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Astrocytic A20 ameliorates experimental autoimmune encephalomyelitis by inhibiting NF-κB- and STAT1-dependent chemokine production in astrocytes

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Abstract

Single-nucleotide polymorphisms in the tumor necrosis factor, alpha-induced protein 3 gene, which encodes the ubiquitin-modifying protein A20, are linked to susceptibility to multiple sclerosis (MS), a demyelinating autoimmune disease of the central nervous system (CNS). Since it is unresolved how A20 regulates MS pathogenesis, we examined its function in a murine model of MS, namely experimental autoimmune encephalomyelitis (EAE). Deletion of A20 in neuroectodermal cells (astrocytes, neurons, and oligodendrocytes; Nestin-Cre A20fl/fl mice) or selectively in astrocytes (GFAP-Cre A20fl/fl mice) resulted in more severe EAE as compared to control animals. In Nestin-Cre A20fl/fl and GFAP-Cre A20fl/fl mice demyelination and recruitment of inflammatory leukocytes were increased as compared to A20fl/fl control mice. Importantly, numbers of encephalitogenic CD4+ T cells producing interferon (IFN)-γ, interleukin (IL)-17, and granulocyte–macrophage colony-stimulating factor (GM-CSF), respectively, as well as mRNA production of IFN-γ, IL-17, tumor necrosis factor (TNF), GM-CSF, IL-6, CXCL1, CCL2, and CXCL10 were significantly increased in spinal cords of Nestin-Cre A20fl/fl and GFAP-Cre A20fl/fl mice, respectively. Compared to A20-sufficient astrocytes, A20-deficient astrocytes displayed stronger activation of nuclear factor kappa-light-chain enhancer of activated B cells (NF-κB) in response to TNF, IL-17, and GM-CSF, and of signal transducer and activator of transcription 1 (STAT1) upon IFN-γ stimulation. Due to NF-κB and STAT1 hyperactivation, A20-deficient astrocytes produced significantly more chemokines in response to these key encephalitogenic cytokines of autoimmune CD4+ T cells resulting in an amplification of CD4+ T cell recruitment to the CNS. Thus, astrocytic A20 is an important inhibitor of autoimmune-mediated demyelination in the CNS.

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References

  1. Anderton SM, Liblau RS (2008) Regulatory T cells in the control of inflammatory demyelinating diseases of the central nervous system. Curr Opin Neurol 21(3):248–254

    Article  PubMed  CAS  Google Scholar 

  2. Bajenaru ML, Zhu Y, Hedrick NM, Donahoe J, Parada LF, Gutmann DH (2002) Astrocyte-specific inactivation of the neurofibromatosis 1 gene (NF1) is insufficient for astrocytoma formation. Mol Cell Biol 22(14):5100–5113

    Article  PubMed  CAS  Google Scholar 

  3. Baxter AG (2007) The origin and application of experimental autoimmune encephalomyelitis. Nat Rev Immunol 7(11):904–912

    Article  PubMed  CAS  Google Scholar 

  4. Brambilla R, Persaud T, Hu X, Karmally S, Shestopalov VI, Dvoriantchikova G, Ivanov D, Nathanson L, Barnum SR, Bethea JR (2009) Transgenic inhibition of astroglial NF-kappa B improves functional outcome in experimental autoimmune encephalomyelitis by suppressing chronic central nervous system inflammation. J Immunol 182(5):2628–2640

    Article  PubMed  CAS  Google Scholar 

  5. Brück W, Pförtner R, Pham T, Zhang J, Hayardeny L, Piryatinsky V, Hanisch UK, Regen T, van Rossum D, Brakelmann L, Hagemeier K, Kuhlmann T, Stadelmann C, John GR, Kramann N, Wegner C (2012) Reduced astrocytic NF-kappaB activation by laquinimod protects from cuprizone-induced demyelination. Acta Neuropathol 124(3):411–424

    Article  PubMed  Google Scholar 

  6. Brüstle A, Brenner D, Knobbe CB, Lang PA, Virtanen C, Hershenfield BM, Reardon C, Lacher SM, Ruland J, Ohashi PS, Mak TW (2012) The NF-kappaB regulator MALT1 determines the encephalitogenic potential of Th17 cells. J Clin Invest 122(12):4698–4709

    Article  PubMed  Google Scholar 

  7. Bulek K, Liu C, Swaidani S, Wang L, Page RC, Gulen MF, Herjan T, Abbadi A, Qian W, Sun D, Lauer M, Hascall V, Misra S, Chance MR, Aronica M, Hamilton T, Li X (2011) The inducible kinase IKKi is required for IL-17-dependent signaling associated with neutrophilia and pulmonary inflammation. Nat Immunol 12(9):844–852

    Article  PubMed  CAS  Google Scholar 

  8. Chu Y, Vahl JC, Kumar D, Heger K, Bertossi A, Wojtowicz E, Soberon V, Schenten D, Mack B, Reutelshöfer M, Beyaert R, Amann K, van Loo G, Schmidt-Supprian M (2011) B cells lacking the tumor suppressor TNFAIP3/A20 display impaired differentiation and hyperactivation and cause inflammation and autoimmunity in aged mice. Blood 117(7):2227–2236

    Article  PubMed  CAS  Google Scholar 

  9. Codarri L, Gyülveszi G, Tosevski V, Hesske L, Fontana A, Magnenat L, Suter T, Becher B (2011) RORgammat drives production of the cytokine GM-CSF in helper T cells, which is essential for the effector phase of autoimmune neuroinflammation. Nat Immunol 12(6):560–567

    Article  PubMed  CAS  Google Scholar 

  10. De Jager PL, Jia X, Wang J, de Bakker PI, Ottoboni L, Aggarwal NT, Piccio L, Raychaudhuri S, Tran D, Aubin C, Briskin R, Romano S, Baranzini SE, McCauley JL, Pericak-Vance MA, Haines JL, Gibson RA, Naeglin Y, Uitdehaag B, Matthews PM, Kappos L, Polman C, McArdle WL, Strachan DP, Evans D, Cross AH, Daly MJ, Compston A, Sawcer SJ, Weiner HL, Hauser SL, Hafler DA, Oksenberg JR (2009) Meta-analysis of genome scans and replication identify CD6, IRF8 and TNFRSF1A as new multiple sclerosis susceptibility loci. Nat Genet 41(7):776–782

    Article  PubMed  Google Scholar 

  11. Decker T, Kovarik P (2000) Serine phosphorylation of STATs. Oncogene 19(21):2628–2637

    Article  PubMed  CAS  Google Scholar 

  12. Domingues HS, Mues M, Lassmann H, Wekerle H, Krishnamoorthy G (2010) Functional and pathogenic differences of Th1 and Th17 cells in experimental autoimmune encephalomyelitis. PLoS One 5(11):e15531

    Article  PubMed  CAS  Google Scholar 

  13. Dong Y, Benveniste EN (2001) Immune function of astrocytes. Glia 36(2):180–190

    Article  PubMed  CAS  Google Scholar 

  14. Engelhardt B, Sorokin L (2009) The blood–brain and the blood–cerebrospinal fluid barriers: function and dysfunction. Semin Immunopathol 31(4):497–511

    Article  PubMed  Google Scholar 

  15. Fung EY, Smyth DJ, Howson JM, Cooper JD, Walker NM, Stevens H, Wicker LS, Todd JA (2009) Analysis of 17 autoimmune disease-associated variants in type 1 diabetes identifies 6q23/TNFAIP3 as a susceptibility locus. Genes Immun 10(2):188–191

    Article  PubMed  CAS  Google Scholar 

  16. Gilli F, Navone ND, Perga S, Marnetto F, Caldano M, Capobianco M, Pulizzi A, Malucchi S, Bertolotto A (2011) Loss of braking signals during inflammation: a factor affecting the development and disease course of multiple sclerosis. Arch Neurol 68(7):879–888

    Article  PubMed  Google Scholar 

  17. Graus-Porta D, Blaess S, Senften M, Littlewood-Evans A, Damsky C, Huang Z, Orban P, Klein R, Schittny JC, Müller U (2001) Beta1-class integrins regulate the development of laminae and folia in the cerebral and cerebellar cortex. Neuron 31(3):367–379

    Article  PubMed  CAS  Google Scholar 

  18. Hammer GE, Turer EE, Taylor KE, Fang CJ, Advincula R, Oshima S, Barrera J, Huang EJ, Hou B, Malynn BA, Reizis B, DeFranco A, Criswell LA, Nakamura MC, Ma A (2011) Expression of A20 by dendritic cells preserves immune homeostasis and prevents colitis and spondyloarthritis. Nat Immunol 12(12):1184–1193

    Article  PubMed  CAS  Google Scholar 

  19. Händel U, Brunn A, Drögemüller K, Müller W, Deckert M, Schlüter D (2012) Neuronal gp130 expression is crucial to prevent neuronal loss, hyperinflammation, and lethal course of murine Toxoplasma encephalitis. Am J Pathol 181(1):163–173

    Article  PubMed  Google Scholar 

  20. Haroon F, Drögemüller K, Händel U, Brunn A, Reinhold D, Nishanth G, Mueller W, Trautwein C, Ernst M, Deckert M, Schlüter D (2011) Gp130-dependent astrocytic survival is critical for the control of autoimmune central nervous system inflammation. J Immunol 186(11):6521–6531

    Article  PubMed  CAS  Google Scholar 

  21. Hövelmeyer N, Reissig S, Xuan NT, Adams-Quack P, Lukas D, Nikolaev A, Schlüter D, Waisman A (2011) A20 deficiency in B cells enhances B-cell proliferation and results in the development of autoantibodies. Eur J Immunol 41(3):595–601

    Article  PubMed  Google Scholar 

  22. International Multiple Sclerosis Genetics Conssortium (IMSGC) (2010) IL12A, MPHOSPH9/CDK2AP1 and RGS1 are novel multiple sclerosis susceptibility loci. Genes Immun 11(5):397–405

    Article  Google Scholar 

  23. Kang Z, Altuntas CZ, Gulen MF, Liu C, Giltiay N, Qin H, Liu L, Qian W, Ransohoff RM, Bergmann C, Stohlman S, Tuohy VK, Li X (2010) Astrocyte-restricted ablation of interleukin-17-induced Act1-mediated signaling ameliorates autoimmune encephalomyelitis. Immunity 32(3):414–425

    Article  PubMed  CAS  Google Scholar 

  24. Kebir H, Kreymborg K, Ifergan I, Dodelet-Devillers A, Cayrol R, Bernard M, Giuliani F, Arbour N, Becher B, Prat A (2007) Human TH17 lymphocytes promote blood–brain barrier disruption and central nervous system inflammation. Nat Med 13(10):1173–1175

    Article  PubMed  CAS  Google Scholar 

  25. Kool M, van Loo G, Waelput W, De Prijck S, Muskens F, Sze M, van Praet J, Branco-Madeira F, Janssens S, Reizis B, Elewaut D, Beyaert R, Hammad H, Lambrecht BN (2011) The ubiquitin-editing protein A20 prevents dendritic cell activation, recognition of apoptotic cells, and systemic autoimmunity. Immunity 35(1):82–96

    Article  PubMed  CAS  Google Scholar 

  26. Langrish CL, Chen Y, Blumenschein WM, Mattson J, Basham B, Sedgwick JD, McClanahan T, Kastelein RA, Cua DJ (2005) IL-23 drives a pathogenic T cell population that induces autoimmune inflammation. J Exp Med 201(2):233–240

    Article  PubMed  CAS  Google Scholar 

  27. Le DG, Kular L, Nicot AB, Calmel C, Melik-Parsadaniantz S, Kitabgi P, Laurent M, Martinerie C (2010) NOV/CCN3 upregulates CCL2 and CXCL1 expression in astrocytes through beta1 and beta5 integrins. Glia 58(12):1510–1521

    Google Scholar 

  28. Lee EG, Boone DL, Chai S, Libby SL, Chien M, Lodolce JP, Ma A (2000) Failure to regulate TNF-induced NF-kappaB and cell death responses in A20-deficient mice. Science 289(5488):2350–2354

    Article  PubMed  CAS  Google Scholar 

  29. Lees JR, Golumbek PT, Sim J, Dorsey D, Russell JH (2008) Regional CNS responses to IFN-gamma determine lesion localization patterns during EAE pathogenesis. J Exp Med 205(11):2633–2642

    Article  PubMed  CAS  Google Scholar 

  30. Linden M, Khademi M, Lima Bomfim I, Piehl F, Jagodic M, Kockum I, Olsson T (2013) Multiple sclerosis risk genotypes correlate with an elevated cerebrospinal fluid level of the suggested prognostic marker CXCL13. Mult Scler 19(7):863–870

    Article  PubMed  CAS  Google Scholar 

  31. Liu Y, Teige I, Birnir B, Issazadeh-Navikas S (2006) Neuron-mediated generation of regulatory T cells from encephalitogenic T cells suppresses EAE. Nat Med 12(5):518–525

    Article  PubMed  CAS  Google Scholar 

  32. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) method. Methods 25(4):402–408

    Article  PubMed  CAS  Google Scholar 

  33. Ma A, Malynn BA (2012) A20: linking a complex regulator of ubiquitylation to immunity and human disease. Nat Rev Immunol 12(11):774–785

    Article  PubMed  CAS  Google Scholar 

  34. Matmati M, Jacques P, Maelfait J, Verheugen E, Kool M, Sze M, Geboes L, Louagie E, Mc Guire C, Vereecke L, Chu Y, Boon L, Staelens S, Matthys P, Lambrecht BN, Schmidt-Supprian M, Pasparakis M, Elewaut D, Beyaert R, van Loo G (2011) A20 (TNFAIP3) deficiency in myeloid cells triggers erosive polyarthritis resembling rheumatoid arthritis. Nat Genet 43(9):908–912

    Article  PubMed  CAS  Google Scholar 

  35. McFarland HF, Martin R (2007) Multiple sclerosis: a complicated picture of autoimmunity. Nat Immunol 8(9):913–919

    Article  PubMed  CAS  Google Scholar 

  36. McGeachy MJ, Stephens LA, Anderton SM (2005) Natural recovery and protection from autoimmune encephalomyelitis: contribution of CD4+CD25+ regulatory cells within the central nervous system. J Immunol 175(5):3025–3032

    PubMed  CAS  Google Scholar 

  37. Meads MB, Li ZW, Dalton WS (2010) A novel TNF receptor-associated factor 6 binding domain mediates NF-kappa B signaling by the common cytokine receptor beta subunit. J Immunol 185(3):1606–1615

    Article  PubMed  CAS  Google Scholar 

  38. Miljkovic D, Timotijevic G, Mostarica SM (2011) Astrocytes in the tempest of multiple sclerosis. FEBS Lett 585(23):3781–3788

    Article  PubMed  CAS  Google Scholar 

  39. Musone SL, Taylor KE, Lu TT, Nititham J, Ferreira RC, Ortmann W, Shifrin N, Petri MA, Kamboh MI, Manzi S, Seldin MF, Gregersen PK, Behrens TW, Ma A, Kwok PY, Criswell LA (2008) Multiple polymorphisms in the TNFAIP3 region are independently associated with systemic lupus erythematosus. Nat Genet 40(9):1062–1064

    Article  PubMed  CAS  Google Scholar 

  40. Musone SL, Taylor KE, Nititham J, Chu C, Poon A, Liao W, Lam ET, Ma A, Kwok PY, Criswell LA (2011) Sequencing of TNFAIP3 and association of variants with multiple autoimmune diseases. Genes Immun 12(3):176–182

    Article  PubMed  CAS  Google Scholar 

  41. Nair A, Frederick TJ, Miller SD (2008) Astrocytes in multiple sclerosis: a product of their environment. Cell Mol Life Sci 65(17):2702–2720

    Article  PubMed  CAS  Google Scholar 

  42. Nair RP, Duffin KC, Helms C, Ding J, Stuart PE, Goldgar D, Gudjonsson JE, Li Y, Tejasvi T, Feng BJ, Ruether A, Schreiber S, Weichenthal M, Gladman D, Rahman P, Schrodi SJ, Prahalad S, Guthery SL, Fischer J, Liao W, Kwok PY, Menter A, Lathrop GM, Wise CA, Begovich AB, Voorhees JJ, Elder JT, Krueger GG, Bowcock AM, Abecasis GR (2009) Genome-wide scan reveals association of psoriasis with IL-23 and NF-kappaB pathways. Nat Genet 41(2):199–204

    Article  PubMed  CAS  Google Scholar 

  43. Ozawa K, Suchanek G, Breitschopf H, Bruck W, Budka H, Jellinger K, Lassmann H (1994) Patterns of oligodendroglia pathology in multiple sclerosis. Brain 117(Pt 6):1311–1322

    Article  PubMed  Google Scholar 

  44. Petermann F, Korn T (2011) Cytokines and effector T cell subsets causing autoimmune CNS disease. FEBS Lett 585(23):3747–3757

    Article  PubMed  CAS  Google Scholar 

  45. Reboldi A, Coisne C, Baumjohann D, Benvenuto F, Bottinelli D, Lira S, Uccelli A, Lanzavecchia A, Engelhardt B, Sallusto F (2009) C–C chemokine receptor 6-regulated entry of TH-17 cells into the CNS through the choroid plexus is required for the initiation of EAE. Nat Immunol 10(5):514–523

    Article  PubMed  CAS  Google Scholar 

  46. Shembade N, Ma A, Harhaj EW (2010) Inhibition of NF-kappaB signaling by A20 through disruption of ubiquitin enzyme complexes. Science 327(5969):1135–1139

    Article  PubMed  CAS  Google Scholar 

  47. Skaug B, Chen J, Du F, He J, Ma A, Chen ZJ (2011) Direct, noncatalytic mechanism of IKK inhibition by A20. Mol Cell 44(4):559–571

    Article  PubMed  CAS  Google Scholar 

  48. Sofroniew MV, Vinters HV (2010) Astrocytes: biology and pathology. Acta Neuropathol 119(1):7–35

    Article  PubMed  Google Scholar 

  49. Stromnes IM, Cerretti LM, Liggitt D, Harris RA, Goverman JM (2008) Differential regulation of central nervous system autoimmunity by T(H)1 and T(H)17 cells. Nat Med 14(3):337–342

    Article  PubMed  CAS  Google Scholar 

  50. Tavares RM, Turer EE, Liu CL, Advincula R, Scapini P, Rhee L, Barrera J, Lowell CA, Utz PJ, Malynn BA, Ma A (2010) The ubiquitin modifying enzyme A20 restricts B cell survival and prevents autoimmunity. Immunity 33(2):181–191

    Article  PubMed  CAS  Google Scholar 

  51. Tronche F, Kellendonk C, Kretz O, Gass P, Anlag K, Orban PC, Bock R, Klein R, Schutz G (1999) Disruption of the glucocorticoid receptor gene in the nervous system results in reduced anxiety. Nat Genet 23(1):99–103

    Article  PubMed  CAS  Google Scholar 

  52. van Loo G, De Lorenzi R, Schmidt H, Huth M, Mildner A, Schmidt-Supprian M, Lassmann H, Prinz MR, Pasparakis M (2006) Inhibition of transcription factor NF-kappaB in the central nervous system ameliorates autoimmune encephalomyelitis in mice. Nat Immunol 7(9):954–961

    Article  PubMed  Google Scholar 

  53. Varinou L, Ramsauer K, Karaghiosoff M, Kolbe T, Pfeffer K, Müller M, Decker T (2003) Phosphorylation of the Stat1 transactivation domain is required for full-fledged IFN-gamma-dependent innate immunity. Immunity 19(6):793–802

    Article  PubMed  CAS  Google Scholar 

  54. Voskuhl RR, Peterson RS, Song B, Ao Y, Morales LB, Tiwari-Woodruff S, Sofroniew MV (2009) Reactive astrocytes form scar-like perivascular barriers to leukocytes during adaptive immune inflammation of the CNS. J Neurosci 29(37):11511–11522

    Article  PubMed  CAS  Google Scholar 

  55. Vucic D, Dixit VM, Wertz IE (2011) Ubiquitylation in apoptosis: a post-translational modification at the edge of life and death. Nat Rev Mol Cell Biol 12(7):439–452

    Article  PubMed  CAS  Google Scholar 

  56. Wang X, Haroon F, Karray S, Martina D, Schlüter D (2013) Astrocytic Fas ligand expression is required to induce T-cell apoptosis and recovery from experimental autoimmune encephalomyelitis. Eur J Immunol 43(1):115–124

    Article  PubMed  CAS  Google Scholar 

  57. Wertz IE, O’Rourke KM, Zhou H, Eby M, Aravind L, Seshagiri S, Wu P, Wiesmann C, Baker R, Boone DL, Ma A, Koonin EV, Dixit VM (2004) De-ubiquitination and ubiquitin ligase domains of A20 downregulate NF-kappaB signalling. Nature 430(7000):694–699

    Article  PubMed  CAS  Google Scholar 

  58. Yuan C, Qi J, Zhao X, Gao C (2012) Smurf1 protein negatively regulates interferon-gamma signaling through promoting STAT1 protein ubiquitination and degradation. J Biol Chem 287(21):17006–17015

    Article  PubMed  CAS  Google Scholar 

  59. Zepp J, Wu L, Li X (2011) IL-17 receptor signaling and T helper 17-mediated autoimmune demyelinating disease. Trends Immunol 32(5):232–239

    Article  PubMed  CAS  Google Scholar 

  60. Zhu Y, Romero MI, Ghosh P, Ye Z, Charnay P, Rushing EJ, Marth JD, Parada LF (2001) Ablation of NF1 function in neurons induces abnormal development of cerebral cortex and reactive gliosis in the brain. Genes Dev 15(7):859–876

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

The authors thank Elena Fischer, Nadja Schlüter, and Annette Sohnekind for technical assistance. This work was supported by grants of the Deutsche Forschungsgemeinschaft (GRK 1167 to DS; SFB 854/TP5 to DS and MN).

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Authors and Affiliations

  1. Institute of Medical Microbiology, Otto-von-Guericke University Magdeburg, Leipzigerstrasse 44, 39120, Magdeburg, Saxony-Anhalt, Germany

    Xu Wang, Nguyen Thi Xuan, Gopala Nishanth, Sissy Just & Dirk Schlüter

  2. Department of Neuropathology, University Hospital Cologne, 50931, Cologne, Germany

    Martina Deckert

  3. Institute for Molecular Medicine, University Medical Center of the Johannes-Gutenberg University of Mainz, 55131, Mainz, Germany

    Ari Waisman

  4. Institute of Experimental Internal Medicine, Otto-von-Guericke University Magdeburg, 39120, Magdeburg, Germany

    Michael Naumann

  5. Helmholtz Centre for Infection Research, 38124, Braunschweig, Germany

    Dirk Schlüter

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  1. Xu Wang
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Correspondence to Dirk Schlüter.

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401_2013_1183_MOESM1_ESM.tif

Online Resource 1 Upregulation of A20 in the spinal cord during EAE mRNA transcription of A20 in the spinal cord was evaluated by quantitative RT-PCR in untreated and MOG35-55-immunized C57BL/6 mice at day 15 and 22 p.i (n = 3). Data are expressed as the increase of A20 mRNA of immunized over unimmunized mice normalized to HPRT (mean + SEM * p < 0.05) (TIFF 118 kb)

401_2013_1183_MOESM2_ESM.tif

Online Resource 2 EAE was not aggravated in Nestin-Cre A20wt/wt mice Clinical scores of Nestin-Cre A20wt/wt (n = 12) and A20wt/wt (n = 11) mice after active immunization with MOG35-55 peptide. Data show the mean clinical scores + SEM (TIFF 195 kb)

401_2013_1183_MOESM3_ESM.tif

Online Resource 3 EAE was not aggravated in GFAP-Cre A20wt/wt mice and Synapsin-Cre A20fl/fl mice a WB analysis for A20 expression in FACS-sorted microglia and cultured neurons from GFAP-Cre A20fl/fl and A20fl/fl mice. b Clinical scores of EAE in GFAP-Cre A20wt/wt (n = 5) and A20wt/wt (n = 5) mice induced by MOG35-55-immunization. Data show the mean clinical scores + SEM. c WB analysis for A20 expression in cultured neurons from neuron-restricted A20 deficient (Synapsin-Cre A20fl/fl) and control (A20fl/fl) mice. The right panel shows the relative quantification of A20 normalized to GAPDH. Data show the mean + SEM. d Clinical scores of EAE in Synapsin-Cre A20fl/fl (n = 8) and A20fl/fl (n = 6) mice induced by MOG35-55-immunization. Data show the mean clinical scores + SEM (TIFF 1013 kb)

401_2013_1183_MOESM4_ESM.tif

Online Resource 4 Normal apoptosis of A20-deficient and -sufficient astrocytes after TNF stimulation Astrocytes isolated from GFAP-Cre A20fl/fl and A20fl/fl mice were treated with 20 or 100 ng/ml TNF, respectively, for 24 h. Apoptosis was detected by PI and Annexin staining (TIFF 583 kb)

401_2013_1183_MOESM5_ESM.tif

Online Resource 5 STAT1 expression was upregulated in astrocytes after A20 siRNA treatment a Astrocytes derived from C57BL/6 mice were transfected with siRNA targeting A20 or nonsense siRNA. Sixty hours later, WB analysis was performed on total cell lysates for A20, STAT1 and GAPDH. b Relative quantification of A20 normalized to GAPDH. Data show the mean + SEM. c Relative quantification of STAT1 normalized to GAPDH. Data show the mean + SEM (TIFF 246 kb)

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Wang, X., Deckert, M., Xuan, N.T. et al. Astrocytic A20 ameliorates experimental autoimmune encephalomyelitis by inhibiting NF-κB- and STAT1-dependent chemokine production in astrocytes. Acta Neuropathol 126, 711–724 (2013). https://doi.org/10.1007/s00401-013-1183-9

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