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Characterization of tetraspanin-2 expressing cells in the mouse
Abstracts
mice, trying to establish a functional correlation between MG activation and neuronal damage. Finally, we attempted to polarize MG cells in Sod1G93A mice toward the M2 paradigm by using a gene therapy approach based on the administration of lentiviruses encoding IL4. The over expression of IL4 successfully triggered the polarization of microglia from an M1 proinflammatory state to an M2 antinflammatory phenotype. This condition mediates positively effects in term of survival and progression of the pathology, thus indicating that a modulation of neuroinflammation could be beneficial for ALS treatment. doi:10.1016/j.jneuroim.2014.08.458
26 The NKG2D signaling pathway is a possible therapeutic target for autoimmune myositis Tobias Ruck, Ali Maisam Afzali, Stefan Bittner, Sven G. Meuth, Heinz Wiendl Department of Neurology, University of Münster, Münster, Germany Abstract Muscle cells actively participate in immune reactions through the expression of cytokines, chemokines, major histocompatibility complex (MHC) and costimulatory molecules. NKG2D is a costimulatory receptor on NK and T cells, which has been previously shown to play an important role in a number of immune-mediated circumstances such as autoimmune diseases or anti-tumor responses. We here aim to investigate a putative role of this pathway in autoimmune myositis. NKG2D ligands were expressed on primary human myoblasts and were found upregulated upon stimulation. In parallel, production of shedded MICA (sMICA) decreased upon stimulation leading to diminished inhibition of NKG2D signaling. In an in vitro killing system, cultivation of pre-stimulated CD8 cells together with myoblast resulted in a dosedependent lysis of myoblasts which was significantly inhibited by a NKG2D-blocking antibody. However, no significant changes were found in the peripheral blood of patients with polymyositis compared to healthy donors concerning levels of sMICA or expression of NKG2D on different immune cell subtypes. In contrast, histological stainings showed an expression of the NKG2D ligand MICA/B in muscle biopsy specimens of polymyositis patients but not in healthy controls supporting the involvement of this pathway in autoimmune myositis potentially opening new therapeutic avenues. doi:10.1016/j.jneuroim.2014.08.459
171
Hospital Hygiene, University of Düsseldorf, Düsseldorf, Germany; fInstitute of Molecular Psychiatry, University of Bonn, Bonn, Germany; gInstitute of Neuropathology, University Hospital Münster, Münster, Germany; h Department of Immunology and Environment, Life & Medical Sciences Institute (LIMES), University of Bonn, Bonn, Germany; iInstitute of Infection Immunology, TWINCORE, Centre for Experimental and Clinical Infection Research, Hannover, Germany; jCells-in-Motion Cluster of Excellence (EXC 1003-CiM), University of Münster, Germany Tetraspanins (Tspans) are a family of transmembrane proteins that modulate immune cell responses and the migration and differentiation of CNS cells by complex formation with neighboring cell surface proteins. A newly described tetraspanin-2 (Tspan-2) in the mouse was suggested to regulate astrocyte and microglia activation in the CNS. However, neither its expression pattern nor its functional role in immune responses has been studied up to now. Here we defined the expression pattern of Tspan-2 by transcriptional analyses and employed a newly generated BAC transgenic mouse model that expresses a diphtheria toxin receptor (DTR)-EGFP fusion protein under the gene regulatory elements of the murine Tspan-2 locus (Tspan-2/DTR/EGFP). Tspan-2 mRNA was identified in GMCSF-differentiated bone-marrow dendritic cells (BM-DCs), lymphoid tissues and at high levels in the brain. Flow cytometric analysis of Tspan-2/DTR/EGFP mice revealed EGFP/Tspan-2 expression in around 8% of all spleen cells, and less than 3% of all lymph node cells. The major fraction of EGFP/Tspan-2 expressing cells in spleen and lymph nodes comprised a subset of classical CD11c+ dendritic cells positive for MHC II and 33D1. In addition, a minor subset of Ly6Ghi neutrophils and CD4+ T cells expressed EGFP/Tspan-2 in the spleen. Upon TLR ligand stimulation expression of EGFP/Tspan-2 was no longer detectable in BM-DCs suggesting that its expression is regulated by pathogen sensing. In the CNS of Tspan-2/DTR/EGFP mice EGFP/Tspan-2 localized to mature MBP+ and CNP+ oligodendrocytes. Expression of Tspan-2 in dendritic cells and oligodendrocytes, two central cellular mediators of inflammatory CNS responses, supports its potential significance for neuroinflammatory disorders. doi:10.1016/j.jneuroim.2014.08.460
405 RGC-32 regulates TGF-beta-induced extracellular matrix expression in reactive astrocytes Cosmin Teglaa, Cornelia Cudricia, Dallas Boodhooa, Alvaro Martina, Adam Sugarmana, Jacob Danoffa, Violeta Rusb, Horea Rusa a
452 Characterization of tetraspanin-2 expressing cells in the mouse Christina Rulanda, Julia Patzigb, Katharina Lahlc, Veronika Lukacs-Kornekd, Arezoo Fattahi-Mehra, Stefanie Scheue, Andreas Zimmerf, Tanja Kuhlmanng, Hauke Wernerb, Irmgard Försterh, Tim Sparwasseri, Judith Alferinka,j a
Department of Psychiatry, University Hospital Münster, Münster, Germany; Department of Neurogenetics, Max Planck Institute of Experimental Medicine, Göttingen, Germany; cLaboratory of Immunology and Vascular Biology, Department of Pathology, Stanford University School of Medicine, Stanford, CA, United States; dDepartment of Medicine II, Saarland University Medical Center, Homburg, Germany; eInstitute of Medical Microbiology and b
University of Maryland, School of Medicine, Department of Neurology, Baltimore, United States; bUniversity of Maryland, School of Medicine, Department of Internal Medicine, Baltimore, United States Excessive extracellular matrix (ECM) deposition in active demyelinating multiple sclerosis (MS) lesions may impede axonal regeneration and can modify immune reactions. RGC-32 plays an important role in the mediation of TGF-beta downstream effects but its role in gliosis has not been investigated. Our objective was to investigate the role of RGC-32 in the mediation of TGF-beta effects on astrocytes ECM production and expression of reactive markers. We examined first the expression of collagen types I to V, decorin and RGC-32 in MS brains by immunohistochemistry and correlate its expression with that of astrocytes. To gain more insight into the role played by RGC-32 in gliosis we then examined the role of RGC-32 in the mediation of ECM and reactive astrocyte markers' expression in cultured astrocytes. To silence RGC-32 expression, astrocytes were
mice, trying to establish a functional correlation between MG activation and neuronal damage. Finally, we attempted to polarize MG cells in Sod1G93A mice toward the M2 paradigm by using a gene therapy approach based on the administration of lentiviruses encoding IL4. The over expression of IL4 successfully triggered the polarization of microglia from an M1 proinflammatory state to an M2 antinflammatory phenotype. This condition mediates positively effects in term of survival and progression of the pathology, thus indicating that a modulation of neuroinflammation could be beneficial for ALS treatment. doi:10.1016/j.jneuroim.2014.08.458
26 The NKG2D signaling pathway is a possible therapeutic target for autoimmune myositis Tobias Ruck, Ali Maisam Afzali, Stefan Bittner, Sven G. Meuth, Heinz Wiendl Department of Neurology, University of Münster, Münster, Germany Abstract Muscle cells actively participate in immune reactions through the expression of cytokines, chemokines, major histocompatibility complex (MHC) and costimulatory molecules. NKG2D is a costimulatory receptor on NK and T cells, which has been previously shown to play an important role in a number of immune-mediated circumstances such as autoimmune diseases or anti-tumor responses. We here aim to investigate a putative role of this pathway in autoimmune myositis. NKG2D ligands were expressed on primary human myoblasts and were found upregulated upon stimulation. In parallel, production of shedded MICA (sMICA) decreased upon stimulation leading to diminished inhibition of NKG2D signaling. In an in vitro killing system, cultivation of pre-stimulated CD8 cells together with myoblast resulted in a dosedependent lysis of myoblasts which was significantly inhibited by a NKG2D-blocking antibody. However, no significant changes were found in the peripheral blood of patients with polymyositis compared to healthy donors concerning levels of sMICA or expression of NKG2D on different immune cell subtypes. In contrast, histological stainings showed an expression of the NKG2D ligand MICA/B in muscle biopsy specimens of polymyositis patients but not in healthy controls supporting the involvement of this pathway in autoimmune myositis potentially opening new therapeutic avenues. doi:10.1016/j.jneuroim.2014.08.459
171
Hospital Hygiene, University of Düsseldorf, Düsseldorf, Germany; fInstitute of Molecular Psychiatry, University of Bonn, Bonn, Germany; gInstitute of Neuropathology, University Hospital Münster, Münster, Germany; h Department of Immunology and Environment, Life & Medical Sciences Institute (LIMES), University of Bonn, Bonn, Germany; iInstitute of Infection Immunology, TWINCORE, Centre for Experimental and Clinical Infection Research, Hannover, Germany; jCells-in-Motion Cluster of Excellence (EXC 1003-CiM), University of Münster, Germany Tetraspanins (Tspans) are a family of transmembrane proteins that modulate immune cell responses and the migration and differentiation of CNS cells by complex formation with neighboring cell surface proteins. A newly described tetraspanin-2 (Tspan-2) in the mouse was suggested to regulate astrocyte and microglia activation in the CNS. However, neither its expression pattern nor its functional role in immune responses has been studied up to now. Here we defined the expression pattern of Tspan-2 by transcriptional analyses and employed a newly generated BAC transgenic mouse model that expresses a diphtheria toxin receptor (DTR)-EGFP fusion protein under the gene regulatory elements of the murine Tspan-2 locus (Tspan-2/DTR/EGFP). Tspan-2 mRNA was identified in GMCSF-differentiated bone-marrow dendritic cells (BM-DCs), lymphoid tissues and at high levels in the brain. Flow cytometric analysis of Tspan-2/DTR/EGFP mice revealed EGFP/Tspan-2 expression in around 8% of all spleen cells, and less than 3% of all lymph node cells. The major fraction of EGFP/Tspan-2 expressing cells in spleen and lymph nodes comprised a subset of classical CD11c+ dendritic cells positive for MHC II and 33D1. In addition, a minor subset of Ly6Ghi neutrophils and CD4+ T cells expressed EGFP/Tspan-2 in the spleen. Upon TLR ligand stimulation expression of EGFP/Tspan-2 was no longer detectable in BM-DCs suggesting that its expression is regulated by pathogen sensing. In the CNS of Tspan-2/DTR/EGFP mice EGFP/Tspan-2 localized to mature MBP+ and CNP+ oligodendrocytes. Expression of Tspan-2 in dendritic cells and oligodendrocytes, two central cellular mediators of inflammatory CNS responses, supports its potential significance for neuroinflammatory disorders. doi:10.1016/j.jneuroim.2014.08.460
405 RGC-32 regulates TGF-beta-induced extracellular matrix expression in reactive astrocytes Cosmin Teglaa, Cornelia Cudricia, Dallas Boodhooa, Alvaro Martina, Adam Sugarmana, Jacob Danoffa, Violeta Rusb, Horea Rusa a
452 Characterization of tetraspanin-2 expressing cells in the mouse Christina Rulanda, Julia Patzigb, Katharina Lahlc, Veronika Lukacs-Kornekd, Arezoo Fattahi-Mehra, Stefanie Scheue, Andreas Zimmerf, Tanja Kuhlmanng, Hauke Wernerb, Irmgard Försterh, Tim Sparwasseri, Judith Alferinka,j a
Department of Psychiatry, University Hospital Münster, Münster, Germany; Department of Neurogenetics, Max Planck Institute of Experimental Medicine, Göttingen, Germany; cLaboratory of Immunology and Vascular Biology, Department of Pathology, Stanford University School of Medicine, Stanford, CA, United States; dDepartment of Medicine II, Saarland University Medical Center, Homburg, Germany; eInstitute of Medical Microbiology and b
University of Maryland, School of Medicine, Department of Neurology, Baltimore, United States; bUniversity of Maryland, School of Medicine, Department of Internal Medicine, Baltimore, United States Excessive extracellular matrix (ECM) deposition in active demyelinating multiple sclerosis (MS) lesions may impede axonal regeneration and can modify immune reactions. RGC-32 plays an important role in the mediation of TGF-beta downstream effects but its role in gliosis has not been investigated. Our objective was to investigate the role of RGC-32 in the mediation of TGF-beta effects on astrocytes ECM production and expression of reactive markers. We examined first the expression of collagen types I to V, decorin and RGC-32 in MS brains by immunohistochemistry and correlate its expression with that of astrocytes. To gain more insight into the role played by RGC-32 in gliosis we then examined the role of RGC-32 in the mediation of ECM and reactive astrocyte markers' expression in cultured astrocytes. To silence RGC-32 expression, astrocytes were