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1.
Proc Natl Acad Sci U S A ; 114(7): E1168-E1177, 2017 02 14.
Artigo em Inglês | MEDLINE | ID: mdl-28137846

RESUMO

Disruption of the blood-brain barrier (BBB) is a defining and early feature of multiple sclerosis (MS) that directly damages the central nervous system (CNS), promotes immune cell infiltration, and influences clinical outcomes. There is an urgent need for new therapies to protect and restore BBB function, either by strengthening endothelial tight junctions or suppressing endothelial vesicular transcytosis. Although wingless integrated MMTV (Wnt)/ß-catenin signaling plays an essential role in BBB formation and maintenance in healthy CNS, its role in BBB repair in neurologic diseases such as MS remains unclear. Using a Wnt/ß-catenin reporter mouse and several downstream targets, we demonstrate that the Wnt/ß-catenin pathway is up-regulated in CNS endothelial cells in both human MS and the mouse model experimental autoimmune encephalomyelitis (EAE). Increased Wnt/ß-catenin activity in CNS blood vessels during EAE progression correlates with up-regulation of neuronal Wnt3 expression, as well as breakdown of endothelial cell junctions. Genetic inhibition of the Wnt/ß-catenin pathway in CNS endothelium before disease onset exacerbates the clinical presentation of EAE, CD4+ T-cell infiltration into the CNS, and demyelination by increasing expression of vascular cell adhesion molecule-1 and the transcytosis protein Caveolin-1 and promoting endothelial transcytosis. However, Wnt signaling attenuation does not affect the progressive degradation of tight junction proteins or paracellular BBB leakage. These results suggest that reactivation of Wnt/ß-catenin signaling in CNS vessels during EAE/MS partially restores functional BBB integrity and limits immune cell infiltration into the CNS.


Assuntos
Linfócitos T CD4-Positivos/metabolismo , Células Endoteliais/metabolismo , Esclerose Múltipla/metabolismo , Via de Sinalização Wnt , beta Catenina/metabolismo , Animais , Barreira Hematoencefálica/metabolismo , Caveolina 1/metabolismo , Sistema Nervoso Central/metabolismo , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/metabolismo , Feminino , Humanos , Camundongos Congênicos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Esclerose Múltipla/genética , Transcitose , beta Catenina/genética
2.
PLoS Biol ; 14(5): e1002467, 2016 05.
Artigo em Inglês | MEDLINE | ID: mdl-27213272

RESUMO

Growth factors of the gp130 family promote oligodendrocyte differentiation, and viability, and myelination, but their mechanisms of action are incompletely understood. Here, we show that these effects are coordinated, in part, by the transcriptional activator Krüppel-like factor-6 (Klf6). Klf6 is rapidly induced in oligodendrocyte progenitors (OLP) by gp130 factors, and promotes differentiation. Conversely, in mice with lineage-selective Klf6 inactivation, OLP undergo maturation arrest followed by apoptosis, and CNS myelination fails. Overlapping transcriptional and chromatin occupancy analyses place Klf6 at the nexus of a novel gp130-Klf-importin axis, which promotes differentiation and viability in part via control of nuclear trafficking. Klf6 acts as a gp130-sensitive transactivator of the nuclear import factor importin-α5 (Impα5), and interfering with this mechanism interrupts step-wise differentiation. Underscoring the significance of this axis in vivo, mice with conditional inactivation of gp130 signaling display defective Klf6 and Impα5 expression, OLP maturation arrest and apoptosis, and failure of CNS myelination.


Assuntos
Sistema Nervoso Central/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Animais , Diferenciação Celular , Sobrevivência Celular/genética , Cromatina/metabolismo , Receptor gp130 de Citocina/genética , Receptor gp130 de Citocina/metabolismo , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Fator 6 Semelhante a Kruppel , Fatores de Transcrição Kruppel-Like/genética , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Bainha de Mielina/metabolismo , Oligodendroglia/metabolismo , Proteínas Proto-Oncogênicas/genética , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais , Células-Tronco/metabolismo , alfa Carioferinas/metabolismo
3.
Mult Scler ; 24(4): 449-458, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-28273783

RESUMO

BACKGROUND: Exosomes are small extracellular vesicles that provide cell-to-cell communication and are involved in immunoregulation. OBJECTIVE: To investigate serum exosomes for the presence of myelin proteins outside the central nervous system (CNS) and their role in multiple sclerosis (MS). METHODS: Serum, cerebrospinal fluid (CSF), and peripheral blood mononuclear cell (PBMC) samples were collected from 45 patients with relapsing-remitting MS (RRMS), 30 patients with secondary progressive MS (SPMS), and 45 healthy controls. Exosomes were isolated using a polymer formulation method, and their size, concentration, and CNS myelin protein contents were measured by a nanoparticle tracking analysis, enzyme-linked immunosorbent assays, and Western blot. RESULTS: We found that exosomes expressed three major myelin proteins, myelin basic protein, proteolipid protein, and myelin oligodendrocyte glycoprotein (MOG). Exosomal content of MOG strongly correlated with disease activity and was highest in RRMS patients in relapse and in SPMS patients. Serum-derived exosomes induced proliferation of MOG-T cell receptor transgenic T cells confirming that serum exosomes maintained MOG immunogenicity. CONCLUSION: Exosomes isolated outside CNS tissue expressed myelin proteins, and the presence of MOG correlated strongly with disease activity. We conclude that exosomes might enhance and/or perpetuate anti-myelin immune reactions in MS and may provide novel markers of disease activity.


Assuntos
Exossomos/metabolismo , Esclerose Múltipla/sangue , Proteína Básica da Mielina/sangue , Glicoproteína Mielina-Oligodendrócito/sangue , Adulto , Feminino , Humanos , Leucócitos Mononucleares/metabolismo , Masculino , Pessoa de Meia-Idade , Esclerose Múltipla/imunologia , Esclerose Múltipla Recidivante-Remitente/sangue
4.
Brain ; 140(2): 399-413, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-28007993

RESUMO

Astrocytes are key players in the pathology of multiple sclerosis and can assume beneficial and detrimental roles during lesion development. The triggers and timing of the different astroglial responses in acute lesions remain unclear. Astrocytes in acute multiple sclerosis lesions have been shown previously to contain myelin debris, although its significance has not been examined. We hypothesized that myelin phagocytosis by astrocytes is an early event during lesion formation and leads to astroglial immune responses. We examined multiple sclerosis lesions and other central nervous system pathologies with prominent myelin injury, namely, progressive multifocal leukoencephalopathy, metachromatic leukodystrophy and subacute infarct. In all conditions, we found that myelin debris was present in most astrocytes at sites of acute myelin breakdown, indicating that astroglial myelin phagocytosis is an early and prominent feature. Functionally, myelin debris was taken up by astrocytes through receptor-mediated endocytosis and resulted in astroglial NF-κB activation and secretion of chemokines. These in vitro results in rats were validated in human disease where myelin-positive hypertrophic astrocytes showed increased nuclear localization of NF-κB and elevated chemokine expression compared to myelin-negative, reactive astrocytes. Thus, our data suggest that myelin uptake is an early response of astrocytes in diseases with prominent myelin injury that results in recruitment of immune cells. This first line response of astrocytes to myelin injury may exert beneficial or detrimental effects on the lesion pathology, depending on the inflammatory context. Modulating this response might be of therapeutic relevance in multiple sclerosis and other demyelinating conditions.


Assuntos
Astrócitos/metabolismo , Doenças Autoimunes Desmielinizantes do Sistema Nervoso Central/patologia , Bainha de Mielina/metabolismo , Bainha de Mielina/patologia , Fagocitose/fisiologia , Adulto , Idoso , Animais , Animais Recém-Nascidos , Astrócitos/patologia , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Pré-Escolar , Cultura , Citocinas/metabolismo , Endocitose/efeitos dos fármacos , Endocitose/fisiologia , Feminino , Humanos , Hidrazonas/farmacologia , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Pessoa de Meia-Idade , Ratos , Ratos Sprague-Dawley , Acidente Vascular Cerebral/patologia , Fatores de Tempo , Fator de Crescimento Transformador beta/farmacologia
5.
J Neurosci Res ; 95(10): 1984-1992, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28480981

RESUMO

A lot of available data on lipid immunology in multiple sclerosis (MS) have been derived from studies using synthetic lipids, therefore the role of lipids in the immunopathogenesis of MS remains poorly defined. The present study on the lipid response in MS was performed on native lipids from autopsied brain tissue. For this, lipid fractions (n = 9) were prepared from MS (n = 3) and control (n = 2) white matter according to the Folch procedure and were characterized depending on their solubility in chloroform/methanol. TLC showed that, in brain from MS cases, neutral lipids were rich in cholesterol and cholesterol esters while lipids from control brains displayed a predominance of phospholipids. MS serum IgG and IgM were found to bind to MS brain lipid fractions with a higher efficacy (p < 0.05) than the control serum. F(ab)2 fractionation revealed that MS serum IgG binding depended on a specific antibody-type of recognition. Pre-adsorption of serum with cholesterol, galactocerebrosides, sulfitides, and phosphatidylinositol prior to ELISA with MS brain lipids, showed that cholesterol diminished IgG and IgM binding up to 70%. Experiments with synthetic lipids confirmed the predominance of cholesterol binding by MS serum. Our results demonstrate that IgG and IgM fractions from MS serum specifically and predominantly recognize native cholesterol and cholesterol esters isolated from the brain tissue of patients with MS. © 2017 Wiley Periodicals, Inc.


Assuntos
Autoanticorpos/imunologia , Autoantígenos/imunologia , Encéfalo/imunologia , Colesterol/imunologia , Esclerose Múltipla Recidivante-Remitente/imunologia , Adulto , Autoanticorpos/sangue , Feminino , Humanos , Lipídeos/imunologia , Masculino , Pessoa de Meia-Idade , Esclerose Múltipla Recidivante-Remitente/sangue
6.
Brain ; 138(Pt 6): 1548-67, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25805644

RESUMO

In inflammatory central nervous system conditions such as multiple sclerosis, breakdown of the blood-brain barrier is a key event in lesion pathogenesis, predisposing to oedema, excitotoxicity, and ingress of plasma proteins and inflammatory cells. Recently, we showed that reactive astrocytes drive blood-brain barrier opening, via production of vascular endothelial growth factor A (VEGFA). Here, we now identify thymidine phosphorylase (TYMP; previously known as endothelial cell growth factor 1, ECGF1) as a second key astrocyte-derived permeability factor, which interacts with VEGFA to induce blood-brain barrier disruption. The two are co-induced NFκB1-dependently in human astrocytes by the cytokine interleukin 1 beta (IL1B), and inactivation of Vegfa in vivo potentiates TYMP induction. In human central nervous system microvascular endothelial cells, VEGFA and the TYMP product 2-deoxy-d-ribose cooperatively repress tight junction proteins, driving permeability. Notably, this response represents part of a wider pattern of endothelial plasticity: 2-deoxy-d-ribose and VEGFA produce transcriptional programs encompassing angiogenic and permeability genes, and together regulate a third unique cohort. Functionally, each promotes proliferation and viability, and they cooperatively drive motility and angiogenesis. Importantly, introduction of either into mouse cortex promotes blood-brain barrier breakdown, and together they induce severe barrier disruption. In the multiple sclerosis model experimental autoimmune encephalitis, TYMP and VEGFA co-localize to reactive astrocytes, and correlate with blood-brain barrier permeability. Critically, blockade of either reduces neurologic deficit, blood-brain barrier disruption and pathology, and inhibiting both in combination enhances tissue preservation. Suggesting importance in human disease, TYMP and VEGFA both localize to reactive astrocytes in multiple sclerosis lesion samples. Collectively, these data identify TYMP as an astrocyte-derived permeability factor, and suggest TYMP and VEGFA together promote blood-brain barrier breakdown.


Assuntos
Astrócitos/metabolismo , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Timidina Fosforilase/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Barreira Hematoencefálica/fisiopatologia , Células Cultivadas , Córtex Cerebral/efeitos dos fármacos , Desoxirribose/fisiologia , Encefalomielite Autoimune Experimental/metabolismo , Encefalomielite Autoimune Experimental/patologia , Encefalomielite Autoimune Experimental/fisiopatologia , Endotélio Vascular/metabolismo , Humanos , Interleucina-1beta/farmacologia , Camundongos , Camundongos Transgênicos , Esclerose Múltipla/metabolismo , Esclerose Múltipla/patologia , Esclerose Múltipla/fisiopatologia , Timidina Fosforilase/antagonistas & inibidores , Timidina Fosforilase/farmacologia , Ativação Transcricional/efeitos dos fármacos , Ativação Transcricional/fisiologia , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator A de Crescimento do Endotélio Vascular/farmacologia
7.
J Neurosci ; 34(25): 8646-58, 2014 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-24948818

RESUMO

The CNS is considered an immune privileged site because its repertoire of highly immunogenic molecules remains unseen by the immune system under normal conditions. However, the mechanism underlying the inhibition of immune reactions within the CNS environment is not known, particularly in regions containing myelin, which contains several potent proteins and lipids that are invariably recognized as foreign by immune system cells. Sulfatides constitute a major component of myelin glycolipids and are known to be capable of raising an immune response. In this study, the effect of sulfatides on mouse T cell function and differentiation was analyzed in vitro and in vivo. We found profound inhibition of sulfatide-dependent T cell proliferation which was particularly pronounced in naive T helper (Th) cells. The inhibitory effect of sulfatides on T cell function was CD1d-independent and was not related to apoptosis or necrosis but did involve the induction of anergy as confirmed by the upregulation of early growth response 2 transcription factor. A glycolipid 3-sulfate group was essential for the T cell suppression, and the T cell inhibition was galectin-4-dependent. Sulfatide stimulation in vitro led to prominent suppression of Th17 differentiation, and this was related to a decrease in susceptibility to disease in a mouse model of multiple sclerosis, experimental autoimmune encephalomyelitis. Thus, we have defined a novel mechanism of negative regulation of T cell function by endogenous brain-derived glycolipids, a family of molecules traditionally deemphasized in favor of myelin proteins in studies of CNS autoimmunity.


Assuntos
Encéfalo/patologia , Doenças Autoimunes Desmielinizantes do Sistema Nervoso Central/patologia , Doenças Autoimunes Desmielinizantes do Sistema Nervoso Central/prevenção & controle , Glicolipídeos/fisiologia , Linfócitos T Auxiliares-Indutores/patologia , Animais , Encéfalo/imunologia , Diferenciação Celular/fisiologia , Células Cultivadas , Doenças Autoimunes Desmielinizantes do Sistema Nervoso Central/imunologia , Feminino , Glicolipídeos/administração & dosagem , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Linfócitos T Auxiliares-Indutores/imunologia
8.
Eur J Immunol ; 44(10): 3026-44, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25092109

RESUMO

Heat shock proteins (Hsps) interact with the immune system and have been shown to contribute to immunoregulation. As efficient chaperones, Hsps bind many peptides and these complexes have many yet-to-be-clarified functions. We have shown that Hsp70 is complexed within the mouse CNS with peptide CLAFHDISPQAPTHFLVIPK derived from histidine triad nucleotide-binding protein-1 (HINT138₋57/Hsp70). Only this complex, in contrast to other peptides complexed with Hsp70, was able to prevent experimental autoimmune encephalomyelitis (EAE) by induction of immunoregulatory mechanisms dependent on NK cells. Pretreatment of proteolipid protein peptide 139₋151(PLP139₋151) sensitized SJL/J mice with HINT138₋57/Hsp70 prevented the development of EAE, suppressed PLP139₋151-induced T-cell proliferation, and blocked secretion of IL-17. HINT138₋57 /Hsp70 stimulation of NK cells depended on synergistic activation of two NK-cell receptors, CD94 and NKG2D. NK cells with depleted CD94 or with blocked NKG2D did not inhibit PLP139₋151-induced spleen cell (SC) proliferation. The HINT138₋57/Hsp70 complex enhanced surface expression of the NKG2D ligand-H60. Downstream signaling of CD94 and NKG2D converged at the adaptor proteins DAP10 and DAP12, and in response to HINT138₋57 /Hsp70 stimulation, expression of DAP10 and DAP12 was significantly increased in NK cells. Thus, we have shown that the HINT138₋57 /Hsp70 complex affects NK-cell function by enhancing NK-cell-dependent immunoregulation in the EAE model of autoimmune demyelination.


Assuntos
Encefalomielite Autoimune Experimental/imunologia , Proteínas de Choque Térmico HSP70/imunologia , Células Matadoras Naturais/imunologia , Proteínas do Tecido Nervoso/imunologia , Neuroimunomodulação/imunologia , Animais , Western Blotting , Separação Celular , Técnicas de Cocultura , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Ativação Linfocitária/imunologia , Espectrometria de Massas , Camundongos , Peptídeos/imunologia , Ligação Proteica , Reação em Cadeia da Polimerase em Tempo Real
9.
Glia ; 62(4): 580-91, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24470341

RESUMO

The oligodendrocyte (OL), the myelinating cell of the central nervous system, undergoes dramatic changes in the organization of its cytoskeleton as it differentiates from a precursor (oligodendrocyte precursor cells) to a myelin-forming cell. These changes include an increase in its branching cell processes, a phenomenon necessary for OL to myelinate multiple axon segments. We have previously shown that levels and activity of non-muscle myosin II (NMII), a regulator of cytoskeletal contractility, decrease as a function of differentiation and that inhibition of NMII increases branching and myelination of OL in coculture with neurons. We have also found that mixed glial cell cultures derived from NMIIB knockout mice display an increase in mature myelin basic protein-expressing OL compared with wild-type cultures. We have now extended our studies to investigate the role of NMIIB ablation on myelin repair following focal demyelination by lysolecithin. To this end, we generated an oligodendrocyte-specific inducible knockout model using a Plp-driven promoter in combination with a temporally activated CRE-ER fusion protein. Our data indicate that conditional ablation of NMII in adult mouse brain, expedites lesion resolution and remyelination by Plp+ oligodendrocyte-lineage cells when compared with that observed in control brains. Taken together, these data validate the function of NMII as that of a negative regulator of OL myelination in vivo and provide a novel target for promoting myelin repair in conditions such as multiple sclerosis.


Assuntos
Doenças Autoimunes Desmielinizantes do Sistema Nervoso Central/fisiopatologia , Regeneração Nervosa/fisiologia , Miosina não Muscular Tipo IIB/deficiência , Animais , Antígenos/metabolismo , Proteínas Relacionadas à Autofagia , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Corpo Caloso/patologia , Doenças Autoimunes Desmielinizantes do Sistema Nervoso Central/genética , Doenças Autoimunes Desmielinizantes do Sistema Nervoso Central/patologia , Modelos Animais de Doenças , Proteína Glial Fibrilar Ácida/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Luminescentes/genética , Lisofosfatidilcolinas , Camundongos , Camundongos Transgênicos , Proteína Básica da Mielina/metabolismo , Proteína Proteolipídica de Mielina/genética , Proteína Proteolipídica de Mielina/metabolismo , Bainha de Mielina/patologia , Proteínas do Tecido Nervoso/metabolismo , Miosina não Muscular Tipo IIB/genética , Fator de Transcrição 2 de Oligodendrócitos , Oligodendroglia/patologia , Proteoglicanas/metabolismo
10.
Nature ; 451(7182): 1076-81, 2008 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-18278032

RESUMO

Understanding the neuropathology of multiple sclerosis (MS) is essential for improved therapies. Therefore, identification of targets specific to pathological types of MS may have therapeutic benefits. Here we identify, by laser-capture microdissection and proteomics, proteins unique to three major types of MS lesions: acute plaque, chronic active plaque and chronic plaque. Comparative proteomic profiles identified tissue factor and protein C inhibitor within chronic active plaque samples, suggesting dysregulation of molecules associated with coagulation. In vivo administration of hirudin or recombinant activated protein C reduced disease severity in experimental autoimmune encephalomyelitis and suppressed Th1 and Th17 cytokines in astrocytes and immune cells. Administration of mutant forms of recombinant activated protein C showed that both its anticoagulant and its signalling functions were essential for optimal amelioration of experimental autoimmune encephalomyelitis. A proteomic approach illuminated potential therapeutic targets selective for specific pathological stages of MS and implicated participation of the coagulation cascade.


Assuntos
Perfilação da Expressão Gênica , Esclerose Múltipla/metabolismo , Esclerose Múltipla/patologia , Proteômica , Adulto , Animais , Coagulação Sanguínea , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/metabolismo , Encefalomielite Autoimune Experimental/patologia , Feminino , Humanos , Inflamação/metabolismo , Inflamação/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Esclerose Múltipla/classificação , Esclerose Múltipla/tratamento farmacológico , Proteína C/genética , Proteína C/metabolismo , Proteína C/farmacologia , Células Th1/imunologia , Células Th2/imunologia , Trombina/antagonistas & inibidores , Trombina/metabolismo
11.
Glia ; 61(4): 453-65, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23322421

RESUMO

Among the constituent cell types of the multiple sclerosis (MS) plaque, the astrocyte has been the least considered as a player in the pathogenesis of the lesion. Traditionally, it has been assigned a secondary scarring role with little or no role in lesion formation or repair. However, the recent upsurge of interest in the demyelinating condition neuromyelitis optica (NMO) has resulted in NMO being identified as the first disease of myelin in which primary damage to astrocytes, resulting from a humoral immune response that forms against the water channel aquaporin-4, has been documented. This finding in NMO prompted us to re-examine data and material from cases of MS displaying active lesions. Our reappraisal revealed unambiguous early damage to perivascular astrocyte end-feet and to hypertrophic astrocytes in the adjacent parenchyma, but whether this was a primary event was difficult to evaluate due to concomitant edema and inflammation in these acute lesions. The astrocyte damage was long-lasting since resolving lesions displaying remyelination also showed defects in the integrity of the astrocytic covering around blood vessels. Analysis of our findings and of the astrocytic literature supports multiple roles for the astrocyte in the evolution of changes encountered in MS depending upon lesion stage and lesion topography. At variance with the somewhat inhibitory role of the astrocyte is the abundant and growing evidence for this cell to actively participate in both lesion development and repair. We propose that the unequivocal selective early involvement of the astrocyte in MS lesions may have therapeutic relevance


Assuntos
Astrócitos/patologia , Esclerose Múltipla/patologia , Animais , Barreira Hematoencefálica/patologia , Encéfalo/patologia , Doenças Desmielinizantes/epidemiologia , Doenças Desmielinizantes/patologia , Humanos , Esclerose Múltipla/epidemiologia , Neuromielite Óptica/epidemiologia , Neuromielite Óptica/patologia , Medula Espinal/patologia
12.
J Immunol ; 187(3): 1129-41, 2011 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-21709156

RESUMO

Current therapies for multiple sclerosis target inflammation but do not directly address oligodendrocyte protection or myelin repair. The gp130 family cytokines ciliary neurotrophic factor, leukemia inhibitory factor, and IL-11 have been identified as oligodendrocyte growth factors, and IL-11 is also strongly immunoregulatory, but their underlying mechanisms of action are incompletely characterized. In this study, we demonstrate that these effects of IL-11 are mediated via differential regulation of apoptosis in oligodendrocytes versus Ag-presenting dendritic cells (DCs), and are dependent on lineage-specific activity of the transcription factors Stat1 versus Stat3. Focal demyelinating lesions induced in cerebral cortices of IL-11Rα(-/-) mice using stereotactic microinjection of lysolecithin were larger than in controls, and remyelination was delayed. In IL-11Rα(-/-) mice, lesions displayed extensive oligodendrocyte loss and axonal transection, and increased infiltration by inflammatory cells including CD11c(+) DCs, CD3(+) lymphocytes, and CD11b(+) phagocytes. In oligodendrocyte progenitor cell (OPC) cultures, IL-11 restricted caspase 9 activation and apoptosis, and it increased myelination in OPC-neuron cocultures. Importantly, siRNA inhibition of Stat1 enhanced the antiapoptotic effects of IL-11 on OPCs, but IL-11 induced apoptosis in the presence of Stat3 silencing. In contrast, IL-11 augmented caspase activation and apoptosis in cultures of CD11c(+) DCs, but not in CD11b(+) or CD3(+) cells. Inhibition of Stat3 exacerbated the proapoptotic effects of IL-11 on DCs, whereas they were ablated in Stat1(-/-) cultures. Collectively, these findings reveal novel mechanisms underlying the actions of a neuroprotective and immunoregulatory member of the gp130 cytokine family, suggesting avenues to enhance oligodendrocyte viability and restrict CNS inflammation in multiple sclerosis.


Assuntos
Proteínas Reguladoras de Apoptose/fisiologia , Interleucina-11/uso terapêutico , Fármacos Neuroprotetores/uso terapêutico , Fator de Transcrição STAT1/fisiologia , Fator de Transcrição STAT3/fisiologia , Animais , Linhagem da Célula/genética , Linhagem da Célula/imunologia , Sobrevivência Celular/genética , Sobrevivência Celular/imunologia , Células Cultivadas , Técnicas de Cocultura , Doenças Desmielinizantes/imunologia , Doenças Desmielinizantes/patologia , Doenças Desmielinizantes/terapia , Células Dendríticas/imunologia , Células Dendríticas/patologia , Modelos Animais de Doenças , Marcação de Genes/métodos , Interleucina-11/deficiência , Interleucina-11/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Esclerose Múltipla/imunologia , Esclerose Múltipla/patologia , Esclerose Múltipla/terapia , Oligodendroglia/imunologia , Oligodendroglia/metabolismo , Oligodendroglia/patologia , Ratos , Ratos Sprague-Dawley , Células-Tronco/imunologia , Células-Tronco/metabolismo , Células-Tronco/patologia
13.
J Neurosci Res ; 90(10): 1941-8, 2012 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-22715030

RESUMO

Heat shock proteins (HSPs) are stress-responsive proteins that serve as important molecules contributing to cellular "protein triage." We and others have reported an increase of selected HSPs in multiple sclerosis (MS) lesions. However, the exact expression pattern of HSP family genes in MS is not known. The aim of our research was to assess global transcriptional changes of all gene members of the HSP families within MS lesions and associated normal-appearing white matter (NAWM). To this end, we used laser capture microdissection (LCM) to isolate defined regions of chronic-active MS lesions (n = 5), one of the most common types of MS lesions. To identify changes in HSP genes in relation to different areas of the plaque, we used genome-wide microarray analysis. We detected a significant change in the transcriptional profile of the demyelinated region compared with NAWM. In particular, overall expression of different HSP genes was upregulated in different areas of chronic-active lesion. These changes were linked to an upregulation of heat shock factor 4 (HSF4). This is the first global analysis of transcriptional changes in HSPs in the central nervous system during MS. The results support a relationship between HSP activation and lesion activity.


Assuntos
Proteínas de Choque Térmico/genética , Esclerose Múltipla/genética , Esclerose Múltipla/metabolismo , Idoso , Autopsia , Encéfalo/patologia , Biologia Computacional , Proteínas de Ligação a DNA/biossíntese , Doenças Desmielinizantes/genética , Feminino , Perfilação da Expressão Gênica , Fatores de Transcrição de Choque Térmico , Humanos , Microdissecção e Captura a Laser , Masculino , Análise em Microsséries , Esclerose Múltipla/patologia , RNA/genética , Fatores de Transcrição/biossíntese , Ativação Transcricional/genética , Regulação para Cima/fisiologia
14.
Proc Natl Acad Sci U S A ; 106(45): 19162-7, 2009 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-19855010

RESUMO

In the developing CNS, Notch1 and its ligand, Jagged1, regulate oligodendrocyte differentiation and myelin formation, but their role in repair of demyelinating lesions in diseases such as multiple sclerosis remains unresolved. To address this question, we generated a mouse model in which we targeted Notch1 inactivation to oligodendrocyte progenitor cells (OPCs) using Olig1Cre and a floxed Notch1 allele, Notch1(12f). During CNS development, OPC differentiation was potentiated in Olig1Cre:Notch1(12f/12f) mice. Importantly, in adults, remyelination of demyelinating lesions was also accelerated, at the expense of proliferation within the progenitor population. Experiments in vitro confirmed that Notch1 signaling was permissive for OPC expansion but inhibited differentiation and myelin formation. These studies also revealed that astrocytes exposed to TGF-beta1 restricted OPC maturation via Jagged1-Notch1 signaling. These data suggest that Notch1 signaling is one of the mechanisms regulating OPC differentiation during CNS remyelination. Thus, Notch1 may represent a potential therapeutical avenue for lesion repair in demyelinating disease.


Assuntos
Diferenciação Celular/fisiologia , Sistema Nervoso Central/citologia , Bainha de Mielina/fisiologia , Oligodendroglia/fisiologia , Receptor Notch1/metabolismo , Transdução de Sinais/fisiologia , Animais , Proteínas de Ligação ao Cálcio/metabolismo , Imuno-Histoquímica , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteína Jagged-1 , Proteínas de Membrana/metabolismo , Camundongos , Oligodendroglia/citologia , Proteínas Serrate-Jagged
15.
Proc Natl Acad Sci U S A ; 106(35): 14948-53, 2009 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-19706421

RESUMO

The renin-angiotensin-aldosterone system (RAAS) is a major regulator of blood pressure. The octapeptide angiotensin II (AII) is proteolytically processed from the decapeptide AI by angiotensin-converting enzyme (ACE), and then acts via angiotensin type 1 and type 2 receptors (AT1R and AT2R). Inhibitors of ACE and antagonists of the AT1R are used in the treatment of hypertension, myocardial infarction, and stroke. We now show that the RAAS also plays a major role in autoimmunity, exemplified by multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE). Using proteomics, we observed that RAAS is up-regulated in brain lesions of MS. AT1R was induced in myelin-specific CD4+ T cells and monocytes during autoimmune neuroinflammation. Blocking AII production with ACE inhibitors or inhibiting AII signaling with AT1R blockers suppressed autoreactive TH1 and TH17 cells and promoted antigen-specific CD4+FoxP3+ regulatory T cells (Treg cells) with inhibition of the canonical NF-kappaB1 transcription factor complex and activation of the alternative NF-kappaB2 pathway. Treatment with ACE inhibitors induces abundant CD4+FoxP3+ T cells with sufficient potency to reverse paralytic EAE. Modulation of the RAAS with inexpensive, safe pharmaceuticals used by millions worldwide is an attractive therapeutic strategy for application to human autoimmune diseases.


Assuntos
Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , Encefalomielite Autoimune Experimental/tratamento farmacológico , Linfócitos T Auxiliares-Indutores/imunologia , Linfócitos T Reguladores/imunologia , Animais , Encefalomielite Autoimune Experimental/enzimologia , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/patologia , Feminino , Fatores de Transcrição Forkhead/imunologia , Humanos , Interleucina-17/imunologia , Camundongos , Peptidil Dipeptidase A/metabolismo , Receptor Tipo 1 de Angiotensina/genética , Receptor Tipo 1 de Angiotensina/metabolismo , Receptor Tipo 2 de Angiotensina/metabolismo , Linfócitos T Auxiliares-Indutores/efeitos dos fármacos , Linfócitos T Auxiliares-Indutores/enzimologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/enzimologia
16.
Nat Med ; 8(10): 1115-21, 2002 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-12357247

RESUMO

During mammalian central nervous system (CNS) development, contact-mediated activation of Notch1 receptors on oligodendrocyte precursors by the ligand Jagged1 induces Hes5, which inhibits maturation of these cells. Here we tested whether the Notch pathway is re-expressed in the adult CNS in multiple sclerosis (MS), an inflammatory demyelinating disease in which remyelination is typically limited. We found that transforming growth factor-beta 1 (TGF-beta 1), a cytokine upregulated in MS, specifically re-induced Jagged1 in primary cultures of human astrocytes. Within and around active MS plaques lacking remyelination, Jagged1 was expressed at high levels by hypertrophic astrocytes, whereas Notch1 and Hes5 localized to cells with an immature oligodendrocyte phenotype, and TGF-beta 1 was associated with perivascular extracellular matrix in the same areas. In contrast, there was negligible Jagged1 expression in remyelinated lesions. Experiments in vitro showed that Jagged1 signaling inhibited process outgrowth from primary human oligodendrocytes. These data are the first to implicate the Notch pathway in the limited remyelination in MS. Thus, Notch may represent a potential target for therapeutic intervention in this disease.


Assuntos
Proteínas de Membrana/metabolismo , Esclerose Múltipla/fisiopatologia , Oligodendroglia/fisiologia , Proteínas/metabolismo , Fatores de Transcrição , Animais , Astrócitos/citologia , Astrócitos/metabolismo , Proteínas de Ligação ao Cálcio , Células Cultivadas , Humanos , Peptídeos e Proteínas de Sinalização Intercelular , Proteína Jagged-1 , Esclerose Múltipla/patologia , Bainha de Mielina/metabolismo , Oligodendroglia/citologia , Análise de Sequência com Séries de Oligonucleotídeos , Receptor Notch1 , Receptores de Superfície Celular/metabolismo , Proteínas Serrate-Jagged , Transdução de Sinais/fisiologia , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta1
17.
Nat Med ; 8(5): 500-8, 2002 May.
Artigo em Inglês | MEDLINE | ID: mdl-11984595

RESUMO

Microarray analysis of multiple sclerosis (MS) lesions obtained at autopsy revealed increased transcripts of genes encoding inflammatory cytokines, particularly interleukin-6 and -17, interferon-gamma and associated downstream pathways. Comparison of two poles of MS pathology--acute lesions with inflammation versus 'silent' lesions without inflammation--revealed differentially transcribed genes. Some products of these genes were chosen as targets for therapy of experimental autoimmune encephalomyelitis (EAE) in mice. Granulocyte colony-stimulating factor is upregulated in acute, but not in chronic, MS lesions, and the effect on ameliorating EAE is more pronounced in the acute phase, in contrast to knocking out the immunoglobulin Fc receptor common gamma chain where the effect is greatest on chronic disease. These results in EAE corroborate the microarray studies on MS lesions. Large-scale analysis of transcripts in MS lesions elucidates new aspects of pathology and opens possibilities for therapy.


Assuntos
Encefalomielite Autoimune Experimental/genética , Interferon gama/genética , Interleucina-17/genética , Interleucina-6/genética , Esclerose Múltipla/genética , Análise de Sequência com Séries de Oligonucleotídeos , Doença Aguda , Animais , Autopsia , Doença Crônica , Encefalomielite Autoimune Experimental/patologia , Feminino , Fator Estimulador de Colônias de Granulócitos/fisiologia , Humanos , Inflamação/genética , Inflamação/patologia , Camundongos , Camundongos Endogâmicos C57BL , Esclerose Múltipla/patologia , Receptores Fc/fisiologia , Reprodutibilidade dos Testes , Transcrição Gênica
18.
Artigo em Inglês | MEDLINE | ID: mdl-34385287

RESUMO

BACKGROUND AND OBJECTIVES: To investigate the total circular RNA (circRNA) profile in patients with relapsing-remitting multiple sclerosis (RRMS) and healthy controls (HCs). METHODS: Hybridization microarray was used to define the circRNA profile in peripheral blood mononuclear cells (PBMCs) from 20 untreated patients with RRMS (10 in relapse and 10 in remission) and 10 HCs. We analyzed close to 14,000 individual circRNAs per sample. The discovery set data were validated using quantitative reverse transcription-PCR with an independent cohort of 47 patients with RRMS (19 in relapse and 28 in remission) and 27 HCs. RESULTS: Microarray analysis revealed 914 transcripts to be differentially expressed between patients with RRMS in relapse and HCs (p < 0.05). We validated 3 circRNAs from 5 showing highest levels of differential expression in the RRMS relapse vs HC group: hsa_circRNA_101348, hsa_circRNA_102611, and hsa_circRNA_104361. Their expression was significantly increased during relapse in RRMS (p = 0.0002, FC = 2.9; p = 0.01, FC = 1.6; and p = 0.001, FC = 1.5, respectively) and in patients showing gadolinium enhancement on brain MRI (hsa_circRNA_101348, p = 0.0039, FC = 2.4; hsa_circRNA_104361, p = 0.029, FC = 1.7). Bioinformatic analysis revealed 15 microRNAs interacting with these circRNAs in a complementary manner and led to the discovery and validation of 3 protein-coding RNAs upregulated in patients with RRMS during relapse. Two of these, AK2 and IKZF3, have previously been implicated in B-cell function. DISCUSSION: circRNAs display a distinct profile in PBMCs from patients with RRMS, and our results may implicate circRNA in the known disturbed B-cell activity in RRMS and thus represent a novel biomarker for monitoring relapse activity.


Assuntos
Linfócitos B/imunologia , Esclerose Múltipla Recidivante-Remitente/sangue , Esclerose Múltipla Recidivante-Remitente/imunologia , RNA Circular/sangue , RNA Circular/imunologia , Adulto , Encéfalo/diagnóstico por imagem , Feminino , Humanos , Leucócitos Mononucleares/imunologia , Masculino , Esclerose Múltipla Recidivante-Remitente/diagnóstico , Sensibilidade e Especificidade
19.
Neuron ; 49(1): 67-79, 2006 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-16387640

RESUMO

Endocannabinoids are released after brain injury and believed to attenuate neuronal damage by binding to CB(1) receptors and protecting against excitotoxicity. Such excitotoxic brain lesions initially result in primary destruction of brain parenchyma, which attracts macrophages and microglia. These inflammatory cells release toxic cytokines and free radicals, resulting in secondary neuronal damage. In this study, we show that the endocannabinoid system is highly activated during CNS inflammation and that the endocannabinoid anandamide (AEA) protects neurons from inflammatory damage by CB(1/2) receptor-mediated rapid induction of mitogen-activated protein kinase phosphatase-1 (MKP-1) in microglial cells associated with histone H3 phoshorylation of the mkp-1 gene sequence. As a result, AEA-induced rapid MKP-1 expression switches off MAPK signal transduction in microglial cells activated by stimulation of pattern recognition receptors. The release of AEA in injured CNS tissue might therefore represent a new mechanism of neuro-immune communication during CNS injury, which controls and limits immune response after primary CNS damage.


Assuntos
Ácidos Araquidônicos/metabolismo , Moduladores de Receptores de Canabinoides/metabolismo , Proteínas de Ciclo Celular/biossíntese , Encefalite/fisiopatologia , Endocanabinoides , Proteínas Imediatamente Precoces/biossíntese , Microglia/metabolismo , Fármacos Neuroprotetores/metabolismo , Fosfoproteínas Fosfatases/biossíntese , Proteínas Tirosina Fosfatases/biossíntese , Adulto , Animais , Ácidos Araquidônicos/farmacologia , Moduladores de Receptores de Canabinoides/farmacologia , Sobrevivência Celular , Células Cultivadas , Fosfatase 1 de Especificidade Dupla , Encefalite/patologia , Encefalite/prevenção & controle , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , Pessoa de Meia-Idade , Esclerose Múltipla/metabolismo , Esclerose Múltipla/patologia , Neurônios/patologia , Fármacos Neuroprotetores/farmacologia , Óxido Nítrico/antagonistas & inibidores , Óxido Nítrico Sintase/antagonistas & inibidores , Fosforilação/efeitos dos fármacos , Alcamidas Poli-Insaturadas , Proteína Fosfatase 1 , Ratos , Ratos Wistar , Receptores de Canabinoides/metabolismo , Distribuição Tecidual
20.
J Neurosci ; 29(24): 7743-52, 2009 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-19535586

RESUMO

Astrocytes are coupled via gap junctions (GJs) comprising connexin 43 (Cx43) (Gja1) and Cx30 (Gjb6), which facilitate intercellular exchange of ions. Astrocyte connexins also form heterotypic GJs with oligodendrocytic somata and lamellae. Loss of oligodendrocyte gap junctions results in oligodendrocyte and myelin pathology. However, whether loss of astrocyte GJs affects oligodendrocytes and myelin is not known. To address this question, mice with astrocyte-targeted deletion of Cx43 and global loss of Cx30 [double knock-out (dKO)] were studied using Western blotting, immunohistochemistry, electron microscopy, and functional assays. Commencing around postnatal day 23 and persisting into old age, we found widespread pathology of white matter tracts comprising vacuolated oligodendrocytes and intramyelinic edema. In contrast, gray matter pathology was restricted to the CA1 region of the hippocampus, and consisted of edematous astrocytes. No differences were observed in synaptic density or total NeuN(+) cells in the hippocampus, or olig2(+) cells in the corpus callosum. However, in dKO mice, fewer CC1-positive mature oligodendrocytes were detected, and Western blotting indicated reduced myelin basic protein. Pathology was not noted in mice expressing a single allele of either Cx43 or Cx30. When compared with single connexin knock-outs, dKO mice were impaired in sensorimotor (rotarod, balance beam assays) and spatial memory tasks (object recognition assays). We conclude that loss of astrocytic GJs can result in white matter pathology that has functional consequences.


Assuntos
Astrócitos/metabolismo , Conexina 43/deficiência , Conexinas/deficiência , Doenças Desmielinizantes/patologia , Hipocampo/patologia , Fenótipo , Animais , Animais Recém-Nascidos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Proteínas de Ligação ao Cálcio , Proliferação de Células , Conexina 30 , Proteínas de Ligação a DNA/metabolismo , Doenças Desmielinizantes/complicações , Doenças Desmielinizantes/genética , Modelos Animais de Doenças , Junções Comunicantes/patologia , Proteína Glial Fibrilar Ácida/genética , Marcação In Situ das Extremidades Cortadas/métodos , Transtornos da Memória/genética , Transtornos da Memória/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas dos Microfilamentos , Microscopia Eletrônica/métodos , Proteína Básica da Mielina/metabolismo , Bainha de Mielina/patologia , Proteínas do Tecido Nervoso/metabolismo , Fator de Transcrição 2 de Oligodendrócitos
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