49 results on '"Kurschus FC"'
Search Results
2. IL-17 Signaling in Keratinocytes Orchestrates the Defense against Staphylococcus aureus Skin Infection.
- Author
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Moos S, Regen T, Wanke F, Tian Y, Arendholz LT, Hauptmann J, Heinen AP, Bleul L, Bier K, El Malki K, Reinhardt C, Prinz I, Diefenbach A, Wolz C, Schittek B, Waisman A, and Kurschus FC
- Subjects
- Mice, Animals, Mice, Knockout, Skin, Keratinocytes, Mice, Inbred C57BL, Interleukin-17, Staphylococcus aureus
- Abstract
Keratinocytes (KCs) form the outer epithelial barrier of the body, protecting against invading pathogens. Mice lacking the IL-17RA or both IL-17A and IL-17F develop spontaneous Staphylococcusaureus skin infections. We found a marked expansion of T
17 cells, comprised of RORγt-expressing γδ T cells and T helper 17 cells in the skin-draining lymph nodes of these mice. Contradictory to previous suggestions, this expansion was not a result of a direct negative feedback loop because we found no expansion of T17 cells in mice lacking IL-17 signaling specifically in T cells. Instead, we found that the T17 expansion depended on the microbiota and was observed only when KCs were deficient for IL-17RA signaling. Indeed, mice that lack IL-17RA only in KCs showed an increased susceptibility to experimental epicutaneous infection with S. aureus together with an accumulation of IL-17A-producing γδ T cells. We conclude that deficiency of IL-17RA on KCs leads to microbiota dysbiosis in the skin, which triggers the expansion of IL-17A-producing T cells. Our data show that KCs are the primary target cells of IL-17A and IL-17F, coordinating the defense against microbial invaders in the skin., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)- Published
- 2023
- Full Text
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3. Skin Sodium Accumulates in Psoriasis and Reflects Disease Severity.
- Author
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Maifeld A, Wild J, Karlsen TV, Rakova N, Wistorf E, Linz P, Jung R, Birukov A, Gimenez-Rivera VA, Wilck N, Bartolomaeus T, Dechend R, Kleinewietfeld M, Forslund SK, Krause A, Kokolakis G, Philipp S, Clausen BE, Brand A, Waisman A, Kurschus FC, Wegner J, Schultheis M, Luft FC, Boschmann M, Kelm M, Wiig H, Kuehne T, Müller DN, Karbach S, and Markó L
- Subjects
- Animals, Cell Differentiation, Cells, Cultured, Humans, Lymphocyte Activation, Male, Mice, Mice, Inbred C57BL, Severity of Illness Index, Skin pathology, Sodium Chloride metabolism, Spectrophotometry, Atomic, Spectrum Analysis, Interleukin-17 metabolism, Psoriasis metabolism, Skin metabolism, Sodium analysis, Th17 Cells immunology
- Abstract
Sodium can accumulate in the skin at concentrations exceeding serum levels. A high sodium environment can lead to pathogenic T helper 17 cell expansion. Psoriasis is a chronic inflammatory skin disease in which IL-17‒producing T helper 17 cells play a crucial role. In an observational study, we measured skin sodium content in patients with psoriasis and in age-matched healthy controls by Sodium-23 magnetic resonance imaging. Patients with PASI > 5 showed significantly higher sodium and water content in the skin but not in other tissues than those with lower PASI or healthy controls. Skin sodium concentrations measured by Sodium-23 spectroscopy or by atomic absorption spectrometry in ashed-skin biopsies verified the findings with Sodium-23 magnetic resonance imaging. In vitro T helper 17 cell differentiation of naive CD4
+ cells from patients with psoriasis markedly induced IL-17A expression under increased sodium chloride concentrations. The imiquimod-induced psoriasis mouse model replicated the human findings. Extracellular tracer Chromium-51-EDTA measurements in imiquimod- and sham-treated skin showed similar extracellular volumes, rendering excessive water of intracellular origin. Chronic genetic IL-17A‒driven psoriasis mouse models underlined the role of IL-17A in dermal sodium accumulation and inflammation. Our data describe skin sodium as a pathophysiological feature of psoriasis, which could open new avenues for its treatment., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)- Published
- 2022
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4. Posttranslational modifications by ADAM10 shape myeloid antigen-presenting cell homeostasis in the splenic marginal zone.
- Author
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Diener N, Fontaine JF, Klein M, Hieronymus T, Wanke F, Kurschus FC, Ludwig A, Ware C, Saftig P, Bopp T, Clausen BE, and Backer RA
- Subjects
- ADAM10 Protein physiology, Amyloid Precursor Protein Secretases physiology, Animals, Antigen-Presenting Cells metabolism, CD11c Antigen metabolism, Cell Differentiation, Cell Proliferation, Female, Homeostasis, Lymphoid Tissue metabolism, Macrophages metabolism, Male, Membrane Proteins physiology, Mice, Mice, Inbred C57BL, Myeloid Cells metabolism, Phosphatidylinositol 3-Kinases metabolism, Protein Processing, Post-Translational genetics, Protein Processing, Post-Translational physiology, Signal Transduction, Spleen cytology, Spleen metabolism, ADAM10 Protein metabolism, Amyloid Precursor Protein Secretases metabolism, Dendritic Cells metabolism, Membrane Proteins metabolism
- Abstract
The spleen contains phenotypically and functionally distinct conventional dendritic cell (cDC) subpopulations, termed cDC1 and cDC2, which each can be divided into several smaller and less well-characterized subsets. Despite advances in understanding the complexity of cDC ontogeny by transcriptional programming, the significance of posttranslational modifications in controlling tissue-specific cDC subset immunobiology remains elusive. Here, we identified the cell-surface-expressed A-disintegrin-and-metalloproteinase 10 (ADAM10) as an essential regulator of cDC1 and cDC2 homeostasis in the splenic marginal zone (MZ). Mice with a CD11c-specific deletion of ADAM10 (ADAM10
ΔCD11c ) exhibited a complete loss of splenic ESAMhi cDC2A because ADAM10 regulated the commitment, differentiation, and survival of these cells. The major pathways controlled by ADAM10 in ESAMhi cDC2A are Notch, signaling pathways involved in cell proliferation and survival (e.g., mTOR, PI3K/AKT, and EIF2 signaling), and EBI2-mediated localization within the MZ. In addition, we discovered that ADAM10 is a molecular switch regulating cDC2 subset heterogeneity in the spleen, as the disappearance of ESAMhi cDC2A in ADAM10ΔCD11c mice was compensated for by the emergence of a Clec12a+ cDC2B subset closely resembling cDC2 generally found in peripheral lymph nodes. Moreover, in ADAM10ΔCD11c mice, terminal differentiation of cDC1 was abrogated, resulting in severely reduced splenic Langerin+ cDC1 numbers. Next to the disturbed splenic cDC compartment, ADAM10 deficiency on CD11c+ cells led to an increase in marginal metallophilic macrophage (MMM) numbers. In conclusion, our data identify ADAM10 as a molecular hub on both cDC and MMM regulating their transcriptional programming, turnover, homeostasis, and ability to shape the anatomical niche of the MZ., Competing Interests: The authors declare no competing interest.- Published
- 2021
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5. Laquinimod dampens IL-1β signaling and Th17-polarizing capacity of monocytes in patients with MS.
- Author
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Engel S, Jolivel V, Kraus SH, Zayoud M, Rosenfeld K, Tumani H, Furlan R, Kurschus FC, Waisman A, and Luessi F
- Subjects
- Adult, Cross-Sectional Studies, Female, Humans, Interleukin-1beta metabolism, Lymphocyte Activation drug effects, Male, Middle Aged, Monocytes immunology, Multiple Sclerosis, Relapsing-Remitting immunology, Signal Transduction drug effects, Interleukin-1beta immunology, Monocytes drug effects, Multiple Sclerosis, Relapsing-Remitting drug therapy, Quinolones therapeutic use, Th17 Cells immunology
- Abstract
Objective: To assess the impact of laquinimod treatment on monocytes and to investigate the underlying immunomodulatory mechanisms in MS., Methods: In this cross-sectional study, we performed in vivo and in vitro analyses of cluster of differentiation (CD14
+ ) monocytes isolated from healthy donors (n = 15), untreated (n = 13), and laquinimod-treated patients with MS (n = 14). Their frequency and the expression of surface activation markers were assessed by flow cytometry and the viability by calcein staining. Cytokine concentrations in the supernatants of lipopolysaccharide (LPS)-stimulated monocytes were determined by flow cytometry. The messenger ribonucleic acid (mRNA) expression level of genes involved in cytokine expression was measured by quantitative PCR. The LPS-mediated nuclear factor kappa-light-chain-enhancer of activated B-cell (NF-κB) activation was determined by the quantification of the phosphorylation level of the p65 subunit. Laquinimod-treated monocytes were cocultured with CD4+ T cells, and the resulting cytokine production was analyzed by flow cytometry after intracellular cytokine staining. The interleukin (IL)-17A concentration of the supernatant was assessed by ELISA., Results: Laquinimod did not alter the frequency or viability of circulating monocytes, but led to an upregulation of CD86 expression. LPS-stimulated monocytes of laquinimod-treated patients with MS secreted less IL-1β following a downregulation of IL-1β gene expression. Phosphorylation levels of the NF-κB p65 subunit were reduced after laquinimod treatment, indicating a laquinimod-associated inhibition of the NF-κB pathway. T cells primed with laquinimod-treated monocytes differentiated significantly less into IL-17A-producing T helper (Th)-17 cells., Conclusions: Our findings suggest that inhibited NF-κB signaling and downregulation of IL-1β expression in monocytes contributes to the immunomodulatory effects of laquinimod and that the impairment of Th17 polarization might mediate its disease-modifying activity in MS., (Copyright © 2020 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology.)- Published
- 2020
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6. Interleukin-1 promotes autoimmune neuroinflammation by suppressing endothelial heme oxygenase-1 at the blood-brain barrier.
- Author
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Hauptmann J, Johann L, Marini F, Kitic M, Colombo E, Mufazalov IA, Krueger M, Karram K, Moos S, Wanke F, Kurschus FC, Klein M, Cardoso S, Strauß J, Bolisetty S, Lühder F, Schwaninger M, Binder H, Bechman I, Bopp T, Agarwal A, Soares MP, Regen T, and Waisman A
- Subjects
- Animals, Blood-Brain Barrier immunology, Encephalomyelitis, Autoimmune, Experimental enzymology, Gene Expression Regulation immunology, Mice, Mice, Inbred C57BL, Signal Transduction immunology, Blood-Brain Barrier enzymology, Encephalomyelitis, Autoimmune, Experimental immunology, Endothelial Cells enzymology, Heme Oxygenase-1 metabolism, Inflammation immunology, Interleukin-1 immunology
- Abstract
The proinflammatory cytokine interleukin 1 (IL-1) is crucially involved in the pathogenesis of multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE). Herein, we studied the role of IL-1 signaling in blood-brain barrier (BBB) endothelial cells (ECs), astrocytes and microglia for EAE development, using mice with the conditional deletion of its signaling receptor IL-1R1. We found that IL-1 signaling in microglia and astrocytes is redundant for the development of EAE, whereas the IL-1R1 deletion in BBB-ECs markedly ameliorated disease severity. IL-1 signaling in BBB-ECs upregulated the expression of the adhesion molecules Vcam-1, Icam-1 and the chemokine receptor Darc, all of which have been previously shown to promote CNS-specific inflammation. In contrast, IL-1R1 signaling suppressed the expression of the stress-responsive heme catabolizing enzyme heme oxygenase-1 (HO-1) in BBB-ECs, promoting disease progression via a mechanism associated with deregulated expression of the IL-1-responsive genes Vcam1, Icam1 and Ackr1 (Darc). Mechanistically, our data emphasize a functional crosstalk of BBB-EC IL-1 signaling and HO-1, controlling the transcription of downstream proinflammatory genes promoting the pathogenesis of autoimmune neuroinflammation.
- Published
- 2020
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7. Review-Current Concepts in Inflammatory Skin Diseases Evolved by Transcriptome Analysis: In-Depth Analysis of Atopic Dermatitis and Psoriasis.
- Author
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Schwingen J, Kaplan M, and Kurschus FC
- Subjects
- Computational Biology, Databases, Factual, Dermatitis, Atopic genetics, Gene Expression Profiling methods, Humans, Prognosis, Psoriasis genetics, RNA-Seq methods, Skin metabolism, Skin Diseases classification, Skin Diseases genetics, Dermatitis, Atopic metabolism, Psoriasis metabolism, Skin Diseases metabolism, Transcriptome
- Abstract
During the last decades, high-throughput assessment of gene expression in patient tissues using microarray technology or RNA-Seq took center stage in clinical research. Insights into the diversity and frequency of transcripts in healthy and diseased conditions provide valuable information on the cellular status in the respective tissues. Growing with the technique, the bioinformatic analysis toolkit reveals biologically relevant pathways which assist in understanding basic pathophysiological mechanisms. Conventional classification systems of inflammatory skin diseases rely on descriptive assessments by pathologists. In contrast to this, molecular profiling may uncover previously unknown disease classifying features. Thereby, treatments and prognostics of patients may be improved. Furthermore, disease models in basic research in comparison to the human disease can be directly validated. The aim of this article is not only to provide the reader with information on the opportunities of these techniques, but to outline potential pitfalls and technical limitations as well. Major published findings are briefly discussed to provide a broad overview on the current findings in transcriptomics in inflammatory skin diseases.
- Published
- 2020
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8. IL-17 + CD8 + T cell suppression by dimethyl fumarate associates with clinical response in multiple sclerosis.
- Author
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Lückel C, Picard F, Raifer H, Campos Carrascosa L, Guralnik A, Zhang Y, Klein M, Bittner S, Steffen F, Moos S, Marini F, Gloury R, Kurschus FC, Chao YY, Bertrams W, Sexl V, Schmeck B, Bonetti L, Grusdat M, Lohoff M, Zielinski CE, Zipp F, Kallies A, Brenner D, Berger M, Bopp T, Tackenberg B, and Huber M
- Subjects
- Adolescent, Adult, Animals, CD8-Positive T-Lymphocytes immunology, CD8-Positive T-Lymphocytes metabolism, Dimethyl Fumarate therapeutic use, Encephalomyelitis, Autoimmune, Experimental blood, Encephalomyelitis, Autoimmune, Experimental immunology, Female, Humans, Immunosuppressive Agents, Interleukin-17 immunology, Interleukin-17 metabolism, Longitudinal Studies, Male, Mice, Middle Aged, Multiple Sclerosis blood, Multiple Sclerosis immunology, Th17 Cells drug effects, Th17 Cells immunology, Treatment Outcome, Young Adult, CD8-Positive T-Lymphocytes drug effects, Dimethyl Fumarate pharmacology, Encephalomyelitis, Autoimmune, Experimental drug therapy, Multiple Sclerosis drug therapy
- Abstract
IL-17-producing CD8
+ (Tc17) cells are enriched in active lesions of patients with multiple sclerosis (MS), suggesting a role in the pathogenesis of autoimmunity. Here we show that amelioration of MS by dimethyl fumarate (DMF), a mechanistically elusive drug, associates with suppression of Tc17 cells. DMF treatment results in reduced frequency of Tc17, contrary to Th17 cells, and in a decreased ratio of the regulators RORC-to-TBX21, along with a shift towards cytotoxic T lymphocyte gene expression signature in CD8+ T cells from MS patients. Mechanistically, DMF potentiates the PI3K-AKT-FOXO1-T-BET pathway, thereby limiting IL-17 and RORγt expression as well as STAT5-signaling in a glutathione-dependent manner. This results in chromatin remodeling at the Il17 locus. Consequently, T-BET-deficiency in mice or inhibition of PI3K-AKT, STAT5 or reactive oxygen species prevents DMF-mediated Tc17 suppression. Overall, our data disclose a DMF-AKT-T-BET driven immune modulation and suggest putative therapy targets in MS and beyond.- Published
- 2019
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9. Keratinocyte-derived IκBζ drives psoriasis and associated systemic inflammation.
- Author
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Lorscheid S, Müller A, Löffler J, Resch C, Bucher P, Kurschus FC, Waisman A, Schäkel K, Hailfinger S, Schulze-Osthoff K, and Kramer D
- Subjects
- Adaptor Proteins, Signal Transducing genetics, Animals, Cells, Cultured, Female, Interleukin-17 genetics, Interleukin-17 metabolism, Male, Mice, Mice, Transgenic, Skin cytology, Skin pathology, Adaptor Proteins, Signal Transducing metabolism, Inflammation metabolism, Keratinocytes metabolism, Psoriasis metabolism
- Abstract
The transcriptional activator IκBζ is a key regulator of psoriasis, but which cells mediate its pathogenic effect remains unknown. Here we found that IκBζ expression in keratinocytes triggers not only skin lesions but also systemic inflammation in mouse psoriasis models. Specific depletion of IκBζ in keratinocytes was sufficient to suppress the induction of imiquimod- or IL-36-mediated psoriasis. Moreover, IκBζ ablation in keratinocytes prevented the onset of psoriatic lesions and systemic inflammation in keratinocyte-specific IL-17A-transgenic mice. Mechanistically, this psoriasis protection was mediated by IκBζ deficiency in keratinocytes abrogating the induction of specific proinflammatory target genes, including Cxcl5, Cxcl2, Csf2, and Csf3, in response to IL-17A or IL-36. These IκBζ-dependent genes trigger the generation and recruitment of neutrophils and monocytes that are needed for skin inflammation. Consequently, our data uncover a surprisingly pivotal role of keratinocytes and keratinocyte-derived IκBζ as key mediators of psoriasis and psoriasis-related systemic inflammation.
- Published
- 2019
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10. Dietary tryptophan links encephalogenicity of autoreactive T cells with gut microbial ecology.
- Author
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Sonner JK, Keil M, Falk-Paulsen M, Mishra N, Rehman A, Kramer M, Deumelandt K, Röwe J, Sanghvi K, Wolf L, von Landenberg A, Wolff H, Bharti R, Oezen I, Lanz TV, Wanke F, Tang Y, Brandao I, Mohapatra SR, Epping L, Grill A, Röth R, Niesler B, Meuth SG, Opitz CA, Okun JG, Reinhardt C, Kurschus FC, Wick W, Bode HB, Rosenstiel P, and Platten M
- Subjects
- Animals, Dietary Proteins, Disease Models, Animal, Encephalomyelitis, Autoimmune, Experimental microbiology, Gastrointestinal Microbiome genetics, Mice, Multiple Sclerosis, RNA, Ribosomal, 16S genetics, Autoimmunity immunology, Diet, Encephalomyelitis, Autoimmune, Experimental immunology, Gastrointestinal Microbiome immunology, T-Lymphocytes immunology, Tryptophan
- Abstract
The interaction between the mammalian host and its resident gut microbiota is known to license adaptive immune responses. Nutritional constituents strongly influence composition and functional properties of the intestinal microbial communities. Here, we report that omission of a single essential amino acid - tryptophan - from the diet abrogates CNS autoimmunity in a mouse model of multiple sclerosis. Dietary tryptophan restriction results in impaired encephalitogenic T cell responses and is accompanied by a mild intestinal inflammatory response and a profound phenotypic shift of gut microbiota. Protective effects of dietary tryptophan restriction are abrogated in germ-free mice, but are independent of canonical host sensors of intracellular tryptophan metabolites. We conclude that dietary tryptophan restriction alters metabolic properties of gut microbiota, which in turn have an impact on encephalitogenic T cell responses. This link between gut microbiota, dietary tryptophan and adaptive immunity may help to develop therapeutic strategies for protection from autoimmune neuroinflammation.
- Published
- 2019
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11. Dimethyl fumarate alters intracellular Ca 2+ handling in immune cells by redox-mediated pleiotropic effects.
- Author
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Herrmann AK, Wüllner V, Moos S, Graf J, Chen J, Kieseier B, Kurschus FC, Albrecht P, Vangheluwe P, and Methner A
- Subjects
- Animals, Calcium metabolism, Calcium Signaling drug effects, Disease Models, Animal, Gene Expression Regulation drug effects, Glutathione metabolism, Humans, Lymphocytes drug effects, Mice, Multiple Sclerosis genetics, Multiple Sclerosis pathology, Oxidation-Reduction drug effects, Psoriasis genetics, Psoriasis pathology, Reactive Oxygen Species metabolism, Dimethyl Fumarate pharmacology, Multiple Sclerosis drug therapy, Psoriasis drug therapy, Sarcoplasmic Reticulum Calcium-Transporting ATPases genetics, TRPA1 Cation Channel genetics
- Abstract
Dimethyl fumarate (DMF) is widely used to treat the human autoimmune diseases multiple sclerosis (MS) and psoriasis. DMF causes short-term oxidative stress and activates the antioxidant response via the transcription factor Nrf2 but its immunosuppressive effect is not well understood. Immune cell activation depends on calcium signaling which itself is influenced by the cellular redox state. We therefore measured calcium, reactive oxygen species levels and glutathione content in lymphocytes from immunized mice before onset of experimental autoimmune encephalomyelitis, in peripheral blood mononuclear cells from MS patients treated with DMF, and in mouse splenocytes treated ex vivo with DMF. This demonstrated altered redox states and increased lymphocytic calcium levels in all model systems. DMF caused an immediate influx of calcium from the extracellular space, long-term increased cytosolic calcium levels and reduced calcium stored in intracellular stores. The DMF-elicited current had the electrophysiological characteristics of a transient receptor potential channel and the intracellular calcium levels were normalized by antagonists of TRPA1. Interestingly, the sarco/endoplasmic reticulum Ca
2+ -ATPase SERCA2b was downregulated but more active due to glutathionylation of the redox-sensitive cysteine 674. DMF therefore causes pleiotropic changes in cellular calcium homeostasis which are likely caused by redox-sensitive post-translational modifications. These changes probably contribute to its immunosuppressive effects., (Copyright © 2019 Elsevier Inc. All rights reserved.)- Published
- 2019
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12. Lugdunin amplifies innate immune responses in the skin in synergy with host- and microbiota-derived factors.
- Author
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Bitschar K, Sauer B, Focken J, Dehmer H, Moos S, Konnerth M, Schilling NA, Grond S, Kalbacher H, Kurschus FC, Götz F, Krismer B, Peschel A, and Schittek B
- Subjects
- Animals, Anti-Bacterial Agents therapeutic use, Antimicrobial Cationic Peptides immunology, Cells, Cultured, Disease Models, Animal, Female, Humans, Keratinocytes drug effects, Keratinocytes immunology, Keratinocytes microbiology, Mice, Mice, Inbred C57BL, Mice, Knockout, Microbiota drug effects, Microbiota immunology, Peptides immunology, Peptides, Cyclic therapeutic use, Primary Cell Culture, Skin drug effects, Skin immunology, Skin microbiology, Staphylococcal Infections immunology, Staphylococcal Infections microbiology, Thiazolidines therapeutic use, Cathelicidins, Anti-Bacterial Agents pharmacology, Immunity, Innate drug effects, Peptides, Cyclic pharmacology, Staphylococcal Infections drug therapy, Staphylococcus aureus drug effects, Thiazolidines pharmacology
- Abstract
Recently our groups discovered lugdunin, a new cyclic peptide antibiotic that inhibits Staphylococcus aureus epithelial colonization in humans and rodents. In this work, we analyzed its immuno-modulatory and antimicrobial potential as a single agent or in combination with other microbiota- or host-derived factors. We show that pretreatment of primary human keratinocytes or mouse skin with lugdunin in combination with microbiota-derived factors results in a significant reduction of S. aureus colonization. Moreover, lugdunin increases expression and release of LL-37 and CXCL8/MIP-2 in human keratinocytes and mouse skin, and results in the recruitment of monocytes and neutrophils in vivo, both by a TLR/MyD88-dependent mechanism. Interestingly, S. aureus elimination by lugdunin is additionally achieved by synergistic antimicrobial activity with LL-37 and dermcidin-derived peptides. In summary, our results indicate that lugdunin provides multi-level protection against S. aureus and may thus become a promising treatment option for S. aureus skin infections in the future.
- Published
- 2019
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13. Imiquimod-Induced Psoriasis in Mice Depends on the IL-17 Signaling of Keratinocytes.
- Author
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Moos S, Mohebiany AN, Waisman A, and Kurschus FC
- Subjects
- Animals, Biopsy, Needle, Cells, Cultured, Cytokines metabolism, Disease Models, Animal, Female, Flow Cytometry, Immunohistochemistry, Keratinocytes drug effects, Keratinocytes metabolism, Male, Mice, Mice, Inbred C57BL, Psoriasis chemically induced, Psoriasis pathology, Random Allocation, Real-Time Polymerase Chain Reaction methods, Signal Transduction genetics, Tumor Necrosis Factor-alpha metabolism, Adjuvants, Immunologic pharmacology, Imiquimod pharmacology, Interleukin-17 genetics, Psoriasis genetics, Signal Transduction drug effects
- Abstract
The pathology of psoriasis strongly depends on IL-17A. Monoclonal antibodies blocking either the cytokine or its receptor are among the most efficient treatments for psoriatic patients. Keratinocytes can be activated upon exposure to IL-17A and tumor necrosis factor-α and secrete secondary cytokines and chemokines in the inflamed skin. In psoriasis and its imiquimod-induced mouse model, a strong skin infiltration of neutrophils and inflammatory monocytes can be observed. However, to date, it is not clear how exactly those cellular populations are attracted to the skin and how they contribute to the pathogenesis of the disease. To define the crucial cell type responding to IL-17 and initiating the downstream pathology in psoriasis-like dermatitis, we used mice specifically lacking the IL-17 receptor (IL-17RA) in different cell types. Deletion of IL-17RA in T cells or myeloid had no impact on disease development. Only deletion of this receptor in keratinocytes reflected the full-body deletion of IL-17RA, resulting in strongly reduced dermatitis development. Imiquimod treatment of those IL-17 signaling-deficient mice maintained high monocytic infiltration but failed to attract neutrophils into the skin. We conclude that keratinocytes are a critical cellular target for IL-17A-mediated neutrophil attraction and psoriasis development., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)
- Published
- 2019
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14. Alternative Splice Forms of CYLD Mediate Ubiquitination of SMAD7 to Prevent TGFB Signaling and Promote Colitis.
- Author
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Tang Y, Reissig S, Glasmacher E, Regen T, Wanke F, Nikolaev A, Gerlach K, Popp V, Karram K, Fantini MC, Schattenberg JM, Galle PR, Neurath MF, Weigmann B, Kurschus FC, Hövelmeyer N, and Waisman A
- Subjects
- Animals, Biopsy, Needle, Deubiquitinating Enzyme CYLD, Disease Models, Animal, Flow Cytometry, Humans, Immunohistochemistry, Intestinal Mucosa metabolism, Intestinal Mucosa pathology, Mice, Mice, Inbred C57BL, Mice, Transgenic, Random Allocation, Reference Values, Signal Transduction, Transforming Growth Factor beta1 genetics, Crohn Disease genetics, Crohn Disease pathology, Cysteine Endopeptidases genetics, Smad7 Protein genetics, Ubiquitination genetics
- Abstract
Background & Aims: The CYLD lysine 63 deubiquitinase gene (CYLD) encodes tumor suppressor protein that is mutated in familial cylindromatosus, and variants have been associated with Crohn disease (CD). Splice forms of CYLD that lack exons 7 and 8 regulate transcription factors and functions of immune cells. We examined the expression of splice forms of CYLD in colon tissues from patients with CD and their effects in mice., Methods: We performed immunohistochemical analyses of colon tissues from patients with untreated CD and patients without inflammatory bowel diseases (controls). We obtained mice that expressed splice forms of CYLD (sCYLD mice) without or with SMAD7 (sCYLD/SMAD7 mice) from transgenes and CYLD-knockout mice (with or without transgenic expression of SMAD7) and performed endoscopic analyses. Colitis was induced in Rag1
-/- mice by transfer of CD4+ CD62L+ T cells from C57/Bl6 or transgenic mice. T cells were isolated from mice and analyzed by flow cytometry and quantitative real-time polymerase chain reaction and intestinal tissues were analyzed by histology and immunohistochemistry. CYLD forms were expressed in mouse embryonic fibroblasts, primary T cells, and HEK293T cells, which were analyzed by immunoblot, mobility shift, and immunoprecipitation assays., Results: The colonic lamina propria from patients with CD was infiltrated by T cells and had higher levels of sCYLD (but not full-length CYLD) and SMAD7 than tissues from controls. Incubation of mouse embryonic fibroblasts and T cells with transforming growth factor β increased their production of sCYLD and decreased full-length CYLD. Transgenic expression of sCYLD and SMAD7 in T cells prevented the differentiation of regulatory T cells and T-helper type 17 cells and increased the differentiation of T-helper type 1 cells. The same effects were observed in colon tissues from sCYLD/SMAD7 mice but not in those from CYLD-knockout SMAD7 mice. The sCYLD mice had significant increases in the numbers of T-helper type 1 cells and CD44high CD62Llow memory-effector CD4+ T cells in the spleen and mesenteric lymph nodes compared with wild-type mice; sCYLD/SMAD7 mice had even larger increases. The sCYLD/SMAD7 mice spontaneously developed severe colitis, with infiltration of the colon by dendritic cells, neutrophils, macrophages, and CD4+ T cells and increased levels of Ifng, Il6, Il12a, Il23a, and Tnf mRNAs. Co-transfer of regulatory T cells from wild-type, but not from sCYLD/SMAD7, mice prevented the induction of colitis in Rag1-/- mice by CD4+ T cells. We found increased levels of poly-ubiquitinated SMAD7 in sCYLD CD4+ T cells. CYLD formed a nuclear complex with SMAD3, whereas sCYLD recruited SMAD7 to the nucleus, which inhibited the expression of genes regulated by SMAD3 and SMAD4. We found that sCYLD mediated lysine 63-linked ubiquitination of SMAD7. The sCYLD-SMAD7 complex inhibited transforming growth factor β signaling in CD4+ T cells., Conclusions: Levels of the spliced form of CYLD are increased in colon tissues from patients with CD. sCYLD mediates ubiquitination and nuclear translocation of SMAD7 and thereby decreases transforming growth factor β signaling in T cells. This prevents immune regulatory mechanisms and leads to colitis in mice., (Copyright © 2019 AGA Institute. Published by Elsevier Inc. All rights reserved.)- Published
- 2019
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15. NF-κB inducing kinase (NIK) is an essential post-transcriptional regulator of T-cell activation affecting F-actin dynamics and TCR signaling.
- Author
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Lacher SM, Thurm C, Distler U, Mohebiany AN, Israel N, Kitic M, Ebering A, Tang Y, Klein M, Wabnitz GH, Wanke F, Samstag Y, Bopp T, Kurschus FC, Simeoni L, Tenzer S, and Waisman A
- Subjects
- Actins genetics, Adaptor Proteins, Signal Transducing genetics, Adaptor Proteins, Signal Transducing immunology, Animals, Central Nervous System immunology, Central Nervous System pathology, Encephalomyelitis, Autoimmune, Experimental chemically induced, Encephalomyelitis, Autoimmune, Experimental genetics, Encephalomyelitis, Autoimmune, Experimental pathology, Gene Expression Profiling, Gene Expression Regulation, Lymph Nodes immunology, Lymph Nodes pathology, Membrane Proteins genetics, Membrane Proteins immunology, Mice, Mice, Inbred C57BL, Mice, Knockout, Mitogen-Activated Protein Kinase 1 genetics, Mitogen-Activated Protein Kinase 1 immunology, Mitogen-Activated Protein Kinase 3 genetics, Mitogen-Activated Protein Kinase 3 immunology, Myelin-Oligodendrocyte Glycoprotein administration & dosage, Peptide Fragments administration & dosage, Phospholipase C gamma genetics, Phospholipase C gamma immunology, Phosphoproteins genetics, Phosphoproteins immunology, Primary Cell Culture, Protein Serine-Threonine Kinases deficiency, Protein Serine-Threonine Kinases genetics, Proto-Oncogene Proteins c-akt genetics, Proto-Oncogene Proteins c-akt immunology, Receptors, Antigen, T-Cell genetics, Signal Transduction, Spleen immunology, Spleen pathology, T-Box Domain Proteins genetics, T-Box Domain Proteins immunology, T-Lymphocytes pathology, ZAP-70 Protein-Tyrosine Kinase genetics, ZAP-70 Protein-Tyrosine Kinase immunology, NF-kappaB-Inducing Kinase, Actins immunology, Encephalomyelitis, Autoimmune, Experimental immunology, Lymphocyte Activation, Protein Serine-Threonine Kinases immunology, Receptors, Antigen, T-Cell immunology, T-Lymphocytes immunology
- Abstract
NF-κB inducing kinase (NIK) is the key protein of the non-canonical NF-κB pathway and is important for the development of lymph nodes and other secondary immune organs. We elucidated the specific role of NIK in T cells using T-cell specific NIK-deficient (NIK
ΔT ) mice. Despite showing normal development of lymphoid organs, NIKΔT mice were resistant to induction of CNS autoimmunity. T cells from NIKΔT mice were deficient in late priming, failed to up-regulate T-bet and to transmigrate into the CNS. Proteomic analysis of activated NIK-/- T cells showed de-regulated expression of proteins involved in the formation of the immunological synapse: in particular, proteins involved in cytoskeleton dynamics. In line with this we found that NIK-deficient T cells were hampered in phosphorylation of Zap70, LAT, AKT, ERK1/2 and PLCγ upon TCR engagement. Hence, our data disclose a hitherto unknown function of NIK in T-cell priming and differentiation., (Copyright © 2018. Published by Elsevier Ltd.)- Published
- 2018
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16. EBI2 - Sensor for dihydroxycholesterol gradients in neuroinflammation.
- Author
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Kurschus FC and Wanke F
- Subjects
- Animals, Chemotaxis, Disease Models, Animal, Encephalomyelitis, Autoimmune, Experimental immunology, Encephalomyelitis, Autoimmune, Experimental metabolism, Humans, Immunity, Cellular, Multiple Sclerosis immunology, Multiple Sclerosis metabolism, Hydroxycholesterols metabolism, Inflammation metabolism, Receptors, G-Protein-Coupled metabolism
- Abstract
Dihydroxycholesterols such as 7α,25-dihydroxysterols (7α,25-OHC) and 7α,27-OHC are generated from cholesterol by the enzymes CH25H, CYP7B1 and CYP27A1 in steady state but also in the context of inflammation. The G-protein coupled receptor (GPCR) Epstein-Barr virus-induced gene 2 (EBI2), also known as GPR183, senses these oxysterols and induces chemotactic migration of immune cells towards higher concentrations of these ligands. We recently showed that these ligands are upregulated in the CNS in experimental autoimmune encephalomyelitis (EAE), an animal model for multiple sclerosis and that EBI2 enhanced early infiltration of encephalitogenic T cells into the CNS. In this short-review we discuss the role of dihydroxysterol-sensing by immune cells in neuroinflammation., (Copyright © 2018 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.)
- Published
- 2018
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17. EBI2 in splenic and local immune responses and in autoimmunity.
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Barington L, Wanke F, Niss Arfelt K, Holst PJ, Kurschus FC, and Rosenkilde MM
- Subjects
- Animals, Humans, Autoimmunity immunology, Immunity, Innate immunology, Receptors, G-Protein-Coupled immunology, Spleen immunology
- Abstract
The seven transmembrane G protein-coupled receptor EBV-induced gene 2 (EBI2), also known as GPR183, is expressed in particular in immune cells. Activated by its endogenous ligands, which are a group of oxysterols, it functions as a chemo-attractant receptor, mediating cell migration. In coordination with other receptors, EBI2 plays important roles in controlling the migration of immune cells during the course of a T-dependent Ab response in the spleen. In recent years, it has become clear that EBI2 also has other roles to play in the immune system. Thus, EBI2 seems to be involved in innate immune responses, such as those mediated by TLR signaling, and it has been implicated in regional immune responses, including immune responses in the CNS. In this review, we describe the functions of EBI2 in B cells, T cells, and dendritic cells during the course of a T-dependent Ab response in the spleen. Furthermore, we review the existing evidence supporting a role for EBI2 in local immune responses and in autoimmune diseases, with a special focus on immune responses in the CNS. Finally, we discuss which type of role EBI2 may play in autoimmune diseases, and we give our opinion about the paths of future research in EBI2., (©2018 Society for Leukocyte Biology.)
- Published
- 2018
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18. Expression of IL-17F is associated with non-pathogenic Th17 cells.
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Wanke F, Tang Y, Gronke K, Klebow S, Moos S, Hauptmann J, Shanmugavadivu A, Regen T, Mufazalov IA, Gabriel LA, Reißig S, Diefenbach A, Kurschus FC, and Waisman A
- Subjects
- Animals, Biomarkers, Cell Differentiation immunology, Disease Susceptibility, Encephalomyelitis, Autoimmune, Experimental, Immunophenotyping, Interleukin-17 metabolism, Intestinal Mucosa immunology, Intestinal Mucosa metabolism, Intestinal Mucosa pathology, Mice, Mice, Transgenic, T-Lymphocyte Subsets immunology, T-Lymphocyte Subsets metabolism, Th17 Cells cytology, Th17 Cells immunology, Gene Expression, Interleukin-17 genetics, Th17 Cells metabolism
- Abstract
IL-17A and IL-17F share the highest sequence homology of the IL-17 family and signal via the same IL-17RA/RC receptor heterodimer. To better explore the expression of these two cytokines, we used a double reporter mouse strain (IL-17
DR mice), where IL-17A expressing cells are marked by enhanced green fluorescent protein (eGFP) while red fluorescence protein (RFP) reports the expression of IL-17F. In steady state, we found that Th17 and γδ T cells only expressed IL-17A, while IL-17F expression was restricted to CD8 T cells (Tc17) and innate lymphoid cells (ILC type 3) of the gut. In experimental autoimmune encephalomyelitis, the vast majority of CNS-infiltrating Th17 cells expressed IL-17A but not IL-17F. In contrast, anti-CD3-induced, TGF-β-driven Th17 cells in the gut expressed both of these IL-17 cytokines. In line with this, in vitro differentiation of Th17 cells in the presence of IL-1β led primarily to IL-17A expressing T cells, while TGF-β induced IL-17F co-expressing Th17 cells. Our results suggest that expression of IL-17F is associated with non-pathogenic T cells, pointing to a differential function of IL-17A versus IL-17F., Key Messages: Naïve mice: CD4+ T cells and γδ T cells express IL-17A, and Tc17 cells express IL-17F. Gut ILC3 show differential expression of IL17A and F. Th17 differentiation with TGF-β1 induces IL-17A and F, whereas IL-1β induced cells expressing IL-17A. Th17 cells in EAE in CNS express IL-17A only. Gut Th17 cells induced by anti-CD3 express IL-17A and F together as skin γδ T cells of IMQ-treated mice.- Published
- 2018
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19. The actin remodeling protein cofilin is crucial for thymic αβ but not γδ T-cell development.
- Author
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Seeland I, Xiong Y, Orlik C, Deibel D, Prokosch S, Küblbeck G, Jahraus B, De Stefano D, Moos S, Kurschus FC, Arnold B, and Samstag Y
- Subjects
- Actin Depolymerizing Factors chemistry, Animals, Cell Movement, Gene Knock-In Techniques, Humans, Jurkat Cells, Mice, Mutation genetics, Proline metabolism, T-Lymphocyte Subsets metabolism, T-Lymphocytes metabolism, Thymocytes metabolism, Actin Depolymerizing Factors metabolism, Actins metabolism, Receptors, Antigen, T-Cell, alpha-beta metabolism, Receptors, Antigen, T-Cell, gamma-delta metabolism, Thymus Gland metabolism
- Abstract
Cofilin is an essential actin remodeling protein promoting depolymerization and severing of actin filaments. To address the relevance of cofilin for the development and function of T cells in vivo, we generated knock-in mice in which T-cell-specific nonfunctional (nf) cofilin was expressed instead of wild-type (WT) cofilin. Nf cofilin mice lacked peripheral αβ T cells and showed a severe thymus atrophy. This was caused by an early developmental arrest of thymocytes at the double negative (DN) stage. Importantly, even though DN thymocytes expressed the TCRβ chain intracellularly, they completely lacked TCRβ surface expression. In contrast, nf cofilin mice possessed normal numbers of γδ T cells. Their functionality was confirmed in the γδ T-cell-driven, imiquimod (IMQ)-induced, psoriasis-like murine model. Overall, this study not only highlights the importance of cofilin for early αβ T-cell development but also shows for the first time that an actin-binding protein is differentially involved in αβ versus γδ T-cell development., Competing Interests: The authors have declared that no competing interests exist.
- Published
- 2018
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20. Regulation of IL-22BP in psoriasis.
- Author
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Voglis S, Moos S, Kloos L, Wanke F, Zayoud M, Pelczar P, Giannou AD, Pezer S, Albers M, Luessi F, Huber S, Schäkel K, and Kurschus FC
- Subjects
- Animals, Cells, Cultured, Dendritic Cells immunology, Dendritic Cells pathology, Dinoprostone analysis, Dinoprostone immunology, Disease Models, Animal, Female, Humans, Mice, Mice, Inbred BALB C, Psoriasis pathology, Receptors, Interleukin analysis, Skin immunology, Skin pathology, Psoriasis immunology, Receptors, Interleukin immunology
- Abstract
IL-22 is a potent pro-inflammatory cytokine upregulated in psoriasis and in other inflammatory diseases. The function of IL-22 is regulated by the soluble scavenging receptor, IL-22 binding protein (IL-22BP or IL-22RA2). However, the role and regulation of IL-22BP itself in the pathogenesis of inflammatory disease remain unclear. We used the TLR7 agonist Imiquimod (IMQ) to induce a psoriasis-like skin disease in mice and found a strong downregulation of IL-22BP in the affected skin as well as in the lymph nodes of animals treated with IMQ. We also analysed psoriatic skin of patients and compared this to skin of healthy donors. Interestingly, IL-22BP expression was similarly downregulated in skin biopsies of psoriasis patients compared to the skin of healthy donors. Since IL-22BP is expressed foremost in dendritic cells, we characterized its expression in monocyte-derived dendritic cells (MoDC) during maturation. In this way, we found Prostaglandin E2 (PGE
2 ) to be a potent suppressor of IL-22BP expression in vitro. We conclude that regulation of IL-22BP by inflammatory mediators is an important step for the progression of inflammation in the skin and possibly also in other autoimmune diseases.- Published
- 2018
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21. IL-17 for therapy.
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Kurschus FC and Moos S
- Subjects
- Animals, Antibodies, Monoclonal therapeutic use, Antibodies, Monoclonal, Humanized therapeutic use, Dermatitis pathology, Disease Models, Animal, Humans, Interleukin-17 genetics, Interleukin-17 metabolism, Mice, Mice, Transgenic, Molecular Targeted Therapy methods, Psoriasis immunology, Signal Transduction immunology, Skin immunology, Skin pathology, Spondylitis, Ankylosing drug therapy, Dermatitis immunology, Interleukin-17 antagonists & inhibitors, Interleukin-17 immunology, Psoriasis drug therapy
- Abstract
The cytokine IL-17 is now a target for an array of therapeutic monoclonal antibodies supposed to treat a variety of inflammatory diseases. The forerunner Secukinumab, an IL-17A neutralizing antibody, is meanwhile approved as first-line treatments for moderate-to-severe plaque psoriasis, and as second-line treatment for psoriatic arthritis and ankylosing spondylitis. Ixekizumab and Brodalumab, both also targeting the IL-17 pathway, were also recently approved by the FDA for plaque psoriasis. Using mice overexpressing IL-17A in a tissue of choice, we showed that the ectopic expression of this cytokine in keratinocytes resulted in a spontaneous and very strong form of psoriasis-like dermatitis. Interestingly, this model showed some typical comorbidities found in humans with psoriasis. In this review, we will discuss why IL-17 is a good target especially in psoriasis and what we learned from mouse models about its functions in pathological situations., (Copyright © 2017 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.)
- Published
- 2017
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22. Single-cell profiling reveals GPCR heterogeneity and functional patterning during neuroinflammation.
- Author
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Tischner D, Grimm M, Kaur H, Staudenraus D, Carvalho J, Looso M, Günther S, Wanke F, Moos S, Siller N, Breuer J, Schwab N, Zipp F, Waisman A, Kurschus FC, Offermanns S, and Wettschureck N
- Abstract
GPCR expression was intensively studied in bulk cDNA of leukocyte populations, but limited data are available with respect to expression in individual cells. Here, we show a microfluidic-based single-cell GPCR expression analysis in primary T cells, myeloid cells, and endothelial cells under naive conditions and during experimental autoimmune encephalomyelitis, the mouse model of multiple sclerosis. We found that neuroinflammation induces characteristic changes in GPCR heterogeneity and patterning, and we identify various functionally relevant subgroups with specific GPCR profiles among spinal cord-infiltrating CD4 T cells, macrophages, microglia, or endothelial cells. Using GPCRs CXCR4, S1P1, and LPHN2 as examples, we show how this information can be used to develop new strategies for the functional modulation of Th17 cells and activated endothelial cells. Taken together, single-cell GPCR expression analysis identifies functionally relevant subpopulations with specific GPCR repertoires and provides a basis for the development of new therapeutic strategies in immune disorders.
- Published
- 2017
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23. NG2 plays a role in neuroinflammation but is not expressed by immune cells.
- Author
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Kitic M, Karram K, Israel N, Yogev N, Lacher SM, Tang Y, Yigit H, Bauer J, Wanke F, Knezovic A, Trotter J, Kurschus FC, and Waisman A
- Subjects
- Animals, Antigens genetics, Antigens, CD metabolism, Bacterial Proteins genetics, Bacterial Proteins metabolism, Central Nervous System metabolism, Central Nervous System pathology, Disease Models, Animal, Luminescent Proteins genetics, Luminescent Proteins metabolism, Mice, Mice, Transgenic, Myelin-Oligodendrocyte Glycoprotein immunology, Myelin-Oligodendrocyte Glycoprotein toxicity, Peptide Fragments immunology, Peptide Fragments toxicity, Pertussis Toxin toxicity, Proteoglycans genetics, Receptor, Platelet-Derived Growth Factor beta metabolism, Encephalomyelitis, Autoimmune, Experimental genetics, Encephalomyelitis, Autoimmune, Experimental immunology, Encephalomyelitis, Autoimmune, Experimental pathology, Proteoglycans deficiency, Th1 Cells pathology, Th17 Cells pathology
- Published
- 2017
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24. TGF-β inhibitor Smad7 regulates dendritic cell-induced autoimmunity.
- Author
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Lukas D, Yogev N, Kel JM, Regen T, Mufazalov IA, Tang Y, Wanke F, Reizis B, Müller W, Kurschus FC, Prinz M, Kleiter I, Clausen BE, and Waisman A
- Subjects
- Animals, Basic-Leucine Zipper Transcription Factors metabolism, CD8-Positive T-Lymphocytes metabolism, Cell Differentiation physiology, Cytokines metabolism, Encephalomyelitis, Autoimmune, Experimental metabolism, Immune Tolerance, Indoleamine-Pyrrole 2,3,-Dioxygenase metabolism, Interferon Regulatory Factors metabolism, Mice, Mice, Inbred C57BL, Signal Transduction physiology, Spleen metabolism, T-Lymphocytes, Regulatory metabolism, Autoimmunity physiology, Dendritic Cells metabolism, Smad7 Protein metabolism, Transforming Growth Factor beta metabolism
- Abstract
TGF-β is an anti-inflammatory cytokine whose signaling is negatively controlled by Smad7. Previously, we established a role for Smad7 in the generation of autoreactive T cells; however, the function of Smad7 in dendritic cells (DCs) remains elusive. Here, we demonstrate that DC-specific Smad7 deficiency resulted in elevated expression of the transcription factors Batf3 and IRF8, leading to increased frequencies of CD8
+ CD103+ DCs in the spleen. Furthermore, Smad7-deficient DCs expressed higher levels of indoleamine 2,3-dioxygenase (IDO), an enzyme associated with tolerance induction. Mice devoid of Smad7 specifically in DCs are resistant to the development of experimental autoimmune encephalomyelitis (EAE) as a result of an increase of protective regulatory T cells (Tregs) and reduction of encephalitogenic effector T cells in the central nervous system. In agreement, inhibition of IDO activity or depletion of Tregs restored disease susceptibility. Intriguingly, when Smad7-deficient DCs also lacked the IFN-γ receptor, the mice regained susceptibility to EAE, demonstrating that IFN-γ signaling in DCs mediates their tolerogenic function. Our data indicate that Smad7 expression governs splenic DC subset differentiation and is critical for the promotion of their efficient function in immunity.- Published
- 2017
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25. EBI2 Is Highly Expressed in Multiple Sclerosis Lesions and Promotes Early CNS Migration of Encephalitogenic CD4 T Cells.
- Author
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Wanke F, Moos S, Croxford AL, Heinen AP, Gräf S, Kalt B, Tischner D, Zhang J, Christen I, Bruttger J, Yogev N, Tang Y, Zayoud M, Israel N, Karram K, Reißig S, Lacher SM, Reichhold C, Mufazalov IA, Ben-Nun A, Kuhlmann T, Wettschureck N, Sailer AW, Rajewsky K, Casola S, Waisman A, and Kurschus FC
- Subjects
- Animals, Autoimmunity physiology, Central Nervous System physiology, Cytochrome P450 Family 7 metabolism, Disease Models, Animal, Encephalomyelitis, Autoimmune, Experimental pathology, Female, Interleukin-1beta metabolism, Interleukin-23 metabolism, Male, Mice, Mice, Inbred C57BL, Steroid Hydroxylases metabolism, Th17 Cells metabolism, Th17 Cells physiology, CD4-Positive T-Lymphocytes metabolism, CD4-Positive T-Lymphocytes physiology, Cell Movement physiology, Central Nervous System metabolism, Encephalomyelitis, Autoimmune, Experimental metabolism, Multiple Sclerosis metabolism, Receptors, G-Protein-Coupled metabolism
- Abstract
Arrival of encephalitogenic T cells at inflammatory foci represents a critical step in development of experimental autoimmune encephalomyelitis (EAE), the animal model for multiple sclerosis. EBI2 and its ligand, 7α,25-OHC, direct immune cell localization in secondary lymphoid organs. CH25H and CYP7B1 hydroxylate cholesterol to 7α,25-OHC. During EAE, we found increased expression of CH25H by microglia and CYP7B1 by CNS-infiltrating immune cells elevating the ligand concentration in the CNS. Two critical pro-inflammatory cytokines, interleukin-23 (IL-23) and interleukin-1 beta (IL-1β), maintained expression of EBI2 in differentiating Th17 cells. In line with this, EBI2 enhanced early migration of encephalitogenic T cells into the CNS in a transfer EAE model. Nonetheless, EBI2 was dispensable in active EAE. Human Th17 cells do also express EBI2, and EBI2 expressing cells are abundant within multiple sclerosis (MS) white matter lesions. These findings implicate EBI2 as a mediator of CNS autoimmunity and describe mechanistically its contribution to the migration of autoreactive T cells into inflamed organs., (Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.)
- Published
- 2017
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26. IL-1 signaling is critical for expansion but not generation of autoreactive GM-CSF+ Th17 cells.
- Author
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Mufazalov IA, Schelmbauer C, Regen T, Kuschmann J, Wanke F, Gabriel LA, Hauptmann J, Müller W, Pinteaux E, Kurschus FC, and Waisman A
- Subjects
- Animals, Mice, Pertussis Toxin administration & dosage, Pertussis Toxin toxicity, Cell Proliferation, Encephalomyelitis, Autoimmune, Experimental pathology, Granulocyte-Macrophage Colony-Stimulating Factor analysis, Interleukin-1 metabolism, Th17 Cells chemistry, Th17 Cells physiology
- Abstract
Interleukin-1 (IL-1) is implicated in numerous pathologies, including multiple sclerosis and its animal model experimental autoimmune encephalomyelitis (EAE). However, the exact mechanism by which IL-1 is involved in the generation of pathogenic T cells and in disease development remains largely unknown. We found that following EAE induction, pertussis toxin administration leads to IL-1 receptor type 1 (IL-1R1)-dependent IL-1β expression by myeloid cells in the draining lymph nodes. This myeloid-derived IL-1β did not vitally contribute to the generation and plasticity of Th17 cells, but rather promoted the expansion of a GM-CSF
+ Th17 cell subset, thereby enhancing its encephalitogenic potential. Lack of expansion of GM-CSF-producing Th17 cells led to ameliorated disease in mice deficient for IL-1R1 specifically in T cells. Importantly, pathogenicity of IL-1R1-deficient T cells was fully restored by IL-23 polarization and expansion in vitro Therefore, our data demonstrate that IL-1 functions as a mitogenic mediator of encephalitogenic Th17 cells rather than qualitative inducer of their generation., (© 2016 The Authors.)- Published
- 2017
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27. T cell mediated pathogenesis in EAE: Molecular mechanisms.
- Author
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Kurschus FC
- Subjects
- Animals, Cytokines immunology, Cytokines metabolism, Humans, B-Lymphocytes immunology, Central Nervous System immunology, Encephalomyelitis, Autoimmune, Experimental immunology, Multiple Sclerosis immunology, T-Lymphocytes immunology
- Abstract
T cells are major initiators and mediators of disease in multiple sclerosis (MS) and in its animal model experimental autoimmune encephalomyelitis (EAE). EAE is an antigen-driven autoimmune model in which immunization against myelin autoantigens elicits strong T cell responses which initiate its pathology with CNS myelin destruction. T cells cause pathogenic events by several mechanisms; some work in a direct fashion in the CNS, such as direct cytokine-induced damage, granzyme-mediated killing, or glutamate-induced neurotoxicity, whereas most are indirect mechanisms, such as activation of other cell types like macrophages, B cells, or neutrophils. This review aims to describe and discuss the molecular effector mechanism by which T cells harm the CNS during EAE.
- Published
- 2015
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28. Improved method to retain cytosolic reporter protein fluorescence while staining for nuclear proteins.
- Author
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Heinen AP, Wanke F, Moos S, Attig S, Luche H, Pal PP, Budisa N, Fehling HJ, Waisman A, and Kurschus FC
- Subjects
- Animals, Cytoplasm metabolism, Fixatives, Fluorescent Dyes, Forkhead Transcription Factors, Mice, Mice, Inbred C57BL, Mice, Transgenic, Mycobacterium tuberculosis immunology, Nuclear Receptor Subfamily 1, Group F, Member 3, Staining and Labeling, T-Box Domain Proteins, T-Lymphocytes cytology, Tissue Fixation methods, Cytokines analysis, Flow Cytometry methods, Nuclear Proteins analysis, Transcription Factors analysis
- Abstract
Staining of transcription factors (TFs) together with retention of fluorescent reporter proteins is hindered by loss of fluorescence using current available methods. In this study, it is shown that current TF staining protocols do not destroy fluorescent proteins (FPs) but rather that fixation is not sufficient to retain FPs in the cytosol of the permeabilized cells. In this article, a simple and reliable protocol is elaborated, which allows efficient TF and cytokine staining while retaining FPs inside fixed cells., (© 2014 International Society for Advancement of Cytometry.)
- Published
- 2014
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29. IL-6 regulates neutrophil microabscess formation in IL-17A-driven psoriasiform lesions.
- Author
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Croxford AL, Karbach S, Kurschus FC, Wörtge S, Nikolaev A, Yogev N, Klebow S, Schüler R, Reissig S, Piotrowski C, Brylla E, Bechmann I, Scheller J, Rose-John S, Thomas Wunderlich F, Münzel T, von Stebut E, and Waisman A
- Subjects
- Abscess drug therapy, Abscess pathology, Animals, Disease Models, Animal, Epidermis immunology, Epidermis metabolism, Epidermis pathology, Gene Expression immunology, Granulocytes immunology, Granulocytes pathology, Interleukin-17 genetics, Interleukin-17 metabolism, Interleukin-6 metabolism, Interleukin-6 pharmacology, Macrophages immunology, Macrophages pathology, Mice, Mice, Knockout, Neutrophils metabolism, Psoriasis drug therapy, Psoriasis pathology, Receptors, Interleukin-6 genetics, Receptors, Interleukin-6 immunology, Signal Transduction drug effects, Signal Transduction immunology, T-Lymphocytes immunology, T-Lymphocytes pathology, Abscess immunology, Interleukin-17 immunology, Interleukin-6 immunology, Neutrophils immunology, Psoriasis immunology
- Abstract
The lack of a generally accepted animal model for human psoriasis has hindered progress with respect to understanding the pathogenesis of the disease. Here we present a model in which transgenic IL-17A expression is targeted to the skin in mice, achievable after crossing our IL-17A(ind) allele to the K14-Cre strain. K14-IL-17A(ind/+) mice invariably develop an overt skin inflammation bearing many hallmark characteristics of human psoriasis including dermal infiltration of effector T cells, formation of neutrophil microabscesses, and hyperkeratosis. IL-17A expression in the skin results in upregulated granulopoiesis and migration of IL-6R-expressing neutrophils into the skin. Neutralization of IL-6 signaling efficiently reduces the observed pathogenesis in skin of IL-17A-overexpressing mice, with marked reductions in epidermal neutrophil abscess formation and epidermal thickening. Thus, IL-6 functions downstream of IL-17A to exacerbate neutrophil microabscess development in psoriasiform lesions.
- Published
- 2014
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30. Inflammatory demyelination induces glia alterations and ganglion cell loss in the retina of an experimental autoimmune encephalomyelitis model.
- Author
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Horstmann L, Schmid H, Heinen AP, Kurschus FC, Dick HB, and Joachim SC
- Subjects
- Animals, Apoptosis immunology, Demyelinating Diseases etiology, Demyelinating Diseases immunology, Demyelinating Diseases metabolism, Encephalomyelitis, Autoimmune, Experimental complications, Encephalomyelitis, Autoimmune, Experimental metabolism, Immunohistochemistry, Inflammation metabolism, Male, Mice, Mice, Inbred C57BL, Neuroglia metabolism, Optic Neuritis etiology, Optic Neuritis metabolism, Optic Neuritis pathology, Retinal Ganglion Cells metabolism, Encephalomyelitis, Autoimmune, Experimental pathology, Inflammation pathology, Neuroglia pathology, Retinal Ganglion Cells pathology
- Abstract
Background: Multiple sclerosis (MS) is often accompanied by optic nerve inflammation. And some patients experience permanent vision loss. We examined if the grade of optic nerve infiltration and demyelination affects the severity of clinical signs in an experimental autoimmune encephalomyelitis (EAE) model. The loss of retinal ganglion cells (RGC) and alterations in glia activity were also investigated., Methods: C57BL/6 mice were immunized with peptide MOG35-55 in complete Freund's adjuvant (CFA) and controls received PBS in CFA. Then 23 days post immunization eyes were prepared for flatmounts and stained with Nissl to evaluated neuronal density. Clinical EAE symptoms as well as cell infiltration and demyelination in the optic nerve were examined. Retinal sections were stained with hematoxylin and eosin and silver stain. Immunohistochemistry was used to label RGCs (Brn-3a), apoptotic cells (caspase 3), macroglia (glial fibrillary acidic protein (GFAP)), microglia (Iba1), macrophages (F 4/80) and interleukin-6 (IL-6) secretion., Results: EAE symptoms started at day 8 and peaked at day 15. Cell infiltrations (P = 0.0047) and demyelination (P = 0.0018) of EAE nerves correlated with the clinical score (r > 0.8). EAE led to a significant loss of RGCs (P< 0.0001). Significantly more caspase 3+ cells were noted in these animals (P = 0.0222). They showed an increased expression of GFAP (P< 0.0002) and a higher number of microglial cells (P< 0.0001). Also more macrophages and IL-6 secretion were observed in EAE mice., Conclusions: MOG immunization leads to optic neuritis and RGC loss. EAE severity is related to the severity of optic nerve inflammation and demyelination. EAE not only affects activation of apoptotic signals, but also causes a glial response in the retina.
- Published
- 2013
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31. Subclinical CNS inflammation as response to a myelin antigen in humanized mice.
- Author
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Zayoud M, El Malki K, Frauenknecht K, Trinschek B, Kloos L, Karram K, Wanke F, Georgescu J, Hartwig UF, Sommer C, Jonuleit H, Waisman A, and Kurschus FC
- Subjects
- Animals, Female, Humans, Inflammation immunology, Inflammation metabolism, Inflammation pathology, Male, Mice, Mice, Inbred NOD, Mice, SCID, T-Lymphocytes immunology, T-Lymphocytes pathology, Antigens, CD toxicity, Encephalomyelitis, Autoimmune, Experimental immunology, Encephalomyelitis, Autoimmune, Experimental pathology, Myelin-Oligodendrocyte Glycoprotein toxicity
- Abstract
Multiple sclerosis is a demyelinating autoimmune disease of the CNS. Its animal model experimental autoimmune encephalomyelitis is commonly induced by active immunization with myelin antigens. To investigate human immune responses against myelin antigens in vivo we established a new subclinical experimental autoimmune encephalomyelitis model in humanized mice. NOD/Scidγc⁻/⁻ animals were transferred with peripheral blood mononuclear cells from healthy human donors and immunized with myelin antigens in complete Freund's adjuvant and antigen-pulsed autologous dendritic cells. Human T cells recovered from these animals reacted specifically to the soluble domain of myelin oligodendrocyte glycoprotein and secreted proinflammatory cytokines. Furthermore, immunized animals developed subclinical CNS inflammation with infiltrating CD4⁺ and CD8⁺ T cells and production of encephalitogenic cytokines. Thus, this model of myelin-induced CNS inflammation by human T cells may allow testing of new human-specific therapeuticals for multiple sclerosis.
- Published
- 2013
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32. An alternative pathway of imiquimod-induced psoriasis-like skin inflammation in the absence of interleukin-17 receptor a signaling.
- Author
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El Malki K, Karbach SH, Huppert J, Zayoud M, Reissig S, Schüler R, Nikolaev A, Karram K, Münzel T, Kuhlmann CR, Luhmann HJ, von Stebut E, Wörtge S, Kurschus FC, and Waisman A
- Subjects
- Adjuvants, Immunologic pharmacology, Animals, Disease Models, Animal, Female, Imiquimod, Interleukin-17 metabolism, Interleukin-6 immunology, Interleukin-6 metabolism, Interleukins immunology, Interleukins metabolism, Macrophages drug effects, Macrophages immunology, Mice, Mice, Knockout, Neutrophil Infiltration drug effects, Neutrophil Infiltration immunology, Psoriasis genetics, Receptors, Interleukin-17 genetics, Receptors, Interleukin-17 metabolism, Signal Transduction drug effects, Skin immunology, Skin metabolism, Skin pathology, Interleukin-22, Aminoquinolines pharmacology, Interleukin-17 immunology, Psoriasis chemically induced, Psoriasis immunology, Receptors, Interleukin-17 immunology, Signal Transduction immunology
- Abstract
Topical application of imiquimod (IMQ) on the skin of mice induces inflammation with common features found in psoriatic skin. Recently, it was postulated that IL-17 has an important role both in psoriasis and in the IMQ model. To further investigate the impact of IL-17RA signaling in psoriasis, we generated IL-17 receptor A (IL-17RA)-deficient mice (IL-17RA(del)) and challenged these mice with IMQ. Interestingly, the disease was only partially reduced and delayed but not abolished when compared with controls. In the absence of IL-17RA, we found persisting signs of inflammation such as neutrophil and macrophage infiltration within the skin. Surprisingly, already in the naive state, the skin of IL-17RA(del) mice contained significantly elevated numbers of Th17- and IL-17-producing γδ T cells, assuming that IL-17RA signaling regulates the population size of Th17 and γδ T cells. Upon IMQ treatment of IL-17RA(del) mice, these cells secreted elevated amounts of tumor necrosis factor-α, IL-6, and IL-22, accompanied by increased levels of the chemokine CXCL2, suggesting an alternative pathway of neutrophil and macrophage skin infiltration. Hence, our findings have major implications in the potential long-term treatment of psoriasis by IL-17-targeting drugs.
- Published
- 2013
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33. TLR-4 ligation of dendritic cells is sufficient to drive pathogenic T cell function in experimental autoimmune encephalomyelitis.
- Author
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Mellanby RJ, Cambrook H, Turner DG, O'Connor RA, Leech MD, Kurschus FC, MacDonald AS, Arnold B, and Anderton SM
- Subjects
- Animals, Antigen-Presenting Cells, Cell Differentiation drug effects, Cytokines metabolism, Dendritic Cells drug effects, Disease Models, Animal, Encephalomyelitis, Autoimmune, Experimental chemically induced, Freund's Adjuvant adverse effects, Histocompatibility Antigens Class II metabolism, Ligation, Lipopolysaccharides toxicity, Lymphocyte Activation drug effects, Mice, Myelin Basic Protein metabolism, Myelin Basic Protein pharmacology, Peptide Fragments pharmacology, T-Lymphocytes drug effects, T-Lymphocytes metabolism, T-Lymphocytes pathology, Toll-Like Receptor 4 immunology, Dendritic Cells metabolism, Encephalomyelitis, Autoimmune, Experimental pathology, T-Lymphocytes immunology, Toll-Like Receptor 4 metabolism
- Abstract
Background: Experimental autoimmune encephalomyelitis (EAE) depends on the initial activation of CD4(+) T cells responsive to myelin autoantigens. The key antigen presenting cell (APC) population that drives the activation of naïve T cells most efficiently is the dendritic cell (DC). As such, we should be able to trigger EAE by transfer of DC that can present the relevant autoantigen(s). Despite some sporadic reports, however, models of DC-driven EAE have not been widely adopted. We sought to test the feasibility of this approach and whether activation of the DC by toll-like receptor (TLR)-4 ligation was a sufficient stimulus to drive EAE., Findings: Host mice were seeded with myelin basic protein (MBP)-reactive CD4+ T cells and then were injected with DC that could present the relevant MBP peptide which had been exposed to lipopolysaccharide as a TLR-4 agonist. We found that this approach induced robust clinical signs of EAE., Conclusions: DC are sufficient as APC to effectively drive the differentiation of naïve myelin-responsive T cells into autoaggressive effector T cells. TLR-4-stimulation can activate the DC sufficiently to deliver the signals required to drive the pathogenic function of the T cell. These models will allow the dissection of the molecular requirements of the initial DC-T cell interaction in the lymphoid organs that ultimately leads to autoimmune pathology in the central nervous system.
- Published
- 2012
- Full Text
- View/download PDF
34. Dendritic cells ameliorate autoimmunity in the CNS by controlling the homeostasis of PD-1 receptor(+) regulatory T cells.
- Author
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Yogev N, Frommer F, Lukas D, Kautz-Neu K, Karram K, Ielo D, von Stebut E, Probst HC, van den Broek M, Riethmacher D, Birnberg T, Blank T, Reizis B, Korn T, Wiendl H, Jung S, Prinz M, Kurschus FC, and Waisman A
- Subjects
- Animals, Antigen Presentation immunology, Autoantigens immunology, Autoimmunity immunology, B7-H1 Antigen immunology, B7-H1 Antigen metabolism, CD11c Antigen, Dendritic Cells metabolism, Mice, Mice, Inbred C57BL, Mice, Knockout, Programmed Cell Death 1 Receptor metabolism, T-Lymphocytes, Regulatory metabolism, Th1 Cells immunology, Th1 Cells metabolism, Th17 Cells immunology, Th17 Cells metabolism, Dendritic Cells immunology, Encephalomyelitis, Autoimmune, Experimental immunology, Immune Tolerance immunology, Programmed Cell Death 1 Receptor immunology, T-Lymphocytes, Regulatory immunology
- Abstract
Mature dendritic cells (DCs) are established as unrivaled antigen-presenting cells (APCs) in the initiation of immune responses, whereas steady-state DCs induce peripheral T cell tolerance. Using various genetic approaches, we depleted CD11c(+) DCs in mice and induced autoimmune CNS inflammation. Unexpectedly, mice lacking DCs developed aggravated disease compared to control mice. Furthermore, when we engineered DCs to present a CNS-associated autoantigen in an induced manner, we found robust tolerance that prevented disease, which coincided with an upregulation of the PD-1 receptor on antigen-specific T cells. Additionally, we showed that PD-1 was necessary for DC-mediated induction of regulatory T cells. Our results show that a reduction of DCs interferes with tolerance, resulting in a stronger inflammatory response, and that other APC populations could compensate for the loss of immunogenic APC function in DC-depleted mice., (Copyright © 2012 Elsevier Inc. All rights reserved.)
- Published
- 2012
- Full Text
- View/download PDF
35. Mouse models for multiple sclerosis: historical facts and future implications.
- Author
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Croxford AL, Kurschus FC, and Waisman A
- Subjects
- Animals, Autoantigens history, Autoantigens immunology, Disease Models, Animal, Encephalomyelitis, Autoimmune, Experimental etiology, Encephalomyelitis, Autoimmune, Experimental genetics, Encephalomyelitis, Autoimmune, Experimental immunology, Gene Targeting history, History, 20th Century, History, 21st Century, Humans, Mice, Multiple Sclerosis immunology, Multiple Sclerosis therapy, Th17 Cells immunology, Encephalomyelitis, Autoimmune, Experimental history, Multiple Sclerosis etiology
- Abstract
Multiple sclerosis (MS) is an inflammatory and demyelinating condition of the CNS, characterized by perivascular infiltrates composed largely of T lymphocytes and macrophages. Although the precise cause remains unknown, numerous avenues of research support the hypothesis that autoimmune mechanisms play a major role in the development of the disease. Pathologically similar lesions to those seen in MS can be induced in laboratory rodents by immunization with CNS-derived antigens. This form of disease induction, broadly termed experimental autoimmune encephalomyelitis, is frequently the starting point in MS research with respect to studying pathogenesis and creating novel treatments. Many different EAE models are available, each mimicking a particular facet of MS. These models all have common ancestry, and have developed from a single concept of immunization with self-antigen. We will discuss the major changes in immunology research, which have shaped the EAE models we use today, and discuss how current animal models of MS have resulted in successful treatments and more open questions for researchers to address., (2010 Elsevier B.V. All rights reserved.)
- Published
- 2011
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36. Modeling a complex disease: multiple sclerosis.
- Author
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Kurschus FC, Wörtge S, and Waisman A
- Subjects
- Animals, Encephalomyelitis, Autoimmune, Experimental immunology, Encephalomyelitis, Autoimmune, Experimental pathology, Encephalomyelitis, Autoimmune, Experimental physiopathology, Humans, Mice, Mice, Inbred C57BL, Nerve Degeneration immunology, Nerve Degeneration pathology, Nerve Degeneration physiopathology, Rats, Rats, Inbred Lew, Disease Models, Animal, Multiple Sclerosis immunology, Multiple Sclerosis pathology, Multiple Sclerosis physiopathology
- Abstract
The recent decades have shown that multiple sclerosis (MS) is not a uniform disease entity with common etiology, but rather a disease or syndrome characterized by a heterogeneous pattern of manifestations and pathological principles. Apart from the older distinctions of the Devic's disease from the standard Western form of relapsing remitting MS or the more Asian form of opticospinal MS, specific pathological patterns indicating distinct etiologies have been established by analyses of biopsies and autopsies. Further, the distinct responses of patients to drugs targeting either specific cell types or immunoregulatory mechanisms such as Rituximab or IFNβ clearly demonstrate the heterogeneity of the disease and their driving mechanisms. Finally, the late neurodegenerative phase, which severe cases of MS patients experience, is now in the focus of research. Here, a mechanism independent of or with low participation of the adaptive immune system takes place, which is therefore not treatable by current immunotargeting drugs. In this review, we will summarize previous and latest efforts to establish new mouse models mirroring these distinct disease patterns and pathways., (Copyright © 2011 Elsevier Inc. All rights reserved.)
- Published
- 2011
- Full Text
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37. Genetic proof for the transient nature of the Th17 phenotype.
- Author
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Kurschus FC, Croxford AL, Heinen AP, Wörtge S, Ielo D, and Waisman A
- Subjects
- Adoptive Transfer, Animals, Cell Differentiation immunology, Gene Expression Regulation genetics, Genes, RAG-1 genetics, Genes, Reporter genetics, Integrases genetics, Interferon-gamma metabolism, Lymphocyte Activation genetics, Lymphocyte Activation immunology, Mice, Mice, Inbred C57BL, Mice, Knockout, Mice, Transgenic, Receptors, Antigen, T-Cell genetics, T-Lymphocyte Subsets immunology, T-Lymphocyte Subsets pathology, Th1 Cells immunology, Th1 Cells pathology, Th17 Cells immunology, Th17 Cells pathology, Encephalomyelitis, Autoimmune, Experimental immunology, Gene Expression Regulation immunology, T-Lymphocyte Subsets metabolism, Th1 Cells metabolism, Th17 Cells metabolism
- Abstract
IL-17-producing CD4(+) T cells (Th17) have been classified as a new T helper cell subset. Using an IL-17 fate mapping mouse strain, which genetically fixes the memory of IL-17 expression, we demonstrate that IL-17A/F-expressing T helper cells generated either in vitro or in vivo are not a stable T-cell subset. Upon adoptive transfer of IL-17F-reporter-positive Th17 cells to RAG-deficient or WT animals, encephalitogenic Th17 cells partially lose IL-17 expression and upregulate IFN-γ. Additionally, we show that Th1 cells can convert in vivo to IL-17A/IFN-γ-coexpressing cells in the mesenteric lymph nodes (mLN). Our data classify IL-17A and IL-17F as cytokines produced transiently in response to the local microenvironment, thus showing that IL-17 expression does not define an end-stage T helper cell subset.
- Published
- 2010
- Full Text
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38. Delivery and therapeutic potential of human granzyme B.
- Author
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Kurschus FC and Jenne DE
- Subjects
- Antineoplastic Agents chemistry, Antineoplastic Agents metabolism, Genetic Therapy, Granzymes antagonists & inhibitors, Granzymes chemistry, Granzymes genetics, Granzymes metabolism, Humans, Immunotoxins chemistry, Immunotoxins genetics, Immunotoxins metabolism, Models, Molecular, Perforin metabolism, Protein Conformation, Protein Transport, Recombinant Fusion Proteins therapeutic use, Serpins metabolism, Antineoplastic Agents therapeutic use, Granzymes therapeutic use, Immunotoxins therapeutic use, Killer Cells, Natural enzymology, T-Lymphocytes, Cytotoxic enzymology
- Abstract
Granzyme B (GzmB) is used by cytotoxic lymphocytes as a molecular weapon for the defense against virus-infected and malignantly transformed host cells. It belongs to a family of small serine proteases that are stored in secretory vesicles of killer cells. After secretion of these cytolytic granules during killer cell attack, GzmB is translocated into the cytosol of target cells with the help of the pore-forming protein perforin. GzmB has adopted similar protease specificity as caspase-8, and once delivered, it activates major executioner apoptosis pathways. Since GzmB is very effective in killing human tumor cell lines that are otherwise resistant against many cytotoxic drugs and since GzmB of human origin can be recombinantly expressed, its use as part of a 'magic bullet' in tumor therapy is a very tempting idea. In this review, we emphasize the peculiar characteristics of GzmB that make it suited for use as an effector domain in potential immunoconjugates. We discuss what is known about its uptake into target cells and the trials performed with GzmB-armed immunoconjugates, and we assess the prospects of its potential therapeutic value.
- Published
- 2010
- Full Text
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39. Gold fluorescent annexin A5 as a novel apoptosis detection tool.
- Author
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Kurschus FC, Pal PP, Bäumler P, Jenne DE, Wiltschi B, and Budisa N
- Subjects
- Annexin A5 genetics, Annexin A5 metabolism, Biomarkers analysis, Biomarkers metabolism, Cell Line, Tumor, Fluorescent Antibody Technique, Fluorescent Dyes metabolism, Gold, Humans, Microscopy, Confocal, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Annexin A5 analysis, Apoptosis, Flow Cytometry, Fluorescent Dyes analysis, Recombinant Fusion Proteins analysis
- Abstract
We describe a golden fluorescent apoptosis detection tool, which we generated by a fusion of golden fluorescent protein (GdFP) with human annexin A5 (anxA5). GdFP was obtained by replacement of tryptophan at position 66 with 4-aminotryptophan in the chromophore of enhanced cyan fluorescent protein. The GdFP-anxA5 construct combines highly desirable features originating from both fusion partners. These include (i) strong binding to membrane phosphatidylserine patches of apoptotic cells in the presence of Ca(2+) which is brought about by anxA5, (ii) the stable and homogeneous monomeric state, (iii) as well as the red-shifted fluorescence maximum at 574 nm originating from GdFP. We found that GdFP-anxA5 is equally well applicable for apoptosis studies as a routinely used fluorescein 5'-isothiocyanate-annexin A5 conjugate. Golden fluorescent annexin A5 represents a new, stable, and homogeneous red-shifted optical probe for the efficient detection of apoptosis by fluorescence microscopy or by flow cytometry., (2009 International Society for Advancement of Cytometry.)
- Published
- 2009
- Full Text
- View/download PDF
40. Spontaneous relapsing-remitting EAE in the SJL/J mouse: MOG-reactive transgenic T cells recruit endogenous MOG-specific B cells.
- Author
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Pöllinger B, Krishnamoorthy G, Berer K, Lassmann H, Bösl MR, Dunn R, Domingues HS, Holz A, Kurschus FC, and Wekerle H
- Subjects
- Animals, Autoantibodies immunology, Autoantigens immunology, B-Lymphocytes cytology, Brain metabolism, Brain pathology, Complement Activation, Disease Models, Animal, Encephalomyelitis, Autoimmune, Experimental pathology, Encephalomyelitis, Autoimmune, Experimental physiopathology, Female, Humans, Immunoglobulins immunology, Interferons immunology, Interleukins immunology, Male, Mice, Multiple Sclerosis immunology, Multiple Sclerosis pathology, Myelin Proteins, Myelin-Associated Glycoprotein genetics, Myelin-Oligodendrocyte Glycoprotein, Peptide Fragments genetics, Spinal Cord metabolism, Spinal Cord pathology, T-Lymphocytes cytology, B-Lymphocytes immunology, Encephalomyelitis, Autoimmune, Experimental immunology, Mice, Transgenic, Myelin-Associated Glycoprotein immunology, Peptide Fragments immunology, Receptors, Antigen, T-Cell genetics, Receptors, Antigen, T-Cell immunology, T-Lymphocytes immunology
- Abstract
We describe new T cell receptor (TCR) transgenic mice (relapsing-remitting [RR] mice) carrying a TCR specific for myelin oligodendrocyte glycoprotein (MOG) peptide 92-106 in the context of I-A(s). Backcrossed to the SJL/J background, most RR mice spontaneously develop RR experimental autoimmune encephalomyelitis (EAE) with episodes often altering between different central nervous system tissues like the cerebellum, optic nerve, and spinal cord. Development of spontaneous EAE depends on the presence of an intact B cell compartment and on the expression of MOG autoantigen. There is no spontaneous EAE development in B cell-depleted mice or in transgenic mice lacking MOG. Transgenic T cells seem to expand MOG autoreactive B cells from the endogenous repertoire. The expanded autoreactive B cells produce autoantibodies binding to a conformational epitope on the native MOG protein while ignoring the T cell target peptide. The secreted autoantibodies are pathogenic, enhancing demyelinating EAE episodes. RR mice constitute the first spontaneous animal model for the most common form of multiple sclerosis (MS), RR MS.
- Published
- 2009
- Full Text
- View/download PDF
41. Myelin-specific T cells also recognize neuronal autoantigen in a transgenic mouse model of multiple sclerosis.
- Author
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Krishnamoorthy G, Saxena A, Mars LT, Domingues HS, Mentele R, Ben-Nun A, Lassmann H, Dornmair K, Kurschus FC, Liblau RS, and Wekerle H
- Subjects
- Amino Acid Sequence, Animals, Cross Reactions, Disease Models, Animal, Encephalomyelitis, Autoimmune, Experimental genetics, Encephalomyelitis, Autoimmune, Experimental immunology, Encephalomyelitis, Autoimmune, Experimental metabolism, Encephalomyelitis, Autoimmune, Experimental pathology, Mice, Mice, Inbred C57BL, Mice, Transgenic, Molecular Sequence Data, Multiple Sclerosis genetics, Multiple Sclerosis metabolism, Multiple Sclerosis pathology, Myelin Proteins, Myelin-Associated Glycoprotein chemistry, Myelin-Associated Glycoprotein deficiency, Myelin-Associated Glycoprotein genetics, Myelin-Associated Glycoprotein metabolism, Myelin-Oligodendrocyte Glycoprotein, Neurofilament Proteins immunology, Receptors, Antigen, T-Cell genetics, Receptors, Antigen, T-Cell immunology, Receptors, Antigen, T-Cell metabolism, Autoantigens immunology, Multiple Sclerosis immunology, Myelin Sheath immunology, T-Lymphocytes immunology
- Abstract
We describe here the paradoxical development of spontaneous experimental autoimmune encephalomyelitis (EAE) in transgenic mice expressing a myelin oligodendrocyte glycoprotein (MOG)-specific T cell antigen receptor (TCR) in the absence of MOG. We report that in Mog-deficient mice (Mog-/-), the autoimmune response by transgenic T cells is redirected to a neuronal cytoskeletal self antigen, neurofilament-M (NF-M). Although components of radically different protein classes, the cross-reacting major histocompatibility complex I-Ab-restricted epitope sequences of MOG35-55 and NF-M18-30 share essential TCR contact positions. This pattern of cross-reaction is not specific to the transgenic TCR but is also commonly seen in MOG35-55-I-Ab-reactive T cells. We propose that in the C57BL/6 mouse, MOG and NF-M response components add up to overcome the general resistance of this strain to experimental induction of autoimmunity. Similar cumulative responses against more than one autoantigen may have a role in spontaneously developing human autoimmune diseases.
- Published
- 2009
- Full Text
- View/download PDF
42. Cutting edge: an IL-17F-CreEYFP reporter mouse allows fate mapping of Th17 cells.
- Author
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Croxford AL, Kurschus FC, and Waisman A
- Subjects
- Adoptive Transfer, Animals, CD8-Positive T-Lymphocytes cytology, CD8-Positive T-Lymphocytes immunology, CD8-Positive T-Lymphocytes pathology, Cells, Cultured, Encephalomyelitis, Autoimmune, Experimental immunology, Encephalomyelitis, Autoimmune, Experimental metabolism, Encephalomyelitis, Autoimmune, Experimental pathology, Gene Expression Regulation immunology, Humans, Immunophenotyping, Interleukin-17 biosynthesis, Interleukin-17 physiology, Mice, Mice, Inbred C57BL, Mice, Inbred DBA, Mice, Transgenic, Proteins genetics, RNA, Untranslated, T-Lymphocytes, Regulatory pathology, T-Lymphocytes, Regulatory transplantation, Bacterial Proteins genetics, Cell Differentiation genetics, Cell Differentiation immunology, Genes, Reporter immunology, Integrases genetics, Interleukin-17 genetics, Luminescent Proteins genetics, T-Lymphocytes, Regulatory immunology, T-Lymphocytes, Regulatory metabolism
- Abstract
The need for reporter lines able to faithfully track Th17 cells in vivo has become an issue of exceptional importance. To address this, we generated a mouse strain in which Cre recombinase is expressed from the IL-17F promoter. Crossing the IL-17F-Cre allele to a conditional enhanced yellow fluorescent protein (EYFP) reporter mouse yielded the IL-17F-Cre(EYFP) strain, in which IL-17F expression is twinned with EYFP in live IL-17F-expressing cells. Although we demonstrate that IL-17F expression is restricted to CD4(+) T cells during experimental autoimmune encephalomyelitis, IL-17F-Cre(EYFP) CD8 T cells robustly expressed IL-17F in response to TGF-beta, IL-6, and IL-23. Fate mapping of IL-17F-expressing reporter T cells revealed a significant down-regulation of Th17 cytokines after homeostatic expansion in RAG1-deficient animals. Despite this loss of effector phenotype, committed Th17 cells were resistant to Foxp3 expression in vitro or in vivo. Thus, the IL-17F-Cre strain furthers our understanding of Th17 biology.
- Published
- 2009
- Full Text
- View/download PDF
43. Granzyme B delivery via perforin is restricted by size, but not by heparan sulfate-dependent endocytosis.
- Author
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Kurschus FC, Fellows E, Stegmann E, and Jenne DE
- Subjects
- Amino Acid Sequence, Apoptosis drug effects, Cell Line, Granzymes chemistry, Granzymes genetics, Heparitin Sulfate pharmacology, Humans, Killer Cells, Natural enzymology, Molecular Weight, Mutation genetics, Protein Binding, Protein Transport, Endocytosis drug effects, Granzymes metabolism, Perforin metabolism
- Abstract
How granzymes gain entry into the cytosol of target cells during killer cell attack has been the subject of several studies in the past, but the effective delivery mechanism during target cell encounter has not been clarified. Here we show that granzyme B (GzmB) mutants lacking binding to negatively charged, essentially heparan-sulfate-containing membrane receptors are poorly endocytosed yet are delivered to the cytosol with efficacy similar to that of WT GzmB. In a cell-based system GzmB-deficient natural killer cells provided perforin (pfn) by natural polarized secretion and synergized with externally added GzmB. Whereas receptor (heparan sulfate)-dependent endocytosis was dispensable, delivery of larger cargo like that of GzmB fusion proteins and GzmB-antibody complexes was restricted by their size. Our data support the model in which granzymes are primarily translocated through repairable membrane pores of finite size and not by the disruption of endocytosed vesicles. We conclude that structurally related translocators, i.e., perforin and cholesterol-dependent cytolysins, deliver deathly cargo across host cell membranes in a similar manner.
- Published
- 2008
- Full Text
- View/download PDF
44. Natural killer cell-derived human granzyme H induces an alternative, caspase-independent cell-death program.
- Author
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Fellows E, Gil-Parrado S, Jenne DE, and Kurschus FC
- Subjects
- Animals, Cell Death drug effects, Cytochromes c metabolism, Granzymes pharmacology, HL-60 Cells, Humans, Immunity, Innate, K562 Cells, Killer Cells, Natural cytology, Killer Cells, Natural immunology, Mitochondria enzymology, Rats, Reactive Oxygen Species metabolism, Caspases metabolism, Cell Death physiology, Granzymes physiology, Killer Cells, Natural physiology
- Abstract
Granzyme H (GzmH) belongs to a family of 5 human serine proteases that are expressed by cytotoxic immune effector cells. Although GzmH is most closely related to the caspase-activating granzyme B (GzmB), neither a natural substrate nor a role in immune defense reactions has been demonstrated for this orphan granzyme. In rodents, multiple related genes exist, but none of these can be regarded as functional homologs. Here we show that host cells are efficiently killed by GzmH after perforin and streptolysin O-mediated delivery into the cytosol. Dying cells show typical hallmarks of programmed cell death, such as mitochondrial depolarization, reactive oxygen species (ROS) generation, DNA degradation, and chromatin condensation. Contrary to GzmB, cell death by GzmH does not involve the activation of executioner caspases, the cleavage of Bid or inhibitor of caspase-activated DNase (ICAD), or the release of cytochrome c. The high expression levels of GzmH in naive natural killer (NK) cells and its potent killing ability strongly support the role of the protease in triggering an alternative cell-death pathway in innate immunity.
- Published
- 2007
- Full Text
- View/download PDF
45. Experimental autoimmune encephalomyelitis in mice expressing the autoantigen MBP 1-10 covalently bound to the MHC class II molecule I-Au.
- Author
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Kurschus FC, Oelert T, Liliensiek B, Buchmann P, Wraith DC, Hämmerling GJ, and Arnold B
- Subjects
- Animals, Autoantigens genetics, Autoimmunity genetics, Autoimmunity immunology, CD4-Positive T-Lymphocytes immunology, Crosses, Genetic, Encephalomyelitis, Autoimmune, Experimental genetics, Gene Expression genetics, Gene Expression immunology, Histocompatibility Antigens Class II genetics, Immune Tolerance genetics, Immune Tolerance immunology, Lymphocyte Activation genetics, Lymphocyte Activation immunology, Mice, Mice, Transgenic, Multiple Sclerosis genetics, Myelin Basic Protein genetics, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins immunology, Autoantigens immunology, Encephalomyelitis, Autoimmune, Experimental immunology, Histocompatibility Antigens Class II immunology, Multiple Sclerosis immunology, Myelin Basic Protein immunology
- Abstract
Most autoantigens implicated in multiple sclerosis (MS) are expressed not only in the central nervous system (CNS) but also in the thymus and the periphery. Nevertheless, these autoantigens might induce a strong autoimmune response leading to severe destruction within the CNS. To investigate the influence of a dominantly presented autoantigen on experimental autoimmune encephalomyelitis (EAE), we generated transgenic mice expressing the autoantigenic peptide MBP 1-10 covalently bound to the MHC class II molecule I-Au. These mice were crossed either with B10.PL or with TCR-transgenic Tg4 mice, specific for the transgenic peptide-MHC combination. In double transgenic mice we found strong thymic deletion and residual peripheral T cells were refractory to antigen stimulation in vitro. Residual peripheral CD4+ T cells expressed activation markers and a high proportion was CD25 positive. Transfer of both CD25-negative and CD25-positive CD4+ T cells from double transgenic animals into B10.PL mice strongly inhibited the progression of EAE. Despite this thorough tolerance induction, some double transgenic mice developed severe signs of EAE after an extended period of time. Our data show that in the circumstances where autoantigenic priming persists, and where the number of antigen-specific T cells is high enough, autoimmunity may prevail over very potent tolerance-inducing mechanisms.
- Published
- 2006
- Full Text
- View/download PDF
46. Membrane receptors are not required to deliver granzyme B during killer cell attack.
- Author
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Kurschus FC, Bruno R, Fellows E, Falk CS, and Jenne DE
- Subjects
- Binding Sites, Cell Line, Cell Membrane Permeability, Granzymes, Heparin analysis, Heparin metabolism, Humans, Killer Cells, Natural metabolism, Lysosomes metabolism, Proteoglycans analysis, Proteoglycans metabolism, Apoptosis, Heparin analogs & derivatives, Killer Cells, Natural physiology, Receptors, Cell Surface physiology, Serine Endopeptidases metabolism
- Abstract
Granzyme B (GzmB), a serine protease of cytotoxic T lymphocytes and natural killer (NK) cells, induces apoptosis by caspase activation after crossing the plasma membrane of target cells. The mechanism of this translocation during killer cell attack, however, is not understood. Killer cells release GzmB and the membrane-disturbing perforin at the contact site after target recognition. Receptor-mediated import of glycosylated GzmB and release from endosomes were suggested, but the role of the cation-independent mannose 6-phosphate receptor was recently refuted. Using recombinant nonglycosylated GzmB, we observed binding of GzmB to cellular membranes in a cell type-dependent manner. The basis and functional impact of surface binding were clarified. GzmB binding was correlated with the surface density of heparan sulfate chains, was eliminated on treatment of target cells with heparinase III or sodium chlorate, and was completely blocked by an excess of catalytically inactive GzmB or GzmK. Although heparan sulfate-bound GzmB was taken up rapidly into intracellular lysosomal compartments, neither of the treatments had an inhibitory influence on apoptosis induced by externally added streptolysin O and GzmB or by natural killer cells. We conclude that membrane receptors for GzmB on target cells are not crucial for killer cell-mediated apoptosis.
- Published
- 2005
- Full Text
- View/download PDF
47. Receptor for advanced glycation end products (RAGE) regulates sepsis but not the adaptive immune response.
- Author
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Liliensiek B, Weigand MA, Bierhaus A, Nicklas W, Kasper M, Hofer S, Plachky J, Gröne HJ, Kurschus FC, Schmidt AM, Yan SD, Martin E, Schleicher E, Stern DM, Hämmerling G Gü, Nawroth PP, and Arnold B
- Subjects
- Animals, Cecum injuries, Immune System immunology, Mice, Mice, Knockout, Peritonitis metabolism, Receptor for Advanced Glycation End Products, Receptors, Immunologic genetics, Receptors, Immunologic immunology, Shock, Septic metabolism, Time Factors, Immune System metabolism, Receptors, Immunologic metabolism, Sepsis metabolism
- Abstract
While the initiation of the adaptive and innate immune response is well understood, less is known about cellular mechanisms propagating inflammation. The receptor for advanced glycation end products (RAGE), a transmembrane receptor of the immunoglobulin superfamily, leads to perpetuated cell activation. Using novel animal models with defective or tissue-specific RAGE expression, we show that in these animal models RAGE does not play a role in the adaptive immune response. However, deletion of RAGE provides protection from the lethal effects of septic shock caused by cecal ligation and puncture. Such protection is reversed by reconstitution of RAGE in endothelial and hematopoietic cells. These results indicate that the innate immune response is controlled by pattern-recognition receptors not only at the initiating steps but also at the phase of perpetuation.
- Published
- 2004
- Full Text
- View/download PDF
48. Killing of target cells by redirected granzyme B in the absence of perforin.
- Author
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Kurschus FC, Kleinschmidt M, Fellows E, Dornmair K, Rudolph R, Lilie H, and Jenne DE
- Subjects
- Animals, Apoptosis physiology, Bacterial Toxins immunology, Bacterial Toxins metabolism, Cell Line, Granzymes, Humans, Lewis Blood Group Antigens genetics, Lewis Blood Group Antigens immunology, Lewis Blood Group Antigens metabolism, Peptide Fragments immunology, Peptide Fragments metabolism, Perforin, Pore Forming Cytotoxic Proteins, Receptors, Cell Surface metabolism, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins immunology, Recombinant Fusion Proteins metabolism, Serine Endopeptidases genetics, Serine Endopeptidases immunology, T-Lymphocytes, Cytotoxic immunology, T-Lymphocytes, Cytotoxic metabolism, Membrane Glycoproteins metabolism, Serine Endopeptidases metabolism
- Abstract
Granzyme B (GzmB) is a potent apoptosis-inducing serine protease of cytotoxic lymphocytes. Following receptor-mediated endocytosis, GzmB is supposed to enter the cytosol through perforin-mediated membrane disruption. We investigated whether retargeting of GzmB to Lewis Y positive surface receptors could lead to perforin-independent target cell death. We coupled recombinant GzmB to the Lewis Y-binding antibody dsFv-B3. Targeting of GzmB to Lewis Y positive cells triggered cell death with similar efficacy as dsFv-B3 targeted Pseudomonas exotoxin fragment 38 (PE38). Since GzmB was only weakly inhibited by plasma proteins, GzmB-based immunoconjugates should be useful as a new class of immunotoxins with low immunogenicity utilizing programmed cell death for therapeutic purposes.
- Published
- 2004
- Full Text
- View/download PDF
49. Crystal structure of the apoptosis-inducing human granzyme A dimer.
- Author
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Hink-Schauer C, Estébanez-Perpiñá E, Kurschus FC, Bode W, and Jenne DE
- Subjects
- Amino Acid Sequence, Crystallography, X-Ray, Dimerization, Granzymes, Humans, Hydrolysis, Models, Molecular, Molecular Sequence Data, Molecular Weight, Protein Conformation, Recombinant Proteins chemistry, Recombinant Proteins metabolism, Sequence Homology, Amino Acid, Serine Endopeptidases metabolism, Serine Endopeptidases physiology, Substrate Specificity, Apoptosis physiology, Serine Endopeptidases chemistry
- Abstract
Granzyme A (GzmA) belongs to a family of trypsin-like serine proteases localized in cytoplasmic granules of activated lymphocytes and natural killer (NK) cells. In contrast to the related granzyme B (GzmB), GzmA forms a stable disulfide-linked homodimer and triggers target-cell death in a caspase-independent way. Limited proteolysis of a high-molecular-mass complex containing SET (also named putative HLA-associated protein II or PHAPII), PHAPI (pp32, leucine-rich acidic nuclear protein) and HMG2 by GzmA liberates NM23-H1, a Mg2+-dependent DNase that causes single-stranded breaks in nuclear DNA. By analyzing the dimeric GzmA structure at a resolution of 2.5 A, we determined the substrate-binding constraints and selective advantages of the two domains arranged as a unique functional tandem. The active sites of the two subunits point in opposite directions and the nearby noncatalytic surfaces can function as exosites, presenting substrates to the active site region of the adjacent partner in a manner analogous to staphylokinase or streptokinase, which present plasminogen to the cofactor-plasmin and cofactor-plasminogen complexes.
- Published
- 2003
- Full Text
- View/download PDF
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