD86, ICAM-1 and CD25 in comparison to mature splenic DC (S-DC). This low expression did not change when the cells where stimulated with different maturation-inducing signals such as replating, LPS, TNF- α and CD40L. Thus, these cells possess a robust phenotype resistant to maturation-inducing stimuli. IL-4 DC take up antigen via endocytosis and are not able to activate naïve T cells or to restimulate antigen-specific T cells. Furthermore, they are able to inhibit and prolongate mature S-DC induced T cell proliferation as well as mature S-DC induced restimulation of antigen-specific T cells, respectively. Thereby, the T cell proliferation was reduced up to 95%. This strong inhibitory effect was mediated within 24 hours in association with a reduced cytokine production (IL-2 about 49% and IFN-γ about 92%). The inhibitory properties of IL-4 DC don´t seem to be caused exclusively by the reduced expression of co-stimulatory molecules. In this study, the detection of the inhibitory molecules PD-L1 and PD-L2 on IL-4 DC suggests they have an impact on mediating inhibitory signals to the T cells. In addition, a suppressive effect of soluble factors was shown. The supernatant of one million IL-4 DC, collected after a 24 hour culture, suppressed mature S-DC induced proliferation of naïve T cells by about 90%. TGF-β, which was detected in the supernatant (up to 300 pg/ml), appears to be the causing soluble factor for this immune inhibition. By contrast, the supernatants of mature S-DC, which did not inhibit the activation of T cells, showed a TGF-β concentration of only about 100 pg/ml. The cytotoxic nitric oxide does not contribute to the IL-4 DC-mediated inhibition of T cell proliferation. The NO synthase inhibitor NMMA reduced the amount of NO by about 50%, but the decreased NO levels did not influence T cell proliferation. Indeed, IL-4 DC are not able to induce T cell proliferation, but this doesn´t mean that there is no change on the molecular level. For instance, T cells co-cultured with IL-4 DC during a first culture are not able to proliferate in the presence of mature S-DC during a second culture. This anergic-like state, however, could be abolished by adding exogenous IL-2. In addition, T cells co-cultured with IL-4 DC are able to inhibit the activation of naïve T cells. Naïve and activated T cells were not able to inhibit the mature S-DC induced T cell proliferation. This observation suggests the induction of Treg cells and was investigated in more detail. Indeed, flow cytometric analysis showed a 1.6-fold expansion of CD4posCD25posFoxp3pos T cells from naturally occurring Treg cells in the presence of IL-4 DC. Thereby, the expansion of CD4posCD25posFoxp3pos T cells occurs independently of the maturation state of DC. Both immature IL-4 DC as well as mature S-DC were able to expand the percentage of naturally occurring Treg cells. However, Treg cells pre-incubated with mature S-DC demonstrated a diminished inhibitory effect compared to Treg cells pre-incubated with IL-4 DC. Treg cells pre-incubated with IL-4 DC were able to inhibit the activation of naïve T cells. In this study it was shown that the regulatory potential of DC cannot be deduced solely by their phenotype or maturation state. Other factors, such as functional properties, need to taken into consideration, too. The induction of Treg cells with suppressive properties induced by in vitro generated tolerogenic IL-4 DC might provide an important mechanism for the maintenance of peripheral tolerance. However, for clinical application further investigation is necessary, not only to understand the interactions between tolerogenic DC and Treg cells, but also to investigate the impact of the transfer of a larger quantity of regulatory cells on the immune system of the recipient. KW - Dendritische Zelle KW - T-Lymphozyt KW - Immuntoleranz KW - Allogene Zelle KW - Transforming Growth Factor beta KW - tolerogen KW - Dendritische Zelle KW - regulatorische T-Zellen KW - allogen KW - TGF-ß KW - tolerogenic KW - dendritic cell KW - regulatory T cells KW - allogenic KW - TGF-ß Y1 - 2010 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-51708 ER - TY - JOUR A1 - Nono, Justin Komguep A1 - Pletinckx, Katrien A1 - Lutz, Manfred B. A1 - Brehm, Klaus T1 - Excretory/Secretory-Products of Echinococcus multilocularis Larvae Induce Apoptosis and Tolerogenic Properties in Dendritic Cells In Vitro JF - PLoS Neglected Tropical Diseases N2 - Background: Alveolar echinococcosis, caused by Echinococcus multilocularis larvae, is a chronic disease associated with considerable modulation of the host immune response. Dendritic cells (DC) are key effectors in shaping the immune response and among the first cells encountered by the parasite during an infection. Although it is assumed that E. multilocularis, by excretory/secretory (E/S)-products, specifically affects DC to deviate immune responses, little information is available on the molecular nature of respective E/S-products and their mode of action. Methodology/Principal Findings: We established cultivation systems for exposing DC to live material from early (oncosphere), chronic (metacestode) and late (protoscolex) infectious stages. When co-incubated with Echinococcus primary cells, representing the invading oncosphere, or metacestode vesicles, a significant proportion of DC underwent apoptosis and the surviving DC failed to mature. In contrast, DC exposed to protoscoleces upregulated maturation markers and did not undergo apoptosis. After pre-incubation with primary cells and metacestode vesicles, DC showed a strongly impaired ability to be activated by the TLR ligand LPS, which was not observed in DC pre-treated with protoscolex E/S-products. While none of the larvae induced the secretion of pro-inflammatory IL-12p70, the production of immunosuppressive IL-10 was elevated in response to primary cell E/S-products. Finally, upon incubation with DC and naive T-cells, E/S-products from metacestode vesicles led to a significant expansion of Foxp3+ T cells in vitro. Conclusions: This is the first report on the induction of apoptosis in DC by cestode E/S-products. Our data indicate that the early infective stage of E. multilocularis is a strong inducer of tolerance in DC, which is most probably important for generating an immunosuppressive environment at an infection phase in which the parasite is highly vulnerable to host attacks. The induction of CD4+CD25+Foxp3+ T cells through metacestode E/S-products suggests that these cells fulfill an important role for parasite persistence during chronic echinococcosis. KW - granulosus KW - hydatid disease KW - metacestode vesicles KW - antigen-B KW - alveoar echinococcosis KW - TGF-BETA KW - regulatory T cells KW - gene expression KW - Brugia Malayi KW - TNF-alpha Y1 - 2012 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-134280 VL - 6 IS - 2 ER -