Publications by authors named "Jochen Huehn"

116 Publications

Generation of Sequencing Libraries for Building Immune Cell Methylomes.

Methods Mol Biol 2021 ;2285:265-276

Department of Experimental Immunology, Helmholtz Centre for Infection Research, Braunschweig, Germany.

The comparison of methylomes from immune cells enables the identification of differentially methylated regions and thereby region-associated gene loci. Those regions can be used to discriminate one immune cell population from the other, as well as help to identify key molecules and major pathways determining the unique phenotypes of immune cell lineages. The combination of bisulfite treatment of genomic DNA and next-generation sequencing provides the basis for studying epigenetic changes in different immune cell populations. Further development of whole-genome bisulfite sequencing resulted in a protocol for sequencing libraries that accept both single- or double-stranded DNA from fixed or nonfixed cells, respectively. Therefore, researchers can include immune cell populations in their methylation studies whose isolation depends on the staining of intracellular molecules.
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http://dx.doi.org/10.1007/978-1-0716-1311-5_21DOI Listing
January 2021

Single-cell chromatin accessibility landscape identifies tissue repair program in human regulatory T cells.

Immunity 2021 Apr 30;54(4):702-720.e17. Epub 2021 Mar 30.

Regensburg Center for Interventional Immunology (RCI); Chair for Immunology, University Regensburg, 93053 Regensburg, Germany. Electronic address:

Murine regulatory T (Treg) cells in tissues promote tissue homeostasis and regeneration. We sought to identify features that characterize human Treg cells with these functions in healthy tissues. Single-cell chromatin accessibility profiles of murine and human tissue Treg cells defined a conserved, microbiota-independent tissue-repair Treg signature with a prevailing footprint of the transcription factor BATF. This signature, combined with gene expression profiling and TCR fate mapping, identified a population of tissue-like Treg cells in human peripheral blood that expressed BATF, chemokine receptor CCR8 and HLA-DR. Human BATFCCR8 Treg cells from normal skin and adipose tissue shared features with nonlymphoid T follicular helper-like (Tfh-like) cells, and induction of a Tfh-like differentiation program in naive human Treg cells partially recapitulated tissue Treg regenerative characteristics, including wound healing potential. Human BATFCCR8 Treg cells from healthy tissue share features with tumor-resident Treg cells, highlighting the importance of understanding the context-specific functions of these cells.
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http://dx.doi.org/10.1016/j.immuni.2021.03.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8050210PMC
April 2021

The microbiota is dispensable for the early stages of peripheral regulatory T cell induction within mesenteric lymph nodes.

Cell Mol Immunol 2021 May 24;18(5):1211-1221. Epub 2021 Mar 24.

Department Experimental Immunology, Helmholtz Centre for Infection Research, Braunschweig, Germany.

Intestinal Foxp3 regulatory T cell (Treg) subsets are crucial players in tolerance to microbiota-derived and food-borne antigens, and compelling evidence suggests that the intestinal microbiota modulates their generation, functional specialization, and maintenance. Selected bacterial species and microbiota-derived metabolites, such as short-chain fatty acids (SCFAs), have been reported to promote Treg homeostasis in the intestinal lamina propria. Furthermore, gut-draining mesenteric lymph nodes (mLNs) are particularly efficient sites for the generation of peripherally induced Tregs (pTregs). Despite this knowledge, the direct role of the microbiota and their metabolites in the early stages of pTreg induction within mLNs is not fully elucidated. Here, using an adoptive transfer-based pTreg induction system, we demonstrate that neither transfer of a dysbiotic microbiota nor dietary SCFA supplementation modulated the pTreg induction capacity of mLNs. Even mice housed under germ-free (GF) conditions displayed equivalent pTreg induction within mLNs. Further molecular characterization of these de novo induced pTregs from mLNs by dissection of their transcriptomes and accessible chromatin regions revealed that the microbiota indeed has a limited impact and does not contribute to the initialization of the Treg-specific epigenetic landscape. Overall, our data suggest that the microbiota is dispensable for the early stages of pTreg induction within mLNs.
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http://dx.doi.org/10.1038/s41423-021-00647-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8093251PMC
May 2021

Lymph node stromal cell subsets-Emerging specialists for tailored tissue-specific immune responses.

Int J Med Microbiol 2021 Apr 25;311(3):151492. Epub 2021 Feb 25.

Department Experimental Immunology, Helmholtz Centre for Infection Research, Inhoffenstr. 7, 38124 Braunschweig, Germany; Cluster of Excellence RESIST (EXC 2155), Hannover Medical School, Carl-Neuberg-Str. 1, 30625 Hannover, Germany. Electronic address:

The effective priming of adaptive immune responses depends on the precise dispatching of lymphocytes and antigens into and within lymph nodes (LNs), which are strategically dispersed throughout the body. Over the past decade, a growing body of evidence has advanced our understanding of lymph node stromal cells (LNSCs) from viewing them as mere accessory cells to seeing them as critical cellular players for the modulation of adaptive immune responses. In this review, we summarize current advances on the pivotal roles that LNSCs play in orchestrating adaptive immune responses during homeostasis and infection, and highlight the imprinting of location-specific information by micro-environmental cues into LNSCs, thereby tailoring tissue-specific immune responses.
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http://dx.doi.org/10.1016/j.ijmm.2021.151492DOI Listing
April 2021

Influenza A virus-induced thymus atrophy differentially affects dynamics of conventional and regulatory T-cell development in mice.

Eur J Immunol 2021 May 17;51(5):1166-1181. Epub 2021 Mar 17.

Department of Experimental Immunology, Helmholtz Centre for Infection Research, Braunschweig, Germany.

Foxp3 Treg cells, which are crucial for maintenance of self-tolerance, mainly develop within the thymus, where they arise from CD25 Foxp3 or CD25 Foxp3 Treg cell precursors. Although it is known that infections can cause transient thymic involution, the impact of infection-induced thymus atrophy on thymic Treg (tTreg) cell development is unknown. Here, we infected mice with influenza A virus (IAV) and studied thymocyte population dynamics post infection. IAV infection caused a massive, but transient thymic involution, dominated by a loss of CD4 CD8 double-positive (DP) thymocytes, which was accompanied by a significant increase in the frequency of CD25 Foxp3 tTreg cells. Differential apoptosis susceptibility could be experimentally excluded as a reason for the relative tTreg cell increase, and mathematical modeling suggested that enhanced tTreg cell generation cannot explain the increased frequency of tTreg cells. Yet, an increased death of DP thymocytes and augmented exit of single-positive (SP) thymocytes was suggested to be causative. Interestingly, IAV-induced thymus atrophy resulted in a significantly reduced T-cell receptor (TCR) repertoire diversity of newly produced tTreg cells. Taken together, IAV-induced thymus atrophy is substantially altering the dynamics of major thymocyte populations, finally resulting in a relative increase of tTreg cells with an altered TCR repertoire.
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http://dx.doi.org/10.1002/eji.202048981DOI Listing
May 2021

Efficient IL-2R signaling differentially affects the stability, function, and composition of the regulatory T-cell pool.

Cell Mol Immunol 2021 Feb 6;18(2):398-414. Epub 2021 Jan 6.

Institute of Immunology, Hannover Medical School, Hannover, Germany.

Signaling via interleukin-2 receptor (IL-2R) is a requisite for regulatory T (Treg) cell identity and function. However, it is not completely understood to what degree IL-2R signaling is required for Treg cell homeostasis, lineage stability and function in both resting and inflammatory conditions. Here, we characterized a spontaneous mutant mouse strain endowed with a hypomorphic Tyr129His variant of CD25, the α-chain of IL-2R, which resulted in diminished receptor expression and reduced IL-2R signaling. Under noninflammatory conditions, Cd25 mice harbored substantially lower numbers of peripheral Treg cells with stable Foxp3 expression that prevented the development of spontaneous autoimmune disease. In contrast, Cd25 Treg cells failed to efficiently induce immune suppression and lost lineage commitment in a T-cell transfer colitis model, indicating that unimpaired IL-2R signaling is critical for Treg cell function in inflammatory environments. Moreover, single-cell RNA sequencing of Treg cells revealed that impaired IL-2R signaling profoundly affected the balance of central and effector Treg cell subsets. Thus, partial loss of IL-2R signaling differentially interferes with the maintenance, heterogeneity, and suppressive function of the Treg cell pool.
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http://dx.doi.org/10.1038/s41423-020-00599-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8027001PMC
February 2021

Alpha-Toxin Limits Type 1 While Fostering Type 3 Immune Responses.

Front Immunol 2020 7;11:1579. Epub 2020 Aug 7.

Department Experimental Immunology, Helmholtz Centre for Infection Research, Braunschweig, Germany.

can cause life-threatening diseases, and hospital- as well as community-associated antibiotic-resistant strains are an emerging global public health problem. Therefore, prophylactic vaccines or immune-based therapies are considered as alternative treatment opportunities. To develop such novel treatment approaches, a better understanding of the bacterial virulence and immune evasion mechanisms and their potential effects on immune-based therapies is essential. One important staphylococcal virulence factor is alpha-toxin, which is able to disrupt the epithelial barrier in order to establish infection. In addition, alpha-toxin has been reported to modulate other cell types including immune cells. Since CD4 T cell-mediated immunity is required for protection against infection, we were interested in the ability of alpha-toxin to directly modulate CD4 T cells. To address this, murine naïve CD4 T cells were differentiated into effector T cell subsets in the presence of alpha-toxin. Interestingly, alpha-toxin induced death of Th1-polarized cells, while cells polarized under Th17 conditions showed a high resistance toward increasing concentrations of this toxin. These effects could neither be explained by differential expression of the cellular alpha-toxin receptor ADAM10 nor by differential activation of caspases, but might result from an increased susceptibility of Th1 cells toward Ca-mediated activation-induced cell death. In accordance with the findings, an alpha-toxin-dependent decrease of Th1 and concomitant increase of Th17 cells was observed during bacteremia. Interestingly, corresponding subsets of innate lymphoid cells and γδ T cells were similarly affected, suggesting a more general effect of alpha-toxin on the modulation of type 1 and type 3 immune responses. In conclusion, we have identified a novel alpha-toxin-dependent immunomodulatory strategy of , which can directly act on CD4 T cells and might be exploited for the development of novel immune-based therapeutic approaches to treat infections with antibiotic-resistant strains.
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http://dx.doi.org/10.3389/fimmu.2020.01579DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7427519PMC
April 2021

Acute neonatal Listeria monocytogenes infection causes long-term, organ-specific changes in immune cell subset composition.

Eur J Microbiol Immunol (Bp) 2020 Jun 19. Epub 2020 Jun 19.

1Department Experimental Immunology, Helmholtz Centre for Infection Research, Braunschweig, Germany.

Listeria monocytogenes (Lm) is a food-borne pathogen with a high chance of infecting neonates, pregnant women, elderly and immunocompromised individuals. Lm infection in neonates can cause neonatal meningitis and sepsis with a high risk of severe neurological and developmental sequelae and high mortality rates. However, whether an acute neonatal Lm infection causes long-term effects on the immune system persisting until adulthood has not been fully elucidated. Here, we established a neonatal Lm infection model and monitored the composition of major immune cell subsets at defined time points post infection (p.i.) in secondary lymphoid organs and the intestine. Twelve weeks p.i., the CD8+ T cell population was decreased in colon and mesenteric lymph nodes (mLNs) with an opposing increase in the spleen. In the colon, we observed an accumulation of CD4+ and CD8+ effector/memory T cells with an increase of T-bet+ T helper 1 (Th1) cells. In addition, 12 weeks p.i. an altered composition of innate lymphoid cell (ILC) and dendritic cell (DC) subsets was still observed in colon and mLNs, respectively. Together, these findings highlight organ-specific long-term consequences of an acute neonatal Lm infection on both the adaptive and innate immune system.
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http://dx.doi.org/10.1556/1886.2020.00007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7391377PMC
June 2020

Salt generates antiinflammatory Th17 cells but amplifies pathogenicity in proinflammatory cytokine microenvironments.

J Clin Invest 2020 09;130(9):4587-4600

Institute of Virology, Technical University of Munich, Munich, Germany.

Th cells integrate signals from their microenvironment to acquire distinct specialization programs for efficient clearance of diverse pathogens or for immunotolerance. Ionic signals have recently been demonstrated to affect T cell polarization and function. Sodium chloride (NaCl) was proposed to accumulate in peripheral tissues upon dietary intake and to promote autoimmunity via the Th17 cell axis. Here, we demonstrate that high-NaCl conditions induced a stable, pathogen-specific, antiinflammatory Th17 cell fate in human T cells in vitro. The p38/MAPK pathway, involving NFAT5 and SGK1, regulated FoxP3 and IL-17A expression in high-NaCl conditions. The NaCl-induced acquisition of an antiinflammatory Th17 cell fate was confirmed in vivo in an experimental autoimmune encephalomyelitis (EAE) mouse model, which demonstrated strongly reduced disease symptoms upon transfer of T cells polarized in high-NaCl conditions. However, NaCl was coopted to promote murine and human Th17 cell pathogenicity, if T cell stimulation occurred in a proinflammatory and TGF-β-low cytokine microenvironment. Taken together, our findings reveal a context-dependent, dichotomous role for NaCl in shaping Th17 cell pathogenicity. NaCl might therefore prove beneficial for the treatment of chronic inflammatory diseases in combination with cytokine-blocking drugs.
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http://dx.doi.org/10.1172/JCI137786DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7456214PMC
September 2020

Vitamin C supports conversion of human γδ T cells into FOXP3-expressing regulatory cells by epigenetic regulation.

Sci Rep 2020 04 16;10(1):6550. Epub 2020 Apr 16.

Institute of Immunology, Christian-Albrechts-University Kiel, D-24105, Kiel, Germany.

Human γδ T cells are potent cytotoxic effector cells, produce a variety of cytokines, and can acquire regulatory activity. Induction of FOXP3, the key transcription factor of regulatory T cells (Treg), by TGF-β in human Vγ9 Vδ2 T cells has been previously reported. Vitamin C is an antioxidant and acts as multiplier of DNA hydroxymethylation. Here we have investigated the effect of the more stable phospho-modified Vitamin C (pVC) on TGF-β-induced FOXP3 expression and the resulting regulatory activity of highly purified human Vγ9 Vδ2 T cells. pVC significantly increased the TGF-β-induced FOXP3 expression and stability and also increased the suppressive activity of Vγ9 Vδ2 T cells. Importantly, pVC induced hypomethylation of the Treg-specific demethylated region (TSDR) in the FOXP3 gene. Genome-wide methylation analysis by Reduced Representation Bisulfite Sequencing additionally revealed differentially methylated regions in several important genes upon pVC treatment of γδ T cells. While Vitamin C also enhances effector functions of Vγ9 Vδ2 T cells in the absence of TGF-β, our results demonstrate that pVC potently increases the suppressive activity and FOXP3 expression in TGF-β-treated Vγ9 Vδ2 T cells by epigenetic modification of the FOXP3 gene.
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http://dx.doi.org/10.1038/s41598-020-63572-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7162875PMC
April 2020

Recirculating IL-1R2 Tregs fine-tune intrathymic Treg development under inflammatory conditions.

Cell Mol Immunol 2021 Jan 27;18(1):182-193. Epub 2020 Jan 27.

Department of Experimental Immunology, Helmholtz Centre for Infection Research, Braunschweig, Germany.

The vast majority of Foxp3 regulatory T cells (Tregs) are generated in the thymus, and several factors, such as cytokines and unique thymic antigen-presenting cells, are known to contribute to the development of these thymus-derived Tregs (tTregs). Here, we report the existence of a specific subset of Foxp3 Tregs within the thymus that is characterized by the expression of IL-1R2, which is a decoy receptor for the inflammatory cytokine IL-1. Detailed flow cytometric analysis of the thymocytes from Foxp3xRAG1 reporter mice revealed that the IL-1R2 Tregs are mainly RAG1 and CCR6CCR7, demonstrating that these Tregs are recirculating cells entering the thymus from the periphery and that they have an activated phenotype. In the spleen, the majority of IL-1R2 Tregs express neuropilin-1 (Nrp-1) and Helios, suggesting a thymic origin for these Tregs. Interestingly, among all tissues studied, the highest frequency of IL-1R2 Tregs was observed in the thymus, indicating preferential recruitment of this Treg subset by the thymus. Using fetal thymic organ cultures (FTOCs), we demonstrated that increased concentrations of exogenous IL-1β blocked intrathymic Treg development, resulting in a decreased frequency of CD25Foxp3 tTregs and an accumulation of CD25Foxp3 Treg precursors. Interestingly, the addition of IL-1R2 Tregs, but not IL-1R2 Tregs, to reaggregated thymic organ cultures (RTOCs) abrogated the IL-1β-mediated blockade, demonstrating that these recirculating IL-1R2 Tregs can quench IL-1 signaling in the thymus and thereby maintain thymic Treg development even under inflammatory conditions.
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http://dx.doi.org/10.1038/s41423-019-0352-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7853075PMC
January 2021

The Transcription Factor MAZR/PATZ1 Regulates the Development of FOXP3 Regulatory T Cells.

Cell Rep 2019 12;29(13):4447-4459.e6

Division of Immunobiology, Institute of Immunology, Center for Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Vienna, Austria. Electronic address:

Forkhead box protein P3 (FOXP3) regulatory T cells (T cells) play a key role in maintaining tolerance and immune homeostasis. Here, we report that a T cell-specific deletion of the transcription factor MAZR (also known as PATZ1) leads to an increased frequency of T cells, while enforced MAZR expression impairs T cell differentiation. Further, MAZR expression levels are progressively downregulated during thymic T cell development and during in-vitro-induced human T cell differentiation, suggesting that MAZR protein levels are critical for controlling T cell development. However, MAZR-deficient T cells show only minor transcriptional changes ex vivo, indicating that MAZR is not essential for establishing the transcriptional program of peripheral T cells. Finally, the loss of MAZR reduces the clinical score in dextran-sodium sulfate (DSS)-induced colitis, suggesting that MAZR activity in T cells controls the extent of intestinal inflammation. Together, these data indicate that MAZR is part of a T cell-intrinsic transcriptional network that modulates T cell development.
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http://dx.doi.org/10.1016/j.celrep.2019.11.089DOI Listing
December 2019

Dynamic Imprinting of the Treg Cell-Specific Epigenetic Signature in Developing Thymic Regulatory T Cells.

Front Immunol 2019 11;10:2382. Epub 2019 Oct 11.

Department Experimental Immunology, Helmholtz Centre for Infection Research, Braunschweig, Germany.

Regulatory T (Treg) cells mainly develop within the thymus and arise from CD25Foxp3 (CD25 TregP) or CD25Foxp3 (Foxp3 TregP) Treg cell precursors resulting in Treg cells harboring distinct transcriptomic profiles and complementary T cell receptor repertoires. The stable and long-term expression of Foxp3 in Treg cells and their stable suppressive phenotype are controlled by the demethylation of Treg cell-specific epigenetic signature genes including an evolutionarily conserved CpG-rich element within the locus, the Treg-specific demethylated region (TSDR). Here we analyzed the dynamics of the imprinting of the Treg cell-specific epigenetic signature genes in thymic Treg cells. We could demonstrate that CD25Foxp3 Treg cells show a progressive demethylation of most signature genes during maturation within the thymus. Interestingly, a partial demethylation of several Treg cell-specific epigenetic signature genes was already observed in Foxp3 TregP but not in CD25 TregP. Furthermore, Foxp3 TregP were very transient in nature and arose at a more mature developmental stage when compared to CD25 TregP. When the two Treg cell precursors were cultured in presence of IL-2, a factor known to be critical for thymic Treg cell development, we observed a major impact of IL-2 on the demethylation of the TSDR with a more pronounced effect on Foxp3 TregP. Together, these results suggest that the establishment of the Treg cell-specific hypomethylation pattern is a continuous process throughout thymic Treg cell development and that the two known Treg cell precursors display distinct dynamics for the imprinting of the Treg cell-specific epigenetic signature genes.
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http://dx.doi.org/10.3389/fimmu.2019.02382DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6797672PMC
October 2020

Guidelines for the use of flow cytometry and cell sorting in immunological studies (second edition).

Eur J Immunol 2019 Oct;49(10):1457-1973

Flow Cytometry Laboratory, Institute of Molecular Toxicology and Pharmacology, Helmholtz Zentrum München, German Research Center for Environmental Health, München, Germany.

These guidelines are a consensus work of a considerable number of members of the immunology and flow cytometry community. They provide the theory and key practical aspects of flow cytometry enabling immunologists to avoid the common errors that often undermine immunological data. Notably, there are comprehensive sections of all major immune cell types with helpful Tables detailing phenotypes in murine and human cells. The latest flow cytometry techniques and applications are also described, featuring examples of the data that can be generated and, importantly, how the data can be analysed. Furthermore, there are sections detailing tips, tricks and pitfalls to avoid, all written and peer-reviewed by leading experts in the field, making this an essential research companion.
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http://dx.doi.org/10.1002/eji.201970107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7350392PMC
October 2019

Chimeric antigen receptor-induced BCL11B suppression propagates NK-like cell development.

J Clin Invest 2019 12;129(12):5108-5122

Department of Pediatric Hematology/Oncology and Blood Stem Cell Transplantation, Hannover Medical School, Hannover, Germany.

The transcription factor B cell CLL/lymphoma 11B (BCL11B) is indispensable for T lineage development of lymphoid progenitors. Here, we show that chimeric antigen receptor (CAR) expression during early phases of ex vivo generation of lymphoid progenitors suppressed BCL11B, leading to suppression of T cell-associated gene expression and acquisition of NK cell-like properties. Upon adoptive transfer into hematopoietic stem cell transplant recipients, CAR-expressing lymphoid progenitors differentiated into CAR-induced killer (CARiK) cells that mediated potent antigen-directed antileukemic activity even across MHC barriers. CD28 and active immunoreceptor tyrosine-based activation motifs were critical for a functional CARiK phenotype. These results give important insights into differentiation of murine and human lymphoid progenitors driven by synthetic CAR transgene expression and encourage further evaluation of ex vivo-generated CARiK cells for targeted immunotherapy.
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http://dx.doi.org/10.1172/JCI126350DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6877334PMC
December 2019

Generation of Foxp3CD25 Regulatory T-Cell Precursors Requires c-Rel and IκB.

Front Immunol 2019 10;10:1583. Epub 2019 Jul 10.

Systems-Oriented Immunology and Inflammation Research Group, Department of Experimental Immunology, Helmholtz Centre for Infection Research, Braunschweig, Germany.

Next to the classical developmental route, in which first CD25 and subsequently Foxp3 are induced to generate thymic regulatory T (Treg) cells, an alternative route has been described. This alternative route is characterized by reciprocal induction of Foxp3 and CD25, with CD25 induction being required to rescue developing Treg cells from Foxp3-induced apoptosis. NF-κB has been demonstrated to be crucial for the development of thymic Treg cells via the classical route. However, its impact on the alternative route is poorly characterized. Using single and double deficient mice for key regulators of the classical route, c-Rel and IκB, we here demonstrate that NF-κB is essential for the generation of alternative CD25Foxp3 precursors, as well. Thus, c-Rel and IκB govern both routes of thymic Treg cell development.
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http://dx.doi.org/10.3389/fimmu.2019.01583DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6635800PMC
June 2020

Microbiome Dependent Regulation of T and Th17 Cells in Mucosa.

Front Immunol 2019 8;10:426. Epub 2019 Mar 8.

Experimental Immunology, Helmholtz Centre for Infection Research, Hamburg, Germany.

Mammals co-exist with resident microbial ecosystem that is composed of an incredible number and diversity of bacteria, viruses and fungi. Owing to direct contact between resident microbes and mucosal surfaces, both parties are in continuous and complex interactions resulting in important functional consequences. These interactions govern immune homeostasis, host response to infection, vaccination and cancer, as well as predisposition to metabolic, inflammatory and neurological disorders. Here, we discuss recent studies on direct and indirect effects of resident microbiota on regulatory T cells (T) and Th17 cells at the cellular and molecular level. We review mechanisms by which commensal microbes influence mucosa in the context of bioactive molecules derived from resident bacteria, immune senescence, chronic inflammation and cancer. Lastly, we discuss potential therapeutic applications of microbiota alterations and microbial derivatives, for improving resilience of mucosal immunity and combating immunopathology.
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http://dx.doi.org/10.3389/fimmu.2019.00426DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6419713PMC
September 2020

miR-181a/b-1 controls thymic selection of Treg cells and tunes their suppressive capacity.

PLoS Biol 2019 03 11;17(3):e2006716. Epub 2019 Mar 11.

Institute of Immunology, Hannover Medical School, Hannover, Germany.

The interdependence of selective cues during development of regulatory T cells (Treg cells) in the thymus and their suppressive function remains incompletely understood. Here, we analyzed this interdependence by taking advantage of highly dynamic changes in expression of microRNA 181 family members miR-181a-1 and miR-181b-1 (miR-181a/b-1) during late T-cell development with very high levels of expression during thymocyte selection, followed by massive down-regulation in the periphery. Loss of miR-181a/b-1 resulted in inefficient de novo generation of Treg cells in the thymus but simultaneously permitted homeostatic expansion in the periphery in the absence of competition. Modulation of T-cell receptor (TCR) signal strength in vivo indicated that miR-181a/b-1 controlled Treg-cell formation via establishing adequate signaling thresholds. Unexpectedly, miR-181a/b-1-deficient Treg cells displayed elevated suppressive capacity in vivo, in line with elevated levels of cytotoxic T-lymphocyte-associated 4 (CTLA-4) protein, but not mRNA, in thymic and peripheral Treg cells. Therefore, we propose that intrathymic miR-181a/b-1 controls development of Treg cells and imposes a developmental legacy on their peripheral function.
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http://dx.doi.org/10.1371/journal.pbio.2006716DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6428341PMC
March 2019

Blimp1 Prevents Methylation of Foxp3 and Loss of Regulatory T Cell Identity at Sites of Inflammation.

Cell Rep 2019 02;26(7):1854-1868.e5

Klinikum Rechts der Isar, Department of Experimental Neuroimmunology, Technical University of Munich, Ismaninger Str. 22, 81675 Munich, Germany; Munich Cluster for Systems Neurology (SyNergy), Feodor-Lynen-Str. 17, 81377 Munich, Germany. Electronic address:

Foxp3 regulatory T (Treg) cells restrict immune pathology in inflamed tissues; however, an inflammatory environment presents a threat to Treg cell identity and function. Here, we establish a transcriptional signature of central nervous system (CNS) Treg cells that accumulate during experimental autoimmune encephalitis (EAE) and identify a pathway that maintains Treg cell function and identity during severe inflammation. This pathway is dependent on the transcriptional regulator Blimp1, which prevents downregulation of Foxp3 expression and "toxic" gain-of-function of Treg cells in the inflamed CNS. Blimp1 negatively regulates IL-6- and STAT3-dependent Dnmt3a expression and function restraining methylation of Treg cell-specific conserved non-coding sequence 2 (CNS2) in the Foxp3 locus. Consequently, CNS2 is heavily methylated when Blimp1 is ablated, leading to a loss of Foxp3 expression and severe disease. These findings identify a Blimp1-dependent pathway that preserves Treg cell stability in inflamed non-lymphoid tissues.
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http://dx.doi.org/10.1016/j.celrep.2019.01.070DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6389594PMC
February 2019

Modulates Regulatory T Cell Stability via Injection of Yersinia Outer Proteins in a Type III Secretion System-Dependent Manner.

Eur J Microbiol Immunol (Bp) 2018 Dec 28;8(4):101-106. Epub 2018 Nov 28.

Experimental Immunology, Helmholtz Centre for Infection Research, Braunschweig, Germany.

Adaptive immunity is essentially required to control acute infection with enteropathogenic (Yptb). We have recently demonstrated that Yptb can directly modulate naïve CD4 T cell differentiation. However, whether fully differentiated forkhead box protein P3 (Foxp3) regulatory T cells (Tregs), fundamental key players to maintain immune homeostasis, are targeted by Yptb remains elusive. Here, we demonstrate that within the CD4 T cell compartment Yptb preferentially targets Tregs and injects Yersinia outer proteins (Yops) in a process that depends on the type III secretion system and invasins. Remarkably, Yop-translocation into ex vivo isolated Foxp3 Tregs resulted in a substantial downregulation of Foxp3 expression and a decreased capacity to express the immunosuppressive cytokine interleukin-10 (IL-10). Together, these findings highlight that invasins are critically required to mediate Yptb attachment to Foxp3 Tregs, which allows efficient Yop-translocation and finally enables the modulation of the Foxp3 Tregs' suppressive phenotype.
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http://dx.doi.org/10.1556/1886.2018.00015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6348704PMC
December 2018

IFN-γ Producing Th1 Cells Induce Different Transcriptional Profiles in Microglia and Astrocytes.

Front Cell Neurosci 2018 10;12:352. Epub 2018 Oct 10.

Department of Neurology, Clinical Neuroimmunology and Neurochemistry, Hannover Medical School, Hannover, Germany.

Autoreactive T cells that infiltrate into the central nervous system (CNS) are believed to have a significant role in mediating the pathology of neuroinflammatory diseases like multiple sclerosis. Their interaction with microglia and astrocytes in the CNS is crucial for the regulation of neuroinflammatory processes. Our previous work demonstrated that effectors secreted by Th1 and Th17 cells have different capacities to influence the phenotype and function of glial cells. We have shown that Th1-derived effectors altered the phenotype and function of both microglia and astrocytes whereas Th17-derived effectors induced direct effects only on astrocytes but not on microglia. Here we investigated if effector molecules associated with IFN-γ producing Th1 cells induced different gene expression profiles in microglia and astrocytes. We performed a microarray analysis of RNA isolated from microglia and astrocytes treated with medium and Th-derived culture supernatants and compared the gene expression data. By using the criteria of 2-fold change and a false discovery rate of 0.01 (corrected < 0.01), we demonstrated that a total of 2,106 and 1,594 genes were differentially regulated in microglia and astrocytes, respectively, in response to Th1-derived factors. We observed that Th1-derived effectors induce distinct transcriptional changes in microglia and astrocytes in addition to commonly regulated transcripts. These distinct transcriptional changes regulate peculiar physiological functions, and this knowledge can help to better understand T cell mediated neuropathologies.
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http://dx.doi.org/10.3389/fncel.2018.00352DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6191492PMC
October 2018

Thymus-derived Foxp3 regulatory T cells upregulate RORγt expression under inflammatory conditions.

J Mol Med (Berl) 2018 12 24;96(12):1387-1394. Epub 2018 Oct 24.

Department Experimental Immunology, Helmholtz Centre for Infection Research, Inhoffenstrasse 7, 38124, Braunschweig, Germany.

Foxp3 regulatory T cells (Tregs) co-expressing the Th17-lineage specification factor RORγt represent a unique Treg subpopulation that has been reported to be induced upon response to gut microbiota within the intestinal immune system. Hence, RORγt Tregs are considered to solely consist of peripherally induced Foxp3 Tregs (pTregs), and the possibility that also thymus-derived Treg (tTregs) can upregulate RORγt expression and contribute to the pool of RORγt Tregs was largely ignored. Here, we expand our knowledge on the origin of RORγt Tregs by demonstrating that also tTregs can attain RORγt expression. In transgenic Foxp3 reporter mice, a substantial fraction of CNS1-independent Tregs, predominantly consisting of tTregs, was found to co-express RORγt. In addition, genuine tTregs isolated from thymi of Foxp3RAG reporter mice initiated RORγt expression both in vitro and in vivo, particularly under inflammatory conditions. In conclusion, our data demonstrate that tTregs can upregulate RORγt expression under inflammatory conditions and that hence RORγt Tregs can be regarded as a heterogeneous population consisting of both pTregs and tTregs. KEY MESSAGES: RORγt cannot be considered as a marker for pTregs. CNS1-independent tTregs within the colon display RORγt expression. RORγt can be induced in genuine tTregs, particularly under inflammatory conditions. RORγt Tregs are a heterogeneous population consisting of both pTregs and tTregs.
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http://dx.doi.org/10.1007/s00109-018-1706-xDOI Listing
December 2018

Neonatally imprinted stromal cell subsets induce tolerogenic dendritic cells in mesenteric lymph nodes.

Nat Commun 2018 09 25;9(1):3903. Epub 2018 Sep 25.

Department Experimental Immunology, Helmholtz Centre for Infection Research, 38124, Braunschweig, Germany.

Gut-draining mesenteric lymph nodes (mLNs) are important for inducing peripheral tolerance towards food and commensal antigens by providing an optimal microenvironment for de novo generation of Foxp3 regulatory T cells (Tregs). We previously identified microbiota-imprinted mLN stromal cells as a critical component in tolerance induction. Here we show that this imprinting process already takes place in the neonatal phase, and renders the mLN stromal cell compartment resistant to inflammatory perturbations later in life. LN transplantation and single-cell RNA-seq uncover stably imprinted expression signatures in mLN fibroblastic stromal cells. Subsetting common stromal cells across gut-draining mLNs and skin-draining LNs further refine their location-specific immunomodulatory functions, such as subset-specific expression of Aldh1a2/3. Finally, we demonstrate that mLN stromal cells shape resident dendritic cells to attain high Treg-inducing capacity in a Bmp2-dependent manner. Thus, crosstalk between mLN stromal and resident dendritic cells provides a robust regulatory mechanism for the maintenance of intestinal tolerance.
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http://dx.doi.org/10.1038/s41467-018-06423-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6156403PMC
September 2018

Transmaternal Helicobacter pylori exposure reduces allergic airway inflammation in offspring through regulatory T cells.

J Allergy Clin Immunol 2019 04 19;143(4):1496-1512.e11. Epub 2018 Sep 19.

Institute of Molecular Cancer Research, University of Zürich, Zurich, Switzerland. Electronic address:

Background: Transmaternal exposure to tobacco, microbes, nutrients, and other environmental factors shapes the fetal immune system through epigenetic processes. The gastric microbe Helicobacter pylori represents an ancestral constituent of the human microbiota that causes gastric disorders on the one hand and is inversely associated with allergies and chronic inflammatory conditions on the other.

Objective: Here we investigate the consequences of transmaternal exposure to H pylori in utero and/or during lactation for susceptibility to viral and bacterial infection, predisposition to allergic airway inflammation, and development of immune cell populations in the lungs and lymphoid organs.

Methods: We use experimental models of house dust mite- or ovalbumin-induced airway inflammation and influenza A virus or Citrobacter rodentium infection along with metagenomics analyses, multicolor flow cytometry, and bisulfite pyrosequencing, to study the effects of H pylori on allergy severity and immunologic and microbiome correlates thereof.

Results: Perinatal exposure to H pylori extract or its immunomodulator vacuolating cytotoxin confers robust protective effects against allergic airway inflammation not only in first- but also second-generation offspring but does not increase susceptibility to viral or bacterial infection. Immune correlates of allergy protection include skewing of regulatory over effector T cells, expansion of regulatory T-cell subsets expressing CXCR3 or retinoic acid-related orphan receptor γt, and demethylation of the forkhead box P3 (FOXP3) locus. The composition and diversity of the gastrointestinal microbiota is measurably affected by perinatal H pylori exposure.

Conclusion: We conclude that exposure to H pylori has consequences not only for the carrier but also for subsequent generations that can be exploited for interventional purposes.
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http://dx.doi.org/10.1016/j.jaci.2018.07.046DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6592617PMC
April 2019

Epigenetic mechanisms regulating T-cell responses.

J Allergy Clin Immunol 2018 09;142(3):728-743

Regensburg Center for Interventional Immunology (RCI), Regensburg, Germany; Chair for Immunology, University Regensburg and University Medical Center Regensburg, Regensburg, Germany. Electronic address:

During the last decade, advances in sequencing technologies allowed production of a wealth of information on epigenetic modifications in T cells. Epigenome maps, in combination with mechanistic studies, have demonstrated that T cells undergo extensive epigenome remodeling in response to signals, which has a strong effect on phenotypic stability and function of lymphocytes. In this review we focus on DNA methylation, histone modifications, and chromatin structure as important epigenetic mechanisms involved in controlling T-cell responses. In particular, we discuss epigenetic processes in light of the development, activation, and differentiation of CD4 T helper (T), regulatory T, and CD8 T cells. As central aspects of the adaptive immune system, we review mechanisms that ensure molecular memory, stability, plasticity, and exhaustion of T cells. We further discuss the effect of the tissue environment on imprinting T-cell epigenomes with potential implications for immunotherapy.
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http://dx.doi.org/10.1016/j.jaci.2018.07.014DOI Listing
September 2018

Intact interleukin-10 receptor signaling protects from hippocampal damage elicited by experimental neurotropic virus infection of SJL mice.

Sci Rep 2018 04 17;8(1):6106. Epub 2018 Apr 17.

Department of Pathology, University of Veterinary Medicine Hannover, Hannover, Germany.

Theiler's murine encephalomyelitis virus (TMEV) infection represents an experimental mouse model to study hippocampal damage induced by neurotropic viruses. IL-10 is a pleiotropic cytokine with profound anti-inflammatory properties, which critically controls immune homeostasis. In order to analyze IL-10R signaling following virus-induced polioencephalitis, SJL mice were intracerebrally infected with TMEV. RNA-based next generation sequencing revealed an up-regulation of Il10, Il10rα and further genes involved in IL-10 downstream signaling, including Jak1, Socs3 and Stat3 in the brain upon infection. Subsequent antibody-mediated blockade of IL-10R signaling led to enhanced hippocampal damage with neuronal loss and increased recruitment of CD3 T cells, CD45R B cells and an up-regulation of Il1α mRNA. Increased expression of Tgfβ and Foxp3 as well as accumulation of Foxp3 regulatory T cells and arginase-1 macrophages/microglia was detected in the hippocampus, representing a potential compensatory mechanism following disturbed IL-10R signaling. Additionally, an increased peripheral Chi3l3 expression was found in spleens of infected mice, which may embody reactive regulatory mechanisms for prevention of excessive immunopathology. The present study highlights the importance of IL-10R signaling for immune regulation and its neuroprotective properties in the context of an acute neurotropic virus infection.
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http://dx.doi.org/10.1038/s41598-018-24378-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5904160PMC
April 2018

Regulation of neuroinflammatory properties of glial cells by T cell effector molecules.

Neural Regen Res 2018 Feb;13(2):234-236

Clinical Neuroimmunology and Neurochemistry, Department of Neurology; Center of Systems Neuroscience, Hannover Medical School, Hannover, Germany.

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http://dx.doi.org/10.4103/1673-5374.226385DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5879891PMC
February 2018

The invasin D protein from selectively binds the Fab region of host antibodies and affects colonization of the intestine.

J Biol Chem 2018 06 13;293(22):8672-8690. Epub 2018 Mar 13.

From the Young Investigator Group Structural Biology of Autophagy, Department of Structure and Function of Proteins,

is a Gram-negative bacterium and zoonotic pathogen responsible for a wide range of diseases, ranging from mild diarrhea, enterocolitis, lymphatic adenitis to persistent local inflammation. The invasin D (InvD) molecule belongs to the invasin (InvA)-type autotransporter proteins, but its structure and function remain unknown. In this study, we present the first crystal structure of InvD, analyzed its expression and function in a murine infection model, and identified its target molecule in the host. We found that InvD is induced at 37 °C and expressed 2-4 days after infection, indicating that InvD is a virulence factor. During infection, InvD was expressed in all parts of the intestinal tract, but not in deeper lymphoid tissues. The crystal structure of the C-terminal adhesion domain of InvD revealed a distinct Ig-related fold that, apart from the canonical β-sheets, comprises various modifications of and insertions into the Ig-core structure. We identified the Fab fragment of host-derived IgG/IgA antibodies as the target of the adhesion domain. Phage display panning and flow cytometry data further revealed that InvD exhibits a preferential binding specificity toward antibodies with VH3/VK1 variable domains and that it is specifically recruited to a subset of B cells. This finding suggests that InvD modulates Ig functions in the intestine and affects direct interactions with a subset of cell surface-exposed B-cell receptors. In summary, our results provide extensive insights into the structure of InvD and its specific interaction with the target molecule in the host.
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http://dx.doi.org/10.1074/jbc.RA117.001068DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5986219PMC
June 2018

Roquin Suppresses the PI3K-mTOR Signaling Pathway to Inhibit T Helper Cell Differentiation and Conversion of Treg to Tfr Cells.

Immunity 2017 12 12;47(6):1067-1082.e12. Epub 2017 Dec 12.

Institute for Immunology, Biomedical Center, Ludwig-Maximilians-Universität München, 82152 Planegg-Martinsried, Germany; Research Unit Molecular Immune Regulation, Helmholtz Zentrum München, 81377 München, Germany. Electronic address:

Roquin proteins preclude spontaneous T cell activation and aberrant differentiation of T follicular helper (Tfh) or T helper 17 (Th17) cells. Here we showed that deletion of Roquin-encoding alleles specifically in regulatory T (Treg) cells also caused the activation of conventional T cells. Roquin-deficient Treg cells downregulated CD25, acquired a follicular Treg (Tfr) cell phenotype, and suppressed germinal center reactions but could not protect from colitis. Roquin inhibited the PI3K-mTOR signaling pathway by upregulation of Pten through interfering with miR-17∼92 binding to an overlapping cis-element in the Pten 3' UTR, and downregulated the Foxo1-specific E3 ubiquitin ligase Itch. Loss of Roquin enhanced Akt-mTOR signaling and protein synthesis, whereas inhibition of PI3K or mTOR in Roquin-deficient T cells corrected enhanced Tfh and Th17 or reduced iTreg cell differentiation. Thereby, Roquin-mediated control of PI3K-mTOR signaling prevents autoimmunity by restraining activation and differentiation of conventional T cells and specialization of Treg cells.
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http://dx.doi.org/10.1016/j.immuni.2017.11.008DOI Listing
December 2017

Effectors of Th1 and Th17 cells act on astrocytes and augment their neuroinflammatory properties.

J Neuroinflammation 2017 Oct 16;14(1):204. Epub 2017 Oct 16.

Clinical Neuroimmunology and Neurochemistry, Department of Neurology, Hannover Medical School, Carl-Neuberg-Str. 1, 30625, Hannover, Germany.

Background: Autoreactive Th1 and Th17 cells are believed to mediate the pathology of multiple sclerosis in the central nervous system (CNS). Their interaction with microglia and astrocytes in the CNS is crucial for the regulation of the neuroinflammation. Previously, we have shown that only Th1 but not Th17 effectors activate microglia. However, it is not clear which cells are targets of Th17 effectors in the CNS.

Methods: To understand the effects driven by Th17 cells in the CNS, we induced experimental autoimmune encephalomyelitis in wild-type mice and CD4 T cell-specific integrin α4-deficient mice where trafficking of Th1 cells into the CNS was affected. We compared microglial and astrocyte response in the brain and spinal cord of these mice. We further treated astrocytes with supernatants from highly pure Th1 and Th17 cultures and assessed the messenger RNA expression of neurotrophic factors, cytokines and chemokines, using real-time PCR. Data obtained was analyzed using the Kruskal-Wallis test.

Results: We observed in α4-deficient mice weak microglial activation but comparable astrogliosis to that of wild-type mice in the regions of the brain populated with Th17 infiltrates, suggesting that Th17 cells target astrocytes and not microglia. In vitro, in response to supernatants from Th1 and Th17 cultures, astrocytes showed altered expression of neurotrophic factors, pro-inflammatory cytokines and chemokines. Furthermore, increased expression of chemokines in Th1- and Th17-treated astrocytes enhanced recruitment of microglia and transendothelial migration of Th17 cells in vitro.

Conclusion: Our results demonstrate the delicate interaction between T cell subsets and glial cells and how they communicate to mediate their effects. Effectors of Th1 act on both microglia and astrocytes whereas Th17 effectors preferentially target astrocytes to promote neuroinflammation.
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http://dx.doi.org/10.1186/s12974-017-0978-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5644084PMC
October 2017