Primary oligodendropathy is not a trigger of CNS autoimmunity

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1 Primary oligodendropathy is not a trigger of CNS autoimmunity Ari Waisman Institute for Molecular Medicine University Medical Center, JGU Mainz 1

2 How is an anti-myelin immune response initiated? Secondary Lymphoid Organs Blood Brain Barrier Invasion Central Nervous System T cells Antigen Presenting Cell T cell activation Neuron secondary oligodendrocyte initial oligodendropathy damage Oligodendrocyte antigen release A T cell model of MS/EAE Immunopathogenesis 1. Activation Th 4. Invasion 3. Attraction 2. Adhesion 5. Reactivation Th1 Periphery BBB CNS Amit Bar-Or 2

3 Importance of oligodendrocytes in EAE/MS Induction To study the importance of ODCs in the induction phase of EAE, we set a system that allows us to damage the ODCs in a non-inflammatory way, then to study how this effect the myelin an EAE induction Generation of the idtr mice Diphtheria toxin: Mode of action The A subunit catalyzes the addition of ADP-Ribose to the Elongation Factor EF- 2 -> suppression of translation -> cell death Buch et al., Nature Methods,

4 MOGi-Cre recombines only in oligodendrocytes NeuN/EGFP CNPase/EGFP CP S100/EGFP Hövelmeyer et al., Journal of Immunology, 2005 (with Hans Lassmann) Induced Death of ODCs - the odtr Mice MOG-iCre MOG cre idtr Rosa26 Cre mediated recombination STOP DT-Receptor pa DT-Receptor pa Oligodendrocyte (ODC) specific cell death (ODCs) MOG-iCre X idtr = odtr Diphteria Toxin (DT) ODC 4

5 Neurological symptoms of odtr mice after DT injection Mice start to shiver after 4-5 weeks Shivering results in complete paralysis. Clinical course of oligodendropathy Weight Neurological score Time on rotarod Grid walk disease onset after 30 d - ataxia - tremor - hunchback phenotype 5

6 DT induced ODC Death aspa PLP LFB LFB 1 week 6 weeks Demyelination and CNS damage odtr MogiCre LFB Spinal cord of odtr mice 5-6 week post DT treatment 6

7 ODC death leads to pronounced demyelination ASPA PLP LFB odtr control Peak of disease Activation and accumulation of microglia and astrocytes week 2 weeks 3 weeks 4 weeks F4/80 GFAP 7

8 MHC II expression on activated microglia control odtr MHC II microglia MHC II microglia NG2 expression upon DT induced ODC death MOGi-Cre odtr cerebellum corpus callosum striatum cerebellum corpus callosum striatum 4 weeks 3 weeks -NG2 8

9 NG2 Expression upon DT induced ODC Death control week 2 weeks 3 weeks 4 weeks cerebellum striatum NG2 Antigen leakage and T Cell Activation odtr MOGi-Cre DT MOG-specific T cells odtr anti-cd40 Accumulation of Lipids shown by OIL RED staining No proliferation of naive MOG-specific Th cells OilRedO pos control neg control odtr spleen Proliferation of co-stimulated (anti- CD40) MOG-specific Th cells in some but not all animals CD4 CFSE lymphnode 9

10 EAE and oligodendropathy immunized with MOG immunized without MOG MogiCre/2D2 odtr/2d2 EAE III 6 5,5 5 4, clinical score 4 3,5 3 2,5 2 1,5 1 0,5 percent mice healthy sick days post immunization 0 no Dtx odtr/2d2 DTx MOG-iCre/2D2 Primary ODC death does not enhance EAE Antigen leakage from DT induced Demyelination is not sufficient for EAE induction EAE and oligodendropathy Demyelination Demyelination + EAE Demyelination + mild EAE (no Pertussis Toxin) Demyelination + Treg ablation ( CD25) Demyelination + demyelinating mab (8-18C5) Demyelination + sublethal irradiation Demyelination + anti-cd40 Demyelination + anti-mog TCR Demyelination + anti-mog TCR + Pertussis/CFA weekly weak insult (50 ng DT) no autoimmunity no significant increase no significant increase no autoimmunity no autoimmunity no autoimmunity no autoimmunity no significant increase no significant increase no disease (so far) 10

11 Conclusions odtr The odtr mice provide a defined system that allow to induce demyelination and follow remyelination We could not detect any antigen drainage at least we found no activation of MOG-specific CD4 cells Likewise, we found no effect on the induction of EAE by ODC death We conclude that induced death of ODCs does not lead to autoimmunity or alters susceptibility to EAE Locatelli, Wörtge, Buch et al., Nature Neuroscience, 2012 MS and pain The most common form of pain in MS is central neuropathic extremity pain, which is distinguished by continuous burning pain as well as mechanical and cold allodynia. Nearly all MS patients who experience central pain report alterations in temperature or pain sensation in the affected region, suggesting contributions of lesions in the spinothalamic tract. 11

12 odtr are noxious cold sensitive odtr are cold but not heat sensitive 12

13 Axonal damage in odtr mice Mechanical and cold sensitivity in odtr mice is independent of microglia activation and inflammation Gritsch, Lu et al., Nature Communications,

14 Conclusions odtr and pain Primary oligodendrocyte loss in the absence of adaptive immunity can induce symptoms of central neuropathic pain, which are similar to those found in MS patients. Neither the microglia inhibitor Minocycline nor FTY720, an inhibitor of lymphocyte infiltration that is highly effective in the reducing CNS inflammation, affected the development or maintenance of nociceptive hypersensitivity upon oligodendrocyte loss. These results suggest that targeting oligodendrocyte function and axonal pathology in addition to immunebased therapies may be of value in the context of pain treatment. More people involved Burkhard Becher Frank Heppner (and Barbara) Giuseppe Locatelli Rohini Kuner; Klaus-Armin Nave 14

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