RESUMEN
Multiple sclerosis (MS) is a multifocal demyelinating disease of the central nervous system (CNS) leading to the progressive destruction of the myelin sheath surrounding axons. It can present with variable clinical and pathological manifestations, which might reflect the involvement of distinct pathogenic processes. Although the mechanisms leading to the development of the disease are not fully understood, numerous evidences indicate that MS is an autoimmune disease, the initiation and progression of which are dependent on an autoimmune response against myelin antigens. In addition, genetic susceptibility and environmental triggers likely contribute to the initiation of the disease. At this time, there is no cure for MS, but several disease-modifying therapies (DMTs) are available to control and slow down disease progression. A good number of these DMTs were identified and tested using animal models of MS referred to as experimental autoimmune encephalomyelitis (EAE). In this review, we will recapitulate the characteristics of EAE models and discuss how they help shed light on MS pathogenesis and help test new treatments for MS patients.
Asunto(s)
Encefalomielitis Autoinmune Experimental/fisiopatología , Esclerosis Múltiple/fisiopatología , Animales , Antígenos/inmunología , Linfocitos B/fisiología , Linfocitos T CD4-Positivos/fisiología , Linfocitos T CD8-positivos/fisiología , Encefalomielitis Autoinmune Experimental/inmunología , Humanos , Esclerosis Múltiple/inmunología , Proteína Básica de Mielina/fisiología , Proteína Proteolipídica de la Mielina/fisiología , Glicoproteína Mielina-Oligodendrócito/fisiologíaRESUMEN
Autoantibodies against various components of host are known to occur in leprosy. Nerve damage is the primary cause of disability associated with leprosy. The aim of this study was to detect the level of autoantibodies and lympho-proliferative response against myelin basic protein (MBP) in leprosy patients (LPs) and their correlation with clinical phenotypes of LPs. Further, probable role of molecular mimicry in nerve damage of LPs was investigated. We observed significantly high level of anti-MBP antibodies in LPs across the spectrum and a positive significant correlation between the level of anti-MBP antibodies and the number of nerves involved in LPs. We report here that 4 B cell epitopes of myelin A1 and Mycobacterium leprae proteins, 50S ribosomal L2 and lysyl tRNA synthetase are cross-reactive. Further, M. leprae sonicated antigen hyperimmunization was responsible for induction of autoantibody response in mice which could be adoptively transferred to naive mice. For the first time our findings suggest the role of molecular mimicry in nerve damage in leprosy.
Asunto(s)
Enfermedades Desmielinizantes/microbiología , Lepra/microbiología , Lisina-ARNt Ligasa/fisiología , Imitación Molecular/fisiología , Mycobacterium leprae/patogenicidad , Proteína Básica de Mielina/fisiología , Proteínas Ribosómicas/fisiología , Animales , Enfermedades Desmielinizantes/complicaciones , Enfermedades Desmielinizantes/etiología , Humanos , Lepra/complicaciones , Lepra/etiología , Ratones , Ratones Endogámicos BALB C/sangre , ConejosRESUMEN
Alzheimer's disease is a progressive neurodegenerative disorder that is characterized by extensive deposition of fibrillar amyloid-ß (Aß) in the brain. Previously, myelin basic protein (MBP) was identified to be a potent inhibitor to Aß fibril formation, and this inhibitory activity was localized to the N-terminal residues 1-64, a fragment designated MBP1. Here, we show that the modest neuronal expression of a fusion protein of the biologically active MBP1 fragment and the enhanced green fluorescent protein (MBP1-EGFP) significantly improved the performance of spatial learning memory in Tg-5xFAD mice, a model of pathologic Aß accumulation in brain. The levels of insoluble Aß and fibrillar amyloid were significantly reduced in bigenic Tg-5xFAD/Tg-MBP1-EGFP mice. Quantitative stereological analysis revealed that the reduction in amyloid was because of a reduction in the size of fibrillar plaques rather than a decrease in plaque numbers. The current findings support previous studies showing that MBP1 inhibits Aß fibril formation in vitro and demonstrate the ability of MBP1 to reduce Aß pathology and improve behavioral performance.
Asunto(s)
Enfermedad de Alzheimer/genética , Péptidos beta-Amiloides/metabolismo , Encéfalo/metabolismo , Proteína Básica de Mielina/fisiología , Agregación Patológica de Proteínas/genética , Enfermedad de Alzheimer/patología , Enfermedad de Alzheimer/psicología , Animales , Conducta , Encéfalo/patología , Modelos Animales de Enfermedad , Ratones Transgénicos , Proteína Básica de Mielina/químicaRESUMEN
Accumulating evidence indicates that the medial prefrontal cortex (mPFC) is a site of myelin and oligodendrocyte abnormalities that contribute to psychotic symptoms of schizophrenia. The development of therapeutic approaches to enhance remyelination, a regenerative process in which new myelin sheaths are formed on demyelinated axons, may be an attractive remedial strategy. Geissoschizine methyl ether (GM) in the Uncaria hook, a galenical constituent of the traditional Japanese medicine yokukansan (Yi-gan san), is one of the active components responsible for the psychotropic effects of yokukansan, though little is known about the mechanisms underlying the effects of either that medicine or GM itself. In the present study, we employed a cuprizone (CPZ)-induced demyelination model and examined the cellular changes in response to GM administration during the remyelination phase in the mPFC of adult mice. Using the mitotic marker 5-bromo-2'-deoxyuridine (BrdU), we demonstrated that CPZ treatment significantly increased the number of BrdU-positive NG2 cells, as well as microglia and mature oligodendrocytes in the mPFC. Newly formed oligodendrocytes were increased by GM administration after CPZ exposure. In addition, GM attenuated a decrease in myelin basic protein immunoreactivity caused by CPZ administration. Taken together, our findings suggest that GM administration ameliorated the myelin deficit by mature oligodendrocyte formation and remyelination in the mPFC of CPZ-fed mice. The present findings provide experimental evidence supporting the role for GM and its possible use as a remedy for schizophrenia symptoms by promoting the differentiation of progenitor cells to and myelination by oligodendrocytes.
Asunto(s)
Cuprizona/farmacología , Indoles/farmacología , Corteza Prefrontal/metabolismo , Animales , Enfermedades Desmielinizantes/inducido químicamente , Enfermedades Desmielinizantes/tratamiento farmacológico , Alcaloides Indólicos , Ratones , Proteína Básica de Mielina/fisiología , Corteza Prefrontal/efectos de los fármacos , Regeneración/efectos de los fármacosRESUMEN
Mutations in the methyl-CpG binding protein 2 gene, Mecp2, affect primarily the brain and lead to a wide range of neuropsychiatric disorders, most commonly Rett syndrome (RTT). Although the neuropathology of RTT is well understood, the cellular and molecular mechanism(s), which lead to the disease initiation and progression, has yet to be elucidated. RTT was initially attributed only to neuronal dysfunction, but our recent studies and those of others show that RTT is not exclusively neuronal but rather also involves interactions between neurons and glia. Importantly, studies have shown that MeCP2-restored astrocytes and microglia are able to attenuate the disease progression in otherwise MeCP2-null mice. Here we show that another type of glia, oligodendrocytes, and their progenitors are also involved in manifestation of specific RTT symptoms. Mice that lost MeCP2 specifically in the oligodendrocyte lineage cells, although overall normal, were more active and developed severe hindlimb clasping phenotypes. Inversely, restoration of MeCP2 in oligodendrocyte lineage cells, in otherwise MeCP2-null mice, although only mildly prolonging their lifespan, significantly improved the locomotor deficits and hindlimb clasping phenotype, both in male and female mice, and fully restored the body weight in male mice. Finally, we found that the level of some myelin-related proteins was impaired in the MeCP2-null mice. Expression of MeCP2 in oligodendrocytes of these mice only partially restored their expression, suggesting that there is a non-cell-autonomous effect by other cell types in the brains on the expression of myelin-related proteins in oligodendrocytes.
Asunto(s)
Linaje de la Célula/fisiología , Proteína 2 de Unión a Metil-CpG/genética , Oligodendroglía/patología , Síndrome de Rett/patología , Animales , Astrocitos/fisiología , Western Blotting , Oscuridad , Femenino , Fuerza de la Mano/fisiología , Miembro Posterior/fisiología , Inmunohistoquímica , Luz , Locomoción/fisiología , Masculino , Proteína 2 de Unión a Metil-CpG/fisiología , Ratones , Mutación/genética , Mutación/fisiología , Proteína Básica de Mielina/fisiología , Fenotipo , Reacción en Cadena de la PolimerasaRESUMEN
The efficacy of cannabinoids in the treatment of multiple sclerosis is widely documented; however their use is limited by psychoactivity mainly ascribed to the activation of the cannabinoid receptor CB1. Emerging findings support as alternative strategy in the treatment of neurodegenerative disorders, the application of compounds targeting the CB2 receptor, since likely unrelated to these side effects. Recently, a novel class of compounds, 1,8-naphthyridine, pyridine and quinoline derivatives have been demonstrated to show high CB2 receptor selectivity and affinity versus the CB1 receptor. Considering that the CB2 receptor is mainly expressed in cell and organs of the immune system, in this study we assessed the potential immune-modulatory effects of these compounds in activated lymphocytes isolated from MS patients with respect to healthy controls. These compounds blocked cell proliferation through a mechanism partially ascribed to the CB2 receptor, down-regulated TNF-α production and did not induce cell death. They also down-regulated Akt, Erk and NF-kB phosphorylation. Despite comparable effects observed in patients and healthy controls, these compounds, in particular, 1,8-naphthyridine and quinoline derivatives inhibited cell activation markers in MS patient derived lymphocytes more efficiently than in healthy control derived cells. Indeed, 1,8-naphthyridin-2-one derivative reduced the levels of Cox-2 in lymphocytes from patients whereas no effect was observed in control cells. Our findings suggest potential application of these drugs in neuro-inflammation, supporting further investigations of the effects of compounds in the therapy of MS, particularly on the aspects regarding activation and inflammation.
Asunto(s)
Antiinflamatorios/farmacología , Esclerosis Múltiple/tratamiento farmacológico , Naftiridinas/farmacología , Piridonas/farmacología , Quinolonas/farmacología , Receptor Cannabinoide CB2/agonistas , Antígenos CD/metabolismo , Antígenos de Diferenciación de Linfocitos T/metabolismo , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Ciclooxigenasa 2/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Femenino , Expresión Génica , Humanos , Inmunosupresores/farmacología , Molécula 1 de Adhesión Intercelular/metabolismo , Lectinas Tipo C/metabolismo , Activación de Linfocitos , Esclerosis Múltiple/inmunología , Proteína Básica de Mielina/fisiología , FN-kappa B/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptor Cannabinoide CB2/metabolismo , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Linfocitos T/metabolismo , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
The classic myelin basic protein (MBP) family of central nervous system (CNS) myelin arises from transcription start site 3 of the Golli (gene of oligodendrocyte lineage) complex and comprises splice isoforms ranging in nominal molecular mass from 14 kDa to (full-length) 21.5 kDa. We have determined here a number of distinct functional differences between the major 18.5-kDa and minor 21.5-kDa isoforms of classic MBP with respect to oligodendrocyte (OLG) proliferation. We have found that, in contrast to 18.5-kDa MBP, 21.5-kDa MBP increases proliferation of early developmental immortalized N19-OLGs by elevating the levels of phosphorylated ERK1/2 and Akt1 kinases and of ribosomal protein S6. Coculture of N2a neuronal cells with N19-OLGs transfected with the 21.5-kDa isoform (or conditioned medium from), but not the 18.5-kDa isoform, caused the N2a cells to have increased neurite outgrowth and process branching complexity. These roles were dependent on subcellular localization of 21.5-kDa MBP to the nucleus and on the exon II-encoded segment, suggesting that the nuclear localization of early minor isoforms of MBP may play a crucial role in regulating and/or initiating myelin and neuronal development in the mammalian CNS.
Asunto(s)
Membrana Celular/metabolismo , Núcleo Celular/metabolismo , Proliferación Celular , Proteína Básica de Mielina/fisiología , Neuritas/fisiología , Oligodendroglía/metabolismo , Animales , Línea Celular Transformada , Membrana Celular/química , Núcleo Celular/química , Núcleo Celular/fisiología , Técnicas de Cocultivo , Ratones , Peso Molecular , Proteína Básica de Mielina/química , Proteína Básica de Mielina/metabolismo , Neuritas/química , Oligodendroglía/química , Isoformas de Proteínas/fisiologíaRESUMEN
BACKGROUND: The myelin sheath provides electrical insulation of mechanosensory Aß-afferent fibers. Myelin-degrading matrix metalloproteinases (MMPs) damage the myelin sheath. The resulting electrical instability of Aß-fibers is believed to activate the nociceptive circuitry in Aß-fibers and initiate pain from innocuous tactile stimulation (mechanical allodynia). The precise molecular mechanisms, responsible for the development of this neuropathic pain state after nerve injury (for example, chronic constriction injury, CCI), are not well understood. METHODS AND RESULTS: Using mass spectrometry of the whole sciatic nerve proteome followed by bioinformatics analyses, we determined that the pathways, which are classified as the Infectious Disease and T-helper cell signaling, are readily activated in the nerves post-CCI. Inhibition of MMP-9/MMP-2 suppressed CCI-induced mechanical allodynia and concomitant TNF-α and IL-17A expression in nerves. MMP-9 proteolysis of myelin basic protein (MBP) generated the MBP84-104 and MBP68-86 digest peptides, which are prominent immunogenic epitopes. In agreement, the endogenous MBP69-86 epitope co-localized with MHCII and MMP-9 in Schwann cells and along the nodes of Ranvier. Administration of either the MBP84-104 or MBP68-86 peptides into the naïve nerve rapidly produced robust mechanical allodynia with a concomitant increase in T cells and MHCII-reactive cell populations at the injection site. As shown by the genome-wide expression profiling, a single intraneural MBP84-104 injection stimulated the inflammatory, immune cell trafficking, and antigen presentation pathways in the injected naïve nerves and the associated spinal cords. Both MBP84-104-induced mechanical allodynia and characteristic pathway activation were remarkably less prominent in the T cell-deficient athymic nude rats. CONCLUSIONS: These data implicate MBP as a novel mediator of pain. Furthermore, the action of MMPs expressed within 1 day post-injury is critical to the generation of tactile allodynia, neuroinflammation, and the immunodominant MBP digest peptides in nerve. These MBP peptides initiate mechanical allodynia in both a T cell-dependent and -independent manner. In the course of Wallerian degeneration, the repeated exposure of the cryptic MBP epitopes, which are normally sheltered from immunosurveillance, may induce the MBP-specific T cell clones and a self-sustaining immune reaction, which may together contribute to the transition of acute pain into a chronic neuropathic pain state.
Asunto(s)
Epítopos de Linfocito T/efectos adversos , Epítopos Inmunodominantes/efectos adversos , Proteína Básica de Mielina/fisiología , Dolor/inmunología , Subgrupos de Linfocitos T/inmunología , Secuencia de Aminoácidos , Animales , Epítopos de Linfocito T/fisiología , Femenino , Células HEK293 , Humanos , Epítopos Inmunodominantes/fisiología , Datos de Secuencia Molecular , Monitorización Inmunológica/efectos adversos , Dolor/etiología , Dolor/patología , Dimensión del Dolor/métodos , Ratas , Ratas Desnudas , Ratas Sprague-Dawley , Subgrupos de Linfocitos T/patologíaRESUMEN
Myelination of axons by oligodendrocytes (OLGs) is essential for proper saltatory nerve conduction, i.e., rapid transmission of nerve impulses. Among others, extracellular matrix (ECM) molecules, neuronal signaling, and axonal adhesion regulate the biogenesis and maintenance of myelin membranes, driven by polarized transport of myelin-specific proteins and lipids. Galectin-4, a tandem-repeat-type lectin with affinity to sulfatide and nonsialylated termini of N-glycans, has the ability to regulate adhesion of cells to ECM components and is also involved in polarized membrane trafficking. We, therefore, anticipated that galectin-4 might play a role in myelination. Here, we show that in developing postnatal rat brains galectin-4 expression is downregulated just before the onset of myelination. Intriguingly, when immature OLGs were treated with galectin-4, OLG maturation was retarded, while a subset of the immature OLGs reverted to a morphologically less complex progenitor stage, displaying concomitantly an increase in proliferation. Similarly, myelination was inhibited when galectin-4 or anti-galectin-4 antibodies were added to co-cultures of dorsal root ganglion neurons and OLGs. Neurons and OLGs were identified as a possible source of galectin-4, both in vitro and in vivo. In culture, neurons but not OLGs released galectin-4. Interestingly, in co-cultures, a reduced release of endogenous galectin-4 correlated with the onset of myelination. Moreover, galectin-4-reactive sites are transiently expressed on processes of premyelinating primary OLGs, but not on neurons. Taken together, these results identify neuronal galectin-4 as a candidate for a soluble regulator of OLG differentiation and, hence, myelination. © 2012 Wiley Periodicals, Inc.
Asunto(s)
Galectina 4/fisiología , Regulación del Desarrollo de la Expresión Génica/fisiología , Proteínas de la Mielina/metabolismo , Vaina de Mielina/metabolismo , Neuronas/metabolismo , Factores de Edad , Animales , Animales Recién Nacidos , Anticuerpos/farmacología , Encéfalo/citología , Bromodesoxiuridina/metabolismo , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/fisiología , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Técnicas de Cocultivo , Galectina 4/genética , Galectina 4/inmunología , Galectina 4/farmacología , Ganglios Espinales/citología , Gangliósidos/metabolismo , Humanos , L-Lactato Deshidrogenasa/metabolismo , Proteína Básica de Mielina/fisiología , Neuronas/fisiología , Oligodendroglía , Ratas , Ratas Wistar , Proteínas Recombinantes/farmacología , Transducción GenéticaRESUMEN
Variations in the gene for the nucleotide-binding oligomerisation domain (NOD) 2 have been associated with Crohn's disease but not multiple sclerosis (MS). Here we investigate the effect of three polymorphisms in the NOD2 gene (rs5743277, rs2066842 and rs5743291) on cytokine production and CD4+ T cell proliferation elicited by human myelin basic protein (MBP) in blood mononuclear cell (MNC) cultures from 29 patients with MS. No polymorphism was observed at rs5743277. No associations with the rs2066842 polymorphism were found. Concerning rs5743291, none were homozygous for the minor allele. Seven of 29 (24%) patients were heterozygous, and five of these (71%) exhibited increased MBP-induced CD4+ T cell proliferation versus four of 22 (18%), who were homozygous for the major allele (p<0.04). Interleukin (IL)-5 was induced by MBP in MNC from the same five carriers versus two (9%) homozygotes (p<0.004); four carriers (57%) versus three non-carriers (14%) exhibited IL-17 responses to MBP (p<0.04). By contrast, we found no association between the polymorphisms investigated and interferon-gamma-, tumor necrosis factor-alpha-, IL-2, -4- or IL-10 responses to MBP. These results indicate that the rs5743291 polymorphism influences T helper (Th) cell 2- and Th17 cell responses in MNC from MS patients.
Asunto(s)
Variación Genética , Esclerosis Múltiple/inmunología , Proteína Básica de Mielina/fisiología , Proteína Adaptadora de Señalización NOD2/fisiología , Células Th17/fisiología , Células Th2/fisiología , Adulto , Femenino , Humanos , Masculino , Esclerosis Múltiple/fisiopatologíaRESUMEN
Regulatory T cells (Tregs) play a vital role in autoimmune disorders. Among several markers, forkhead box p3 (Foxp3) is the most specific with regard to Treg activity. Therefore, understanding mechanisms that regulate Foxp3 expression is a critical step for unraveling the complicacy of autoimmune pathophysiology. The present study was undertaken to investigate the crosstalk between NO and Tregs. Interestingly, after myelin basic protein (MBP) priming, the expression of Foxp3 decreased in MBP-primed T cells. However, blocking NO either by inhibiting inducible NO synthase with l-N(6)-(1-iminoethyl)-lysine hydrochloride or through scavenging with PTIO or by pharmacological drugs, such as pravastatin, sodium benzoate, or gemfibrozil, restored the expression of Foxp3 in MBP-primed T cells. However, this restoration of Foxp3 by pharmacological drugs was reversed by S-nitrosoglutathione, an NO donor. Similarly, NO also decreased the populations of Tregs characterized by CD4(+)CD25(+) and CD25(+)FoxP3(+) phenotypes. We have further confirmed this inverse relationship between NO and Foxp3 by analyzing the mRNA expression of Foxp3 and characterizing CD25(+)FoxP3(+) or CD4(+)Foxp3(+) phenotypes from inducible NO synthase knockout mice. Moreover, this inverse relation between NO and Foxp3 also was observed during priming with myelin oligodendrocyte glycoprotein, another target neuroantigen in multiple sclerosis, as well as collagen, a target autoantigen in rheumatoid arthritis. Finally, we demonstrate that NO inhibited the expression of Foxp3 in MBP-primed T cells via soluble guanylyl cyclase-mediated production of cGMP. Taken together, our data imply a novel role of NO in suppressing Foxp3(+) Tregs via the soluble guanylyl cyclase pathway.
Asunto(s)
Regulación hacia Abajo/inmunología , Factores de Transcripción Forkhead/antagonistas & inhibidores , Proteína Básica de Mielina/fisiología , Óxido Nítrico/fisiología , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Animales , Bovinos , Células Cultivadas , Regulación hacia Abajo/genética , Femenino , Factores de Transcripción Forkhead/genética , Glicoproteínas/administración & dosificación , Glicoproteínas/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos , Ratones Noqueados , Proteína Básica de Mielina/administración & dosificación , Glicoproteína Mielina-Oligodendrócito , Óxido Nítrico Sintasa de Tipo II/deficiencia , Óxido Nítrico Sintasa de Tipo II/genética , Fragmentos de Péptidos/administración & dosificación , Fragmentos de Péptidos/fisiología , Linfocitos T Reguladores/enzimología , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismoRESUMEN
OBJECTIVE: Cortical myelin can be severely affected in patients with demyelinating disorders of the central nervous system. However, the functional implication of cortical demyelination remains elusive. In this study, we investigated whether cortical myelin influences cortical spreading depression (CSD). METHODS: CSD measurements were performed in rodent models of toxic and autoimmune induced cortical demyelination, in neuregulin-1 type I transgenic mice displaying cortical hypermyelination, and in glial fibrillary acidic protein-transgenic mice exhibiting pronounced astrogliosis. RESULTS: Cortical demyelination, but not astrogliosis or inflammation per se, was associated with accelerated CSD. In contrast, hypermyelinated neuregulin-1 type I transgenic mice displayed a decelerated CSD propagation. INTERPRETATION: Cortical myelin may be crucially involved in the stabilization and buffering of extracellular ion content that is decisive for CSD propagation velocity and cortical excitability, respectively. Our data thus indicate that cortical involvement in human demyelinating diseases may lead to relevant alterations of cortical function.
Asunto(s)
Corteza Cerebral/fisiopatología , Depresión de Propagación Cortical/fisiología , Enfermedades Desmielinizantes/fisiopatología , Encefalomielitis Autoinmune Experimental/fisiopatología , Proteína Básica de Mielina/análisis , Vaina de Mielina/fisiología , Animales , Astrocitos , Corteza Cerebral/química , Corteza Cerebral/efectos de los fármacos , Depresión de Propagación Cortical/efectos de los fármacos , Cuprizona/farmacología , Enfermedades Desmielinizantes/inducido químicamente , Enfermedades Desmielinizantes/genética , Electroencefalografía , Femenino , Lateralidad Funcional/fisiología , Proteína Ácida Fibrilar de la Glía/genética , Gliosis , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Transgénicos , Esclerosis Múltiple/fisiopatología , Proteína Básica de Mielina/fisiología , Vaina de Mielina/genética , Neurregulina-1/genética , Ratas , Ratas Endogámicas LewRESUMEN
OPCs (oligodendrocyte precursor cells) express golli proteins which, through regulation of Ca2+ influx, appear to be important in OPC process extension/retraction and migration. The aim of the present study was to examine further the role of golli in regulating OPC development. The effects of golli ablation and overexpression were examined in primary cultures of OPCs prepared from golli-KO (knockout) and JOE (golli J37-overexpressing) mice. In OPCs lacking golli, or overexpressing golli, differentiation induced by growth factor withdrawal was impaired. Proliferation analysis in the presence of PDGF (platelet-derived growth factor), revealed that golli enhanced the mitogen-stimulated proliferation of OPCs through activation of SOCCs (store-operated Ca2+ channels). PDGF treatment induced a biphasic increase in OPC intracellular Ca2+, and golli specifically increased Ca2+ influx during the second SOCC-dependent phase that followed the initial release of Ca2+ from intracellular stores. This store-operated Ca2+ uptake appeared to be essential for cell division, since specific SOCC antagonists completely blocked the effects of PDGF and golli on OPC proliferation. Additionally, in OPCs overexpressing golli, increased cell death was observed after mitogen withdrawal. This phenomenon could be prevented by exposure to VOCC (voltage-operated Ca2+ channel) blockers, indicating that the effect of golli on cell death involved increased Ca2+ influx through VOCCs. The results showed a clear effect of golli on OPC development and support a role for golli in modulating multiple Ca2+-regulatory events through VOCCs and SOCCs. Our results also suggest that PDGF engagement of its receptor resulting in OPC proliferation proceeds through activation of SOCCs.
Asunto(s)
Canales de Calcio/metabolismo , Proliferación Celular , Proteína Básica de Mielina/fisiología , Oligodendroglía/metabolismo , Células Madre/metabolismo , Animales , Muerte Celular/fisiología , Células Cultivadas , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Oligodendroglía/citología , Células Madre/citologíaRESUMEN
Cytokine-Ag fusion proteins represent a novel approach for induction of Ag-specific tolerance and may constitute an efficient therapy for autoimmune disease. This study addressed whether a fusion protein containing rat IFN-beta and the encephalitogenic 73-87 determinant of myelin basic protein (i.e., the neuroantigen, or NAg) could prevent or treat experimental autoimmune encephalomyelitis (EAE) in Lewis rats. The optimal structure of the fusion protein was comprised of the rat IFN-beta cytokine as the N-terminal domain with an enterokinase (EK) linker to the NAg domain. Both cytokine and NAg domains had full biological activity. Subcutaneous administration of 1 nmol of IFNbeta-NAg fusion protein in saline on days -21, -14, and -7 before encephalitogenic challenge on day 0 resulted in a substantial attenuation of EAE. In contrast, administration of IFN-beta or NAg alone did not affect susceptibility to EAE. The covalent attachment of IFN-beta and NAg was not necessary, because separate injections of IFN-beta and NAg at adjacent sites were as effective as injection of IFNbeta-NAg for prevention of disease. When treatment was initiated after disease onset, the rank order of inhibitory activity was as follows: the IFNbeta-NAg fusion protein > or = a mixture of IFN-beta plus NAg > IFN-beta > NAg. The novel finding that IFN-beta acts as a tolerogenic adjuvant as well as a tolerogenic fusion partner may have significance for development of tolerogenic vaccines.
Asunto(s)
Adyuvantes Inmunológicos/administración & dosificación , Encefalomielitis Autoinmune Experimental/inmunología , Encefalomielitis Autoinmune Experimental/prevención & control , Tolerancia Inmunológica , Interferón beta/administración & dosificación , Proteína Básica de Mielina/administración & dosificación , Proteínas Recombinantes de Fusión/administración & dosificación , Adyuvantes Inmunológicos/genética , Secuencia de Aminoácidos , Animales , Línea Celular , Células Cultivadas , Encefalomielitis Autoinmune Experimental/patología , Cobayas , Humanos , Tolerancia Inmunológica/genética , Inmunidad Innata/genética , Interferón beta/genética , Interferón beta/inmunología , Datos de Secuencia Molecular , Proteína Básica de Mielina/genética , Proteína Básica de Mielina/fisiología , Ratas , Ratas Endogámicas Lew , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/inmunología , Vacunas Combinadas/administración & dosificación , Vacunas Combinadas/genética , Vacunas Combinadas/inmunologíaRESUMEN
Multiple sclerosis (MS) is an inflammatory disease where phagocytic cells infiltrate the nerve tissue and act as terminal agents in destruction of the myelin sheath. However, the mechanism that triggers the ability of these cells to recognize myelin remains obscure. We show that myelin basic protein (MBP), a major autoantigen in MS, is a potent and specific ligand for the integrin alpha(M)beta(2) (Mac-1, CD11b/CD18) expressed mainly on phagocytic cells. MBP undergoes a dramatic conformational change when liberated from the lipid-rich environment of the myelin sheath. The MS drug glatiramer acetate mimics the conformationally labile regions of MBP, interacts in the unfolded state strongly with alpha(M)beta(2), and inhibits the MBP binding to alpha(M)beta(2). Our study reveals a link between MBP, glatiramer acetate, and the alpha(M)beta(2) integrin, and suggests a new model for MS pathogenesis based on the recognition of unfolded MBP by the alpha(M)beta(2) integrin.
Asunto(s)
Antígeno de Macrófago-1/metabolismo , Proteína Básica de Mielina/química , Proteína Básica de Mielina/metabolismo , Animales , Autoantígenos/química , Autoantígenos/metabolismo , Autoantígenos/fisiología , Unión Competitiva/inmunología , Bovinos , Adhesión Celular/inmunología , Acetato de Glatiramer , Humanos , Células K562 , Ligandos , Antígeno de Macrófago-1/fisiología , Microesferas , Imitación Molecular/inmunología , Esclerosis Múltiple/metabolismo , Proteína Básica de Mielina/fisiología , Péptidos/química , Péptidos/metabolismo , Unión Proteica/inmunología , Pliegue de ProteínaRESUMEN
We showed previously that the addition to cultured oligodendrocytes (OLs) of multivalent carbohydrate in the form of liposomes containing the two major glycosphingolipids (GSLs) of myelin, galactosylceramide (GalC) and cerebroside sulfate (Sulf), or galactose conjugated to bovine serum albumin caused clustering of GalC on the extracellular surface and myelin basic protein (MBP) on the cytosolic surface. Multivalent carbohydrate also caused depolymerization of actin microfilaments and microtubules, indicating that interaction of the carbohydrate with the OL surface transmits a transmembrane signal to the cytoskeleton. In the present study we show that inhibition of GSL synthesis with fumonisin B1 prevents clustering of MBP in GalC/Sulf-negative oligodendrocytes, suggesting that GSLs are required for the effect. Because the effects of multivalent carbohydrate resemble those caused by the addition of anti-GalC/Sulf antibodies to OLs and because GalC and Sulf can interact with each other by trans carbohydrate-carbohydrate interactions across apposed membranes, these results support the conclusion that the OL receptor for GalC/Sulf in liposomes is GalC/Sulf in the OL membrane. Inhibition of MBP expression using MBP siRNA inhibited GalC clustering, suggesting that MBP is required for the effect. We also investigate the signal transduction pathways involved using a number of enzyme inhibitors. These indicated that the Akt and p42/p44 MAPK pathways, Rho GTPases, and GSK-3beta are involved, consistent with their known involvement in regulation of the cytoskeleton. These interactions between GalC/Sulf-containing liposomes and the OL membrane may mimic interactions between GalC/Sulf-enriched signaling domains when OL cell membranes or the extracellular surfaces of compact myelin come into contact.
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Cerebrósidos/metabolismo , Galactosilceramidas/metabolismo , Liposomas/metabolismo , Proteína Básica de Mielina/fisiología , Oligodendroglía/metabolismo , Transducción de Señal/fisiología , Sulfoglicoesfingolípidos/metabolismo , Animales , Animales Recién Nacidos , Células Cultivadas , Interacciones Farmacológicas , Inhibidores Enzimáticos/farmacología , Oligodendroglía/efectos de los fármacos , ARN Interferente Pequeño/farmacología , Ratas , Ratas Wistar , Transducción de Señal/efectos de los fármacos , Médula Espinal/citología , Transfección/métodosRESUMEN
Deposition of fibrillar amyloid beta-protein (Abeta) in the brain is a prominent pathological feature of Alzheimer disease and related disorders, including familial forms of cerebral amyloid angiopathy (CAA). Mutant forms of Abeta, including Dutch- and Iowa-type Abeta, which are responsible for familial CAA, deposit primarily as fibrillar amyloid along the cerebral vasculature and are either absent or present only as diffuse non-fibrillar plaques in the brain parenchyma. Despite the lack of parenchymal fibril formation in vivo, these CAA mutant Abeta peptides exhibit a markedly increased rate and extent of fibril formation in vitro compared with wild-type Abeta. Based on these conflicting observations, we sought to determine whether brain parenchymal factors that selectively interact with and modulate CAA mutant Abeta fibril assembly exist. Using a combination of immunoaffinity chromatography and mass spectrometry, we identified myelin basic protein (MBP) as a prominent brain parenchymal factor that preferentially binds to CAA mutant Abeta compared with wild-type Abeta. Surface plasmon resonance measurements confirmed that MBP bound more tightly to Dutch/Iowa CAA double mutant Abeta than to wild-type Abeta. Using a combination of biochemical and ultrastructural techniques, we found that MBP inhibited the fibril assembly of CAA mutant Abeta. Together, these findings suggest a possible role for MBP in regulating parenchymal fibrillar Abeta deposition in familial CAA.
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Péptidos beta-Amiloides/metabolismo , Amiloide/antagonistas & inhibidores , Angiopatía Amiloide Cerebral Familiar/metabolismo , Proteína Básica de Mielina/fisiología , Secuencia de Aminoácidos , Amiloide/biosíntesis , Péptidos beta-Amiloides/genética , Animales , Bovinos , Angiopatía Amiloide Cerebral Familiar/genética , Humanos , Ratones , Datos de Secuencia MolecularRESUMEN
Physical development, behavioral reactions, and training capacity were studied in the progeny of female BALB/c mice with high levels of antibodies to myelin basic protein. The proposed protocol of immunization ensures high levels of antibodies to myelin basic protein in this mouse strain. High level of antibodies to myelin basic protein in pregnant females causes an increase in the blood level of these antibodies in the progeny. Inhibitory effect of antibodies to myelin basic protein on physical development, training process, and memory in mouse pups was detected.
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Anticuerpos/inmunología , Conducta Animal , Proteína Básica de Mielina/inmunología , Animales , Animales Recién Nacidos , Anticuerpos/sangre , Bovinos , Femenino , Ratones , Ratones Endogámicos BALB C , Actividad Motora , Proteína Básica de Mielina/fisiología , EmbarazoRESUMEN
Myelin basic protein (MBP) binds to negatively charged lipids on the cytosolic surface of oligodendrocyte membranes and is responsible for adhesion of these surfaces in the multilayered myelin sheath. The pattern of extensive post-translational modifications of MBP is dynamic during normal central nervous system (CNS) development and during myelin degeneration in multiple sclerosis (MS), affecting its interactions with the myelin membranes and with other molecules. In particular, the degree of deimination (or citrullination) of MBP is correlated with the severity of MS, and may represent a primary defect that precedes neurodegeneration due to autoimmune attack. That the degree of MBP deimination is also high in early CNS development indicates that this modification plays major physiological roles in myelin assembly. In this review, we describe the structural and functional consequences of MBP deimination in healthy and diseased myelin.
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Citrulina/fisiología , Proteína Básica de Mielina , Secuencia de Aminoácidos , Animales , Artritis Reumatoide/fisiopatología , Humanos , Hidrolasas/metabolismo , Datos de Secuencia Molecular , Esclerosis Múltiple/fisiopatología , Proteína Básica de Mielina/química , Proteína Básica de Mielina/fisiología , Vaina de Mielina/química , Vaina de Mielina/metabolismo , Conformación Proteica , Procesamiento Proteico-Postraduccional , Desiminasas de la Arginina ProteicaRESUMEN
Fusion proteins incorporating anti-inflammatory cytokines and immunodominant self antigen as separate domains of a single protein may hold promise for development of antigen-specific tolerogenic vaccines. Proteins incorporating rat sequences of IL-1RA, IL-2, IL-4, IL-10, or IL-13 were expressed as fusion proteins containing the major encephalitogenic region of myelin basic protein (MBP). These fusion proteins were expressed via baculovirus (bv) expression systems and were shown to have cytokine-dependent and antigen-specific biological activity. In the case of the IL-2 and IL-4 fusion proteins, covalent linkage of the cytokine and neuroantigen domains resulted in synergistic antigen presentation. These data indicate that the cytokine domain may be able to modulate APC activity and simultaneously target the covalently tethered NAg for enhanced presentation by certain APC subsets. Cytokine/antigen fusion proteins may represent a novel tool for antigen-specific immune modulation in autoimmune disease.