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Leuenberger T.,Johannes Gutenberg University Mainz | Leuenberger T.,Max Delbrueck Center for Molecular Medicine Berlin Buch | Paterka M.,Johannes Gutenberg University Mainz | Paterka M.,Max Delbrueck Center for Molecular Medicine Berlin Buch | And 12 more authors.
Journal of Immunology | Year: 2013

T cells have an essential role in the induction of multiple sclerosis and its animal model experimental autoimmune encephalomyelitis (EAE). Although for CD4+T cells it is well established that they contribute to the disease, less is known about the role of CD8+T cells. Our aim was to determine the individual contribution of CD4+ and CD8+T cells in myelin oligodendrocyte glycoprotein (MOG)35-55-induced EAE. We investigated MOG35-55-activated CD8+T cells to clarify their potential to induce or attenuate EAE. We monitored the behavior of CD8 +T cells and their interaction with CD4+T cells directly at the site of inflammation in the CNS using intravital imaging of the brainstem of EAE-affected living anesthetized mice. We found that mice without CD4 +T cells did not develop relevant clinical signs of disease, although CD8+T cells were present in the CNS of these mice. These CD8 +T cells displayed reduced motility compared with those in the presence of CD4+T cells. In mice that harbored CD4+and CD8+T cells, we saw a similar extent of clinical signs of EAE as in mice with only CD4+T cells. Furthermore, the dynamic motility and viability of CD4+T cells were not disturbed by CD8+T cells in the lesions of these mice. Therefore, we conclude that in MOG35-55-induced EAE, CD8+T cell accumulation in the CNS represents instead an epiphenomenon with no impact on clinical disease or on the effects of CD4 +T cells, the latter being the true inducers of the disease. © 2013 by The American Association of Immunologists, Inc.


Leuenberger T.,Johannes Gutenberg University Mainz | Leuenberger T.,Max Delbrueck Center for Molecular Medicine Berlin Buch | Pfueller C.F.,Charité - Medical University of Berlin | Luessi F.,Johannes Gutenberg University Mainz | And 13 more authors.
PLoS ONE | Year: 2014

The maturation status of dendritic cells determines whether interacting T cells are activated or if they become tolerant. Previously we could induce T cell tolerance by applying a 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase inhibitor (HMGCRI) atorvastatin, which also modulates MHC class II expression and has therapeutic potential in autoimmune disease. Here, we aimed at elucidating the impact of this therapeutic strategy on T cell differentiation as a consequence of alterations in dendritic cell function. We investigated the effect of HMGCRI during differentiation of peripheral human monocytes and murine bone marrow precursors to immature DC in vitro and assessed their phenotype. To examine the stimulatory and tolerogenic capacity of these modulated immature dendritic cells, we measured proliferation and suppressive function of CD4+ T cells after stimulation with the modulated immature dendritic cells. We found that an HMGCRI, atorvastatin, prevents dendrite formation during the generation of immature dendritic cells. The modulated immature dendritic cells had a diminished capacity to take up and present antigen as well as to induce an immune response. Of note, the consequence was an increased capacity to differentiate naïve T cells towards a suppressor phenotype that is less sensitive to proinflammatory stimuli and can effectively inhibit the proliferation of T effector cells in vitro. Thus, manipulation of antigen-presenting cells by HMGCRI contributes to an attenuated immune response as shown by promotion of T cells with suppressive capacities. © 2014 Leuenberger et al.


PubMed | Max Delbrück Center for Molecular Medicine, Charité - Medical University of Berlin, Heinrich Heine University Düsseldorf, Max Delbrueck Center for Molecular Medicine Berlin Buch and 2 more.
Type: Journal Article | Journal: PloS one | Year: 2014

The maturation status of dendritic cells determines whether interacting T cells are activated or if they become tolerant. Previously we could induce T cell tolerance by applying a 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase inhibitor (HMGCRI) atorvastatin, which also modulates MHC class II expression and has therapeutic potential in autoimmune disease. Here, we aimed at elucidating the impact of this therapeutic strategy on T cell differentiation as a consequence of alterations in dendritic cell function. We investigated the effect of HMGCRI during differentiation of peripheral human monocytes and murine bone marrow precursors to immature DC in vitro and assessed their phenotype. To examine the stimulatory and tolerogenic capacity of these modulated immature dendritic cells, we measured proliferation and suppressive function of CD4+ T cells after stimulation with the modulated immature dendritic cells. We found that an HMGCRI, atorvastatin, prevents dendrite formation during the generation of immature dendritic cells. The modulated immature dendritic cells had a diminished capacity to take up and present antigen as well as to induce an immune response. Of note, the consequence was an increased capacity to differentiate nave T cells towards a suppressor phenotype that is less sensitive to proinflammatory stimuli and can effectively inhibit the proliferation of T effector cells in vitro. Thus, manipulation of antigen-presenting cells by HMGCRI contributes to an attenuated immune response as shown by promotion of T cells with suppressive capacities.


Glass R.,Ludwig Maximilians University of Munich | Synowitz M.,Charité - Medical University of Berlin | Synowitz M.,Max Delbrueck Center for Molecular Medicine Berlin Buch
Acta Neuropathologica | Year: 2014

Primary brain tumours (gliomas) initiate a strong host response and can contain large amounts of immune cells (myeloid cells) such as microglia and tumour-infiltrating macrophages. In gliomas the course of pathology is not only controlled by the genetic make-up of the tumour cells, but also depends on the interplay with myeloid cells in the tumour microenvironment. Especially malignant gliomas such as glioblastoma multiforme (GBM) are notoriously immune-suppressive and it is now evident that GBM cells manipulate myeloid cells to support tumour expansion. The protumorigenic effects of glioma-associated myeloid cells comprise a support for angiogenesis as well as tumour cell invasion, proliferation and survival. Different strategies for inhibiting the pathological functions of myeloid cells in gliomas are explored, and blocking the tropism of microglia/macrophages to gliomas or manipulating the signal transduction pathways for immune cell activation has been successful in pre-clinical models. Hence, myeloid cells are now emerging as a promising target for new adjuvant therapies for gliomas. However, it is also becoming evident that some myeloid-directed glioma therapies may only be beneficial for distinct subclasses of gliomas and that a more cell-type-specific manipulation of either microglia or macrophages may improve therapeutic outcomes. © 2014 Springer-Verlag.


Herz J.,Charité - Medical University of Berlin | Herz J.,Max Delbrueck Center for Molecular Medicine Berlin Buch | Zipp F.,Charité - Medical University of Berlin | Zipp F.,Max Delbrueck Center for Molecular Medicine Berlin Buch | And 2 more authors.
Experimental Neurology | Year: 2010

Multiple Sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system, in which the myelin sheath has been considered to be the primary target for many years. However, an increasing number of reports have focused on neurodegenerative aspects of the disease pathogenesis. Recent studies in post-mortem MS biopsies and in the animal model Experimental Autoimmune Encephalomyelitis (EAE) have shown that key features of neurodegeneration, i.e. axonal transection, neuronal cell atrophy and neuronal death already occur in early disease phases. Furthermore, it has become clear that irreversible disability correlates stronger with the neuronal affectation than with demyelination. However the cause of neuronal damage still remains elusive, since both demyelination-dependent and direct immune cell-mediated mechanisms have been suggested so far. Here, we summarize the current concepts and recently identified molecular mechanisms of inflammatory neurodegeneration in autoimmune CNS inflammation and highlight the role of different immune cells in the complex network of interactions leading to neuronal damage. © 2009 Elsevier Inc.


Ahmad F.,U.S. National Institutes of Health | Shen W.,U.S. National Institutes of Health | Vandeput F.,University of Utah | Szabo-Fresnais N.,University of Utah | And 7 more authors.
Journal of Biological Chemistry | Year: 2015

Cyclic nucleotide phosphodiesterase 3A (PDE3) regulates cAMP-mediated signaling in the heart, and PDE3 inhibitors augment contractility in patients with heart failure. Studies in mice showed that PDE3A, not PDE3B, is the subfamily responsible for these inotropic effects and that murine PDE3A1 associates with sarcoplasmic reticulum Ca2+ ATPase 2 (SERCA2), phospholamban (PLB), and AKAP18 in a multiprotein signalosome in human sarcoplasmic reticulum (SR). Immunohistochemical staining demonstrated that PDE3A co-localizes in Z-bands of human cardiac myocytes with desmin, SERCA2, PLB, and AKAP18. In human SR fractions, cAMP increased PLB phosphorylation and SERCA2 activity; this was potentiated by PDE3 inhibition but not by PDE4 inhibition. During gel filtration chromatography of solubilized SR membranes, PDE3 activity was recovered in distinct high molecular weight (HMW) and low molecular weight (LMW) peaks. HMW peaks contained PDE3A1 and PDE3A2, whereas LMW peaks contained PDE3A1, PDE3A2, and PDE3A3. Western blotting showed that endogenous HMW PDE3A1 was the principal PKA-phosphorylated isoform. Phosphorylation of endogenous PDE3A by rPKAc increased cAMP-hydrolytic activity, correlated with shift of PDE3A from LMW to HMW peaks, and increased co-immunoprecipitation of SERCA2, cav3, PKA regulatory subunit (PKARII), PP2A, and AKAP18 with PDE3A. In experiments with recombinant proteins, phosphorylation of recombinant human PDE3A isoforms by recombinant PKA catalytic subunit increased co-immunoprecipitation with rSERCA2 and rat rAKAP18 (recombinant AKAP18). Deletion of the recombinant human PDE3A1/PDE3A2 N terminus blocked interactions with recombinant SERCA2. Serine-to-alanine substitutions identified Ser-292/Ser-293, a site unique to human PDE3A1, as the principal site regulating its interaction with SERCA2. These results indicate that phosphorylation of human PDE3A1 at a PKA site in its unique N-terminal extension promotes its incorporation into SERCA2/AKAP18 signalosomes, where it regulates a discrete cAMP pool that controls contractility by modulating phosphorylation-dependent protein-protein interactions, PLB phosphorylation, and SERCA2 activity. © 2015, American Society for Biochemistry and Molecular Biology Inc. All rights reserved.


Schutze J.,Max Delbrueck Center for Molecular Medicine Berlin Buch | Schutze J.,Humboldt University of Berlin | Wolf J.,Max Delbrueck Center for Molecular Medicine Berlin Buch
BioSystems | Year: 2010

Glycolytic oscillations occur in many cell types and have been intensively studied in yeast. Recent experimental and theoretical research has been focussed on the oscillatory dynamics and the synchronisation mechanism in stirred yeast cell suspensions. Here we are interested in the spatio-temporal organisation of glycolysis in cell layers. To this end we study a grid of a few thousand compartments each containing a cell. The intracellular dynamics is described by a core model of glycolysis. The compartments can exchange metabolites via diffusion. The conditions for oscillatory dynamics in a single compartment are investigated by bifurcation analysis. The spatio-temporal behaviour of the cell layer is studied by simulations. The model predicts the propagation of repetitive wave fronts induced by a substrate gradient. The formation of these waves crucially depends on the diffusive exchange of the reaction product between cells. Depending on the kinetic parameters complex spatio-temporal behaviour such as periodic termination of waves can arise. In these cases the cellular oscillation characteristics depend on the location of the cell in the array. © 2009 Elsevier Ireland Ltd. All rights reserved.


PubMed | Max Delbrueck Center for Molecular Medicine Berlin Buch
Type: Journal Article | Journal: Current drug targets | Year: 2016

A-kinase anchoring proteins (AKAPs) control the localization of cAMP-dependent protein kinase A (PKA) by tethering PKA to distinct cellular compartments. Through additional direct proteinprotein interactions with PKA substrates and other signaling molecules they form multi-protein complexes. Thereby, AKAPs regulate the access of PKA to its substrates in a temporal and spatial manner as well as the local crosstalk of cAMP/PKA with other signaling pathways. Due to the increasing information on their molecular functioning and three-dimensional structures, and their emerging roles in the development of diseases, AKAPs move into the focus as potential drug targets. Targeting AKAP dependent protein-protein interactions for interference with local signal processing inside cells potentially allows for the development of therapeutics with high selectivity and fewer side effects.

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