- Email: [email protected]
Critical role of thymocyte-derived brain-derived neurotrophic factor in T cell maturation
202
Abstracts
(SOCS3, STAT5) pathways. Gene targets within cell cycle regulation pathways (PTPN6, SLC3A2, VAV3, DDR1) and modulators of oxidative stress (MT1A, HMOX1, SLC30A1) were also significantly affected by the CBD treatment. The microarray results were confirmed using qPCR on selected gene targets. Immunoblotting demonstrated that CBD reduces IL-17 by decreasing STAT3 phosphorylation and increasing that of STAT5. In conclusion, our observations increase our understanding of the mechanisms of the anti-inflammatory activities of CBD. doi:10.1016/j.jneuroim.2014.08.541
329 The role of NFkappaB inducing kinase (NIK) in the pathogenicity of EAE Sonja Lachera, Florian Mairb, Nicole Israela, Sabrina Rodewalda, Burkhard Becherb, Ari Waismana a
Institute for Molecular Medicine, University Medical Center of the Johannes Gutenberg, University Mainz, Mainz, Germany; bInstitute of Experimental Immunology, University of Zurich, Zürich, Switzerland Question: NIK knockout mice have been shown to be resistant in the mouse model of multiple sclerosis, experimental autoimmune encephalomyelitis (EAE). As the knockout mice harbor various deficiencies in the immune cell compartment as well as in the structure of lymphoid tissue, it is not possible to use them to study the cell-specific role of NIK. Of special interest is the analysis of the function of NIK in T-cells, which are one of the main players in this disease. To overcome this issue, we generated a conditional NIKfl/fl mouse where we can analyze the role of NIK specifically in T-cells in the pathogenicity of EAE. Methods: We used a conditional NIKfl/fl mouse, which was crossed to the CD4-cre mouse line to specifically knock out NIK in T-cells. We induced the mice with EAE and monitored the course of disease with subsequent flow cytometric analysis and cell culturing of MOG responding cells. Results: Under steady state conditions the T-cell specific NIK knockout mice had a normal lymphoid organization as well as TH1/TH17 cell numbers. However, following EAE induction, we found that these mice were resistant to the disease. This was accompanied by a dramatic reduction of CNS-infiltrating immune cells. Furthermore, we found a decreased generation of MOG-responding T-cells in the periphery. In addition, these cells showed a decreased ability to secrete the effector cytokines IFN gamma and IL-17A. Conclusions: Our results suggest an indispensible role of NIK in T-cells for the induction of an experimental neurodegenerative disease. This might be due to the role of NIK in the generation of encephalitogenic MOG responding T-cells. doi:10.1016/j.jneuroim.2014.08.542
175 Chemokine-induced trafficking of T cells into the meninges through the brain "gates" Antoine Louveau, Sachin Gadani, Tajie Harris, Jonathan Kipnis Center for Brain Immunology and Glia, Department of Neuroscience, University of Virginia, Charlottesville, United States
After being perceived as an immunoprivileged site, it is now accepted that the CNS is under a constant immune surveillance that contributes to the maintenance and homeostasis of brain functions. Particularly, T lymphocytes have an important role in physiological brain function. Previous studies have enlightened the pro-cognitive role of IL-4 secreting CD4 T lymphocytes in the meninges. However, the path of T cell entry under physiological conditions and the signals that drive them remain poorly understood. In this study, we analyzed T cell localization within the meninges and addressed the anatomical and molecular mechanism of T cell entry into the meningeal spaces. Using confocal microscopy, we show that the vast majority of T cells reside in the dura matter. Interestingly, T cells were also found in other meningeal compartments, such as arachnoid, pia and choroid plexus, but at a much lower density. More importantly, after a cognitive task, we measured a specific increase of T cells density within the dura but not in the other areas. Quantitative PCR analysis of whole meninges and choroid plexus from naïve and cognitively challenged mice showed a specific increase of CCL5, CCL8, CXCL9 and VCAM-1 in the meninges after training, whereas ICAM-1, E-Selectin and P-Selectin are decreased in the choroid plexus of trained mice. Those results suggest that T cells enter the meninges through a chemokine-induced signal that most likely does not involve the choroid plexus. Overall, this study suggests that dura matter is a major site for T cell trafficking and function within the meninges. Understanding the physiological path of T cell entry into the meninges may shed a new light on the etiology of neuroinflammatory diseases, such as multiple sclerosis. doi:10.1016/j.jneuroim.2014.08.543
216 Critical role of thymocyte-derived brain-derived neurotrophic factor in T cell maturation Fred Lühdera, Ralf Linkerb, De-hyung Leeb, Holger Reichardtc, Ursula Bommhardtd, Ralf Golde, Alexander Flügela a
Department of Neuroimmunology, Institute for Multiple Sclerosis Research, University of Göttingen, Göttingen, Germany; bDepartment of Neurology, Friedrich-Alexander University Erlangen, Erlangen, Germany; cDepartment of Cellular and Molecular Immunology, University of Göttingen, GöttingenGermany; dInstitute for Molecular and Clinical Immunology, Otto-Guericke University Magdeburg, Magdeburg, Germany; eDepartment of Neurology, St. Josef-Hospital/Ruhr-University Bochum, Bochum, Germany Brain-derived neurotrophic factor (BDNF) is an essential neurotrophin for neuronal survival, axonal growth, generation of new synaptic connections, regulation of neuronal activity as well as synaptic and dendritic plasticity. Emerging evidence also indicates an essential role for BDNF outside the nervous system, for instance in immune cells. In this study we focussed on the effect of BDNF on T cells, using conventional and conditional BDNF knockout (ko) mice lacking BDNF specifically in this lymphoid subset. In both settings, diminished T cell cellularity in peripheral lymphoid organs was observed. In order to get further insight into this phenomenon, we analysed T cell development in the thymus and found diminished total thymocyte numbers accompanied by a significant increase in CD4/CD8 double negative (DN) thymocytes due to a partial block in the transition from the DN3 to the DN4 stage. As underlying molecular mechanisms we could rule out increased thymocyte apoptosis as well as defects in the expression of the TCR -chain or the pre-TCR. In contrast, pERK but not pAKT levels were reduced in DN3 BDNF-deficient thymocytes pointing towards a MAP kinase-based
Abstracts
signalling defect as cause for this developmental T cell retardation. BDNF deficiency did not substantially influence peripheral acute immune responses and homeostatic proliferation of peripheral T cells. Taken together, our data reveal a critical autocrine and/or paracrine role of T cell-derived BDNF in thymocyte maturation involving ERK-mediated TCR signalling pathways.
doi:10.1016/j.jneuroim.2014.08.544
471 Cognitive performance among older individuals associated with the peripheral effector memory CD4+ T cells Cláudia Miranda, Susana Roque, Nadine Santos, Carlos Portugal-nunes, Patrício Costa, Joana Palha, Nuno Sousa, Margarida Correia-neves Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Braga, Portugal Progressive decline in immune function (immunosenescence) and in cognitive performance are common alterations accompanying aging. The interaction between these two systems has been highlighted in recent years mainly in studies using animal models. It has been shown that mice lacking B and T lymphocytes display cognitive impairments that are reverted by T cell replenishment. In fact, aging is accompanied by defined alterations in the adaptive immune system, namely in the T cell compartments. Taking into consideration the link between T cells and cognition, and since both age-associated cognitive decline and immunosenescence vary greatly between individuals we hypothesized that alterations in lymphocytes and cognitive performance are associated. In a cohort of cognitively healthy agers (n = 114), characterized by diverse neurocognitive/psychological performance patterns, we observed that better cognitive performance is associated with lower numbers of effector memory CD4+ T cell, higher numbers of naive CD8+ T cell and higher numbers of B cells in the blood. Furthermore, effector memory CD4+ T cells are predictors of general and executive function and memory, even when factors known to influence cognitive performance in older individuals (e.g. age, gender, education and mood) are taken into account. This is the first study identifying a phenotype of the immune system associated with poorer cognitive performance in normal cognitive aging, which strongly supports the interplay between the immune system and cognitive behavior and open a new perspective for interventions aiming to delay cognitive decline through aging.
doi:10.1016/j.jneuroim.2014.08.545
203
circulate as soluble forms, thereby neutralizing IL-1 activity. Expression of IL-1 is associated with experimental autoimmune encephalomyelitis (EAE), an animal model of human multiple sclerosis, which is believed to be mediated by T cells. In fact, exogenous IL-1 administration exacerbated the disease, while IL-1 knock-out (KO) animals showed resistance in EAE. To better understand the role of IL-1RI and IL-1RII in T cell encephalitogenicity we used the Cre/loxP system to delete these receptors either specifically in conventional T cells or in all cells of the body. For that purpose we crossed mice carrying Il1r1 and Il1r2 conditional alleles to the CD4-Cre or CMV-Cre transgenic mouse strains, respectively. All four groups of KO animals showed normal ontogenesis in steadystate conditions and T cells lacking IL-1R expression were triggered under in vitro polarization conditions towards effector subsets such as Th1 by adding the cytokines IL-2 and IL-12, or towards Th17 in the presence of TGF-beta, IL-6 and IL-23. As expected, T cells with deleted Il1r1 gene were non-responsive to IL-1 beta administration in a cell proliferation assay. Interestingly, mice lacking the IL-1RI developed EAE after active myelin oligodendrocyte glycoprotein (MOG) peptide immunization, although with a milder severity compared to wild type controls. At the peak of disease CNS-infiltrating IL-1RI KO T cells were characterized by significantly lower expression of effector cytokines, such as IL-17A, IFN-gamma and GM-CSF. In contrast, IL-1RII full KO mice showed an increased EAE severity, accompanied by enhanced cytokine expression from T cells. However, IL-1RII T cell specific KO animals developed a normal EAE. In this work we provide the first results obtained from two novel models of conditional IL-1RI and IL-1RII deletion. These mutations lead to opposite effects pointing to the necessity of intact IL-1 signaling for in vivo T cell development and pathogenicity. Thereby, the exacerbated disease in IL-1RII full KO animals suggests a negative contribution of this decoy receptor in EAE development. However, withdrawal of the receptor only from T cells is not enough to influence the disease progression, pointing to a contribution of IL-1RII from non-T cell sources. doi:10.1016/j.jneuroim.2014.08.546
523 Abnormal responses of CD8 + CD161high mucosal associated invariant T (MAIT) cells and CCR2 + CCR5+ CD4 T cells contribute to disrupted balance of effector and regulatory T cells in pediatric-onset MS Mukanthu Nyirendaa, Rui Lia, Craig Moorea, Ayal Rozenberga, Ayman Rezka, Trina Johnsonb, Dessa Sadovnickc, Douglas Arnolda, Ruth Ann Married, Brenda Banwelle,f, Amit Bar-ora,b, on behalf of the Canadian Pediatric Demyelinating Disease Network a
363 IL-1 receptor type I and type II differentially regulate EAE pathogenesis Ilgiz Mufazalov, Carsten Schelmbauer, Janina Kuschmann, Tommy Regen, Christopher Hackenbruch, Ari Waisman
Neuroimmunology Unit, Montreal Neurological Institute, McGill University, Montreal, Canada; bExperimental Therapeutics Program, Montreal Neurological Institute, McGill University, Montreal, Canada; c Division of Neurology, Department of Medical Genetics, University of British Columbia, British Columbia, Canada; dDepartments of Internal Medicine and Community Health, University of Manitoba, Manitoba, Canada; eDepartment of Paediatrics, Hospital For Sick Chilgdren, Toronto, Canada; fDivision of Neurology, Children's Hospital of Philadelphia, Philadelphia, United States
Institue for Molecular Medicine, JGU Medical Center, Mainz, Germany Interleukin-1 (IL-1) can bind to two receptors termed IL-1RI and IL1RII, but only the type I receptor is capable to transduce the signal. In addition, both receptors can be cleaved from the cell surface and
Abstract We wished to examine a carefully selected panel of effector and regulatory T cell (Treg) subsets (previously implicated in studies of adult-onset multiple sclerosis (MS) or other autoimmune diseases), at
Abstracts
(SOCS3, STAT5) pathways. Gene targets within cell cycle regulation pathways (PTPN6, SLC3A2, VAV3, DDR1) and modulators of oxidative stress (MT1A, HMOX1, SLC30A1) were also significantly affected by the CBD treatment. The microarray results were confirmed using qPCR on selected gene targets. Immunoblotting demonstrated that CBD reduces IL-17 by decreasing STAT3 phosphorylation and increasing that of STAT5. In conclusion, our observations increase our understanding of the mechanisms of the anti-inflammatory activities of CBD. doi:10.1016/j.jneuroim.2014.08.541
329 The role of NFkappaB inducing kinase (NIK) in the pathogenicity of EAE Sonja Lachera, Florian Mairb, Nicole Israela, Sabrina Rodewalda, Burkhard Becherb, Ari Waismana a
Institute for Molecular Medicine, University Medical Center of the Johannes Gutenberg, University Mainz, Mainz, Germany; bInstitute of Experimental Immunology, University of Zurich, Zürich, Switzerland Question: NIK knockout mice have been shown to be resistant in the mouse model of multiple sclerosis, experimental autoimmune encephalomyelitis (EAE). As the knockout mice harbor various deficiencies in the immune cell compartment as well as in the structure of lymphoid tissue, it is not possible to use them to study the cell-specific role of NIK. Of special interest is the analysis of the function of NIK in T-cells, which are one of the main players in this disease. To overcome this issue, we generated a conditional NIKfl/fl mouse where we can analyze the role of NIK specifically in T-cells in the pathogenicity of EAE. Methods: We used a conditional NIKfl/fl mouse, which was crossed to the CD4-cre mouse line to specifically knock out NIK in T-cells. We induced the mice with EAE and monitored the course of disease with subsequent flow cytometric analysis and cell culturing of MOG responding cells. Results: Under steady state conditions the T-cell specific NIK knockout mice had a normal lymphoid organization as well as TH1/TH17 cell numbers. However, following EAE induction, we found that these mice were resistant to the disease. This was accompanied by a dramatic reduction of CNS-infiltrating immune cells. Furthermore, we found a decreased generation of MOG-responding T-cells in the periphery. In addition, these cells showed a decreased ability to secrete the effector cytokines IFN gamma and IL-17A. Conclusions: Our results suggest an indispensible role of NIK in T-cells for the induction of an experimental neurodegenerative disease. This might be due to the role of NIK in the generation of encephalitogenic MOG responding T-cells. doi:10.1016/j.jneuroim.2014.08.542
175 Chemokine-induced trafficking of T cells into the meninges through the brain "gates" Antoine Louveau, Sachin Gadani, Tajie Harris, Jonathan Kipnis Center for Brain Immunology and Glia, Department of Neuroscience, University of Virginia, Charlottesville, United States
After being perceived as an immunoprivileged site, it is now accepted that the CNS is under a constant immune surveillance that contributes to the maintenance and homeostasis of brain functions. Particularly, T lymphocytes have an important role in physiological brain function. Previous studies have enlightened the pro-cognitive role of IL-4 secreting CD4 T lymphocytes in the meninges. However, the path of T cell entry under physiological conditions and the signals that drive them remain poorly understood. In this study, we analyzed T cell localization within the meninges and addressed the anatomical and molecular mechanism of T cell entry into the meningeal spaces. Using confocal microscopy, we show that the vast majority of T cells reside in the dura matter. Interestingly, T cells were also found in other meningeal compartments, such as arachnoid, pia and choroid plexus, but at a much lower density. More importantly, after a cognitive task, we measured a specific increase of T cells density within the dura but not in the other areas. Quantitative PCR analysis of whole meninges and choroid plexus from naïve and cognitively challenged mice showed a specific increase of CCL5, CCL8, CXCL9 and VCAM-1 in the meninges after training, whereas ICAM-1, E-Selectin and P-Selectin are decreased in the choroid plexus of trained mice. Those results suggest that T cells enter the meninges through a chemokine-induced signal that most likely does not involve the choroid plexus. Overall, this study suggests that dura matter is a major site for T cell trafficking and function within the meninges. Understanding the physiological path of T cell entry into the meninges may shed a new light on the etiology of neuroinflammatory diseases, such as multiple sclerosis. doi:10.1016/j.jneuroim.2014.08.543
216 Critical role of thymocyte-derived brain-derived neurotrophic factor in T cell maturation Fred Lühdera, Ralf Linkerb, De-hyung Leeb, Holger Reichardtc, Ursula Bommhardtd, Ralf Golde, Alexander Flügela a
Department of Neuroimmunology, Institute for Multiple Sclerosis Research, University of Göttingen, Göttingen, Germany; bDepartment of Neurology, Friedrich-Alexander University Erlangen, Erlangen, Germany; cDepartment of Cellular and Molecular Immunology, University of Göttingen, GöttingenGermany; dInstitute for Molecular and Clinical Immunology, Otto-Guericke University Magdeburg, Magdeburg, Germany; eDepartment of Neurology, St. Josef-Hospital/Ruhr-University Bochum, Bochum, Germany Brain-derived neurotrophic factor (BDNF) is an essential neurotrophin for neuronal survival, axonal growth, generation of new synaptic connections, regulation of neuronal activity as well as synaptic and dendritic plasticity. Emerging evidence also indicates an essential role for BDNF outside the nervous system, for instance in immune cells. In this study we focussed on the effect of BDNF on T cells, using conventional and conditional BDNF knockout (ko) mice lacking BDNF specifically in this lymphoid subset. In both settings, diminished T cell cellularity in peripheral lymphoid organs was observed. In order to get further insight into this phenomenon, we analysed T cell development in the thymus and found diminished total thymocyte numbers accompanied by a significant increase in CD4/CD8 double negative (DN) thymocytes due to a partial block in the transition from the DN3 to the DN4 stage. As underlying molecular mechanisms we could rule out increased thymocyte apoptosis as well as defects in the expression of the TCR -chain or the pre-TCR. In contrast, pERK but not pAKT levels were reduced in DN3 BDNF-deficient thymocytes pointing towards a MAP kinase-based
Abstracts
signalling defect as cause for this developmental T cell retardation. BDNF deficiency did not substantially influence peripheral acute immune responses and homeostatic proliferation of peripheral T cells. Taken together, our data reveal a critical autocrine and/or paracrine role of T cell-derived BDNF in thymocyte maturation involving ERK-mediated TCR signalling pathways.
doi:10.1016/j.jneuroim.2014.08.544
471 Cognitive performance among older individuals associated with the peripheral effector memory CD4+ T cells Cláudia Miranda, Susana Roque, Nadine Santos, Carlos Portugal-nunes, Patrício Costa, Joana Palha, Nuno Sousa, Margarida Correia-neves Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Braga, Portugal Progressive decline in immune function (immunosenescence) and in cognitive performance are common alterations accompanying aging. The interaction between these two systems has been highlighted in recent years mainly in studies using animal models. It has been shown that mice lacking B and T lymphocytes display cognitive impairments that are reverted by T cell replenishment. In fact, aging is accompanied by defined alterations in the adaptive immune system, namely in the T cell compartments. Taking into consideration the link between T cells and cognition, and since both age-associated cognitive decline and immunosenescence vary greatly between individuals we hypothesized that alterations in lymphocytes and cognitive performance are associated. In a cohort of cognitively healthy agers (n = 114), characterized by diverse neurocognitive/psychological performance patterns, we observed that better cognitive performance is associated with lower numbers of effector memory CD4+ T cell, higher numbers of naive CD8+ T cell and higher numbers of B cells in the blood. Furthermore, effector memory CD4+ T cells are predictors of general and executive function and memory, even when factors known to influence cognitive performance in older individuals (e.g. age, gender, education and mood) are taken into account. This is the first study identifying a phenotype of the immune system associated with poorer cognitive performance in normal cognitive aging, which strongly supports the interplay between the immune system and cognitive behavior and open a new perspective for interventions aiming to delay cognitive decline through aging.
doi:10.1016/j.jneuroim.2014.08.545
203
circulate as soluble forms, thereby neutralizing IL-1 activity. Expression of IL-1 is associated with experimental autoimmune encephalomyelitis (EAE), an animal model of human multiple sclerosis, which is believed to be mediated by T cells. In fact, exogenous IL-1 administration exacerbated the disease, while IL-1 knock-out (KO) animals showed resistance in EAE. To better understand the role of IL-1RI and IL-1RII in T cell encephalitogenicity we used the Cre/loxP system to delete these receptors either specifically in conventional T cells or in all cells of the body. For that purpose we crossed mice carrying Il1r1 and Il1r2 conditional alleles to the CD4-Cre or CMV-Cre transgenic mouse strains, respectively. All four groups of KO animals showed normal ontogenesis in steadystate conditions and T cells lacking IL-1R expression were triggered under in vitro polarization conditions towards effector subsets such as Th1 by adding the cytokines IL-2 and IL-12, or towards Th17 in the presence of TGF-beta, IL-6 and IL-23. As expected, T cells with deleted Il1r1 gene were non-responsive to IL-1 beta administration in a cell proliferation assay. Interestingly, mice lacking the IL-1RI developed EAE after active myelin oligodendrocyte glycoprotein (MOG) peptide immunization, although with a milder severity compared to wild type controls. At the peak of disease CNS-infiltrating IL-1RI KO T cells were characterized by significantly lower expression of effector cytokines, such as IL-17A, IFN-gamma and GM-CSF. In contrast, IL-1RII full KO mice showed an increased EAE severity, accompanied by enhanced cytokine expression from T cells. However, IL-1RII T cell specific KO animals developed a normal EAE. In this work we provide the first results obtained from two novel models of conditional IL-1RI and IL-1RII deletion. These mutations lead to opposite effects pointing to the necessity of intact IL-1 signaling for in vivo T cell development and pathogenicity. Thereby, the exacerbated disease in IL-1RII full KO animals suggests a negative contribution of this decoy receptor in EAE development. However, withdrawal of the receptor only from T cells is not enough to influence the disease progression, pointing to a contribution of IL-1RII from non-T cell sources. doi:10.1016/j.jneuroim.2014.08.546
523 Abnormal responses of CD8 + CD161high mucosal associated invariant T (MAIT) cells and CCR2 + CCR5+ CD4 T cells contribute to disrupted balance of effector and regulatory T cells in pediatric-onset MS Mukanthu Nyirendaa, Rui Lia, Craig Moorea, Ayal Rozenberga, Ayman Rezka, Trina Johnsonb, Dessa Sadovnickc, Douglas Arnolda, Ruth Ann Married, Brenda Banwelle,f, Amit Bar-ora,b, on behalf of the Canadian Pediatric Demyelinating Disease Network a
363 IL-1 receptor type I and type II differentially regulate EAE pathogenesis Ilgiz Mufazalov, Carsten Schelmbauer, Janina Kuschmann, Tommy Regen, Christopher Hackenbruch, Ari Waisman
Neuroimmunology Unit, Montreal Neurological Institute, McGill University, Montreal, Canada; bExperimental Therapeutics Program, Montreal Neurological Institute, McGill University, Montreal, Canada; c Division of Neurology, Department of Medical Genetics, University of British Columbia, British Columbia, Canada; dDepartments of Internal Medicine and Community Health, University of Manitoba, Manitoba, Canada; eDepartment of Paediatrics, Hospital For Sick Chilgdren, Toronto, Canada; fDivision of Neurology, Children's Hospital of Philadelphia, Philadelphia, United States
Institue for Molecular Medicine, JGU Medical Center, Mainz, Germany Interleukin-1 (IL-1) can bind to two receptors termed IL-1RI and IL1RII, but only the type I receptor is capable to transduce the signal. In addition, both receptors can be cleaved from the cell surface and
Abstract We wished to examine a carefully selected panel of effector and regulatory T cell (Treg) subsets (previously implicated in studies of adult-onset multiple sclerosis (MS) or other autoimmune diseases), at