RESUMEN
Diabetic neuropathy (DNP), afflicting sensory and motor nerve fibers, is a major complication in diabetes. The underlying cellular mechanisms of axon degeneration are poorly understood. IGFBP5, an inhibitory binding protein for insulin-like growth factor 1 (IGF1) is highly up-regulated in nerve biopsies of patients with DNP. We investigated the pathogenic relevance of this finding in transgenic mice overexpressing IGFBP5 in motor axons and sensory nerve fibers. These mice develop motor axonopathy and sensory deficits similar to those seen in DNP. Motor axon degeneration was also observed in mice in which the IGF1 receptor (IGF1R) was conditionally depleted in motoneurons, indicating that reduced activity of IGF1 on IGF1R in motoneurons is responsible for the observed effect. These data provide evidence that elevated expression of IGFBP5 in diabetic nerves reduces the availability of IGF1 for IGF1R on motor axons, thus leading to progressive neurodegeneration. Inhibition of IGFBP5 could thus offer novel treatment strategies for DNP.
Asunto(s)
Axones/fisiología , Proteínas Portadoras/metabolismo , Diabetes Mellitus Experimental/fisiopatología , Neuropatías Diabéticas/fisiopatología , Neuronas Motoras/fisiología , Degeneración Nerviosa/fisiopatología , Animales , Axones/patología , Aumento de la Célula , Supervivencia Celular/fisiología , Células Cultivadas , Diabetes Mellitus Experimental/patología , Neuropatías Diabéticas/patología , Humanos , Ratones Transgénicos , Actividad Motora/fisiología , Neuronas Motoras/patología , Degeneración Nerviosa/patología , Nervio Frénico/patología , Nervio Frénico/fisiopatología , Receptor IGF Tipo 1/metabolismo , Nervio Ciático/patología , Nervio Ciático/fisiopatología , Sensación/fisiologíaRESUMEN
Insulin-like growth factor I (IGF-I) has been successfully tested in the SOD1-G93A mouse model of familial amyotrophic lateral sclerosis (ALS) and proposed for clinical treatment. However, beneficial effects required gene therapy or intrathecal application. Circumventing the dosing issues we recently found that polyethylene glycol (PEG) modified IGF-I (PEG-IGF-I) modulated neuromuscular function after systemic application, and protected against disease progression in a motor neuron disease model. Here we investigated its effects in two SOD1-G93A mouse lines, the G1L with a milder and the G1H with a more severe phenotype. Results showed that in G1L mice, PEG-IGF-I treatment significantly improved muscle force, motor coordination and animal survival. In contrast, treatment of G1H mice with PEG-IGF-I or IGF-I even at high doses did not beneficially affect survival or functional outcomes despite increased signalling in brain and spinal cord by both agents. In conclusion, the data point towards further investigation of the therapeutic potential of PEG-IGF-I in ALS patients with less severe clinical phenotypes.
Asunto(s)
Esclerosis Amiotrófica Lateral/tratamiento farmacológico , Factor I del Crecimiento Similar a la Insulina/uso terapéutico , Neuronas Motoras/efectos de los fármacos , Esclerosis Amiotrófica Lateral/patología , Animales , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Masculino , Ratones , Ratones Transgénicos , Neuronas Motoras/metabolismo , Índice de Severidad de la EnfermedadRESUMEN
Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS). So far, immunological mechanisms responsible for demyelination have been the focus of interest. However, mechanisms regulating axon maintenance as well as glial precursor-cell proliferation and oligodendrocyte survival might also influence disease outcome. The cytokine ciliary neurotrophic factor (CNTF), which was originally identified as a survival factor for isolated neurons, promotes differentiation, maturation and survival of oligodendrocytes. To investigate the role of endogenous CNTF in inflammatory demyelinating disease, we studied myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE) in CNTF-deficient and wild-type C57BL/6 mice. Disease was more severe in CNTF-deficient mice and recovery was poor, with a 60% decrease in the number of proliferating oligodendrocyte precursor cells (OPCs) and a more than 50% increase in the rate of oligodendrocyte apoptosis. In addition, vacuolar dystrophy of myelin and axonal damage were more severe in CNTF-deficient mice. These specific pathological features could be prevented by treatment with an antiserum against tumor necrosis factor-alpha, suggesting that endogenous CNTF may counterbalance this effect of TNF-alpha (ref. 7). Here we identify a factor that modulates, in an inflammatory environment, glial cell survival and is an outcome determinant of EAE.
Asunto(s)
Factor Neurotrófico Ciliar/fisiología , Enfermedades Desmielinizantes/prevención & control , Encefalomielitis Autoinmune Experimental/patología , Esclerosis Múltiple/prevención & control , Factores de Crecimiento Nervioso/fisiología , Animales , Factor Neurotrófico Ciliar/deficiencia , Factor Neurotrófico Ciliar/genética , Citocinas/fisiología , Inflamación/prevención & control , Ratones , Ratones Endogámicos C57BL , Ratones NoqueadosRESUMEN
Mutations in Ribosomal s6 kinase 2 (Rsk2) are associated with severe neuronal dysfunction in Coffin-Lowry syndrome (CLS) patients, flies and mice. So far, the mechanisms of how Rsk2 regulates development, maintenance and activity of neurons are not understood. We have investigated the consequences of Rsk2 deficiency in mouse spinal motoneurons. Survival of isolated Rsk2 deficient motoneurons is not reduced, but these cells grow significantly longer neurites. Conversely, overexpression of a constitutively active form of Rsk2 leads to reduced axon growth. Increased axon growth in Rsk2 deficient neurons was accompanied by higher Erk 1/2 phosphorylation, and the knockout phenotype could be rescued by pharmacological inhibition of MAPK/Erk kinase (Mek). These data indicate that Rsk2 negatively regulates axon elongation via the MAPK pathway. Thus, the functional defects observed in the nervous system of CLS patients and animal models with Rsk2 deficiency might be caused by dysregulated neurite growth rather than primary neurodegeneration.
Asunto(s)
Axones/fisiología , Neuronas Motoras , Proteínas Quinasas S6 Ribosómicas 90-kDa/metabolismo , Animales , Supervivencia Celular , Femenino , Humanos , Sistema de Señalización de MAP Quinasas/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neuronas Motoras/citología , Neuronas Motoras/fisiología , Proteínas Quinasas S6 Ribosómicas 90-kDa/genética , Médula Espinal/citologíaRESUMEN
Members of the ciliary neurotrophic factor (CNTF)/leukemia inhibitory factor (LIF)/cardiotrophin gene family are potent survival factors for embryonic and lesioned motoneurons. These factors act via receptor complexes involving gp130 and LIFR-beta and ligand binding leads to activation of various signaling pathways, including phosphorylation of Stat3. The role of Stat3 in neuronal survival was investigated in mice by Cre-mediated gene ablation in motoneurons. Cre is expressed under the neurofilament light chain (NF-L) promoter, starting around E12 when these neurons become dependent on neurotrophic support. Loss of motoneurons during the embryonic period of naturally occurring cell death is not enhanced in NF-L-Cre; Stat3(flox/KO) mice although motoneurons isolated from these mice need higher concentrations of CNTF for maximal survival in culture. In contrast, motoneuron survival is significantly reduced after facial nerve lesion in the adult. These neurons, however, can be rescued by the addition of neurotrophic factors, including CNTF. Stat3 is essential for upregulation of Reg-2 and Bcl-xl expression in lesioned motoneurons. Our data show that Stat3 activation plays an essential role for motoneuron survival after nerve lesion in postnatal life but not during embryonic development, indicating that signaling requirements for motoneuron survival change during maturation.
Asunto(s)
Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Traumatismos del Nervio Facial/patología , Neuronas Motoras/citología , Proteínas del Tejido Nervioso , Sistema Nervioso/embriología , Sistema Nervioso/patología , Transducción de Señal , Transactivadores/genética , Transactivadores/metabolismo , Envejecimiento , Animales , Animales Recién Nacidos , Axotomía , Proteínas de Unión al Calcio/metabolismo , Muerte Celular/efectos de los fármacos , Supervivencia Celular , Células Cultivadas , Factor Neurotrófico Ciliar/farmacología , Traumatismos del Nervio Facial/genética , Traumatismos del Nervio Facial/metabolismo , Eliminación de Gen , Integrasas/genética , Integrasas/metabolismo , Litostatina , Ratones , Ratones Noqueados , Neuronas Motoras/metabolismo , Neuronas Motoras/patología , Sistema Nervioso/crecimiento & desarrollo , Sistema Nervioso/metabolismo , Proteínas de Neurofilamentos/genética , Especificidad de Órganos , Regiones Promotoras Genéticas/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factor de Transcripción STAT3 , Proteínas Virales/genética , Proteínas Virales/metabolismo , Proteína bcl-XRESUMEN
Bag1 is a cochaperone for the heat-shock protein Hsp70 that interacts with C-Raf, B-Raf, Akt, Bcl-2, steroid hormone receptors and other proteins. Here we use targeted gene disruption in mice to show that Bag1 has an essential role in the survival of differentiating neurons and hematopoietic cells. Cells of the fetal liver and developing nervous system in Bag1-/- mice underwent massive apoptosis. Lack of Bag1 did not disturb the primary function of Akt or Raf, as phosphorylation of the forkhead transcription factor FKHR and activation of extracellular signal-regulated kinase (Erk)-1/2 were not affected. However, the defect was associated with the disturbance of a tripartite complex formed by Akt, B-Raf and Bag1, in addition to the absence of Bad phosphorylation at Ser136. We also observed reduced expression of members of the inhibitor of apoptosis (IAP) family. Our data show that Bag1 is a physiological mediator of extracellular survival signals linked to the cellular mechanisms that prevent apoptosis in hematopoietic and neuronal progenitor cells.
Asunto(s)
Diferenciación Celular/fisiología , Células Madre Hematopoyéticas/fisiología , Proteínas de la Membrana/fisiología , Neuronas Motoras/fisiología , Médula Espinal/citología , Fosfatasa Alcalina/metabolismo , Análisis de Varianza , Animales , Southern Blotting/métodos , Western Blotting/métodos , Proteínas Portadoras/metabolismo , Recuento de Células/métodos , Supervivencia Celular/fisiología , Células Cultivadas , Proteínas de Unión al ADN , Complejo IV de Transporte de Electrones/metabolismo , Embrión de Mamíferos , Proteínas del Ojo/metabolismo , Fibroblastos/metabolismo , Proteínas HSP70 de Choque Térmico/deficiencia , Proteínas de Homeodominio/metabolismo , Inmunohistoquímica/métodos , Inmunoprecipitación/métodos , Etiquetado Corte-Fin in Situ/métodos , Proteínas de Filamentos Intermediarios/metabolismo , Isoenzimas/metabolismo , Hígado/citología , Hígado/crecimiento & desarrollo , Hígado/metabolismo , Proteínas de la Membrana/deficiencia , Ratones , Ratones Noqueados , Mutación , Proteínas del Tejido Nervioso/metabolismo , Nestina , Proteínas de Neurofilamentos/metabolismo , Factor de Transcripción PAX6 , Factores de Transcripción Paired Box , Propidio , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas B-raf/metabolismo , Proteínas Proto-Oncogénicas c-akt , ARN Mensajero/biosíntesis , ARN Interferente Pequeño/metabolismo , Ratas , Proteínas Represoras/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Médula Espinal/crecimiento & desarrollo , Médula Espinal/metabolismo , Factores de Transcripción , Transfección/métodos , Proteína Letal Asociada a bclRESUMEN
The Hey basic helix-loop-helix transcription factors are downstream effectors of Notch signaling in the cardiovascular system. Mice lacking Hey2 develop cardiac hypertrophy, often associated with congenital heart defects, whereas combined Hey1/Hey2 deficiency leads to severe vascular defects and embryonic lethality around embryonic day E9.5. The molecular basis of these disorders is poorly understood, however, since target genes of Hey transcription factors in the affected tissues remain elusive. To identify genes regulated by Hey factors we have generated a conditional Hey1 knockout mouse. This strain was used to generate paired Hey2- and Hey1/2-deficient embryonic stem cell lines. Comparison of these cell lines by microarray analysis identified GATA4 and GATA6 as differentially expressed genes. Loss of Hey1/2 leads to elevated GATA4/6 and ANF mRNA levels in embryoid bodies, while forced expression of Hey factors strongly represses expression of the GATA4 and GATA6 promoter in various cell lines. In addition, the promoter activity of the GATA4/6 target gene ANF was inhibited by Hey1, Hey2, and HeyL. Protein interaction and mutation analyses suggest that repression is due to direct binding of Hey proteins to GATA4 and GATA6, blocking their transcriptional activity. In Hey2-deficient fetal hearts we observed elevated mRNA levels of ANF and CARP. Expression of ANF and Hey2 is normally restricted to the trabecular and compact myocardial layer, respectively. Intriguingly, loss of Hey2 leads to ectopic ANF expression in the compact layer, suggesting a direct role for Hey2 in limiting ANF expression in this cardiac compartment.
Asunto(s)
Factor Natriurético Atrial/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Proteínas de Ciclo Celular/metabolismo , Corazón Fetal/metabolismo , Factor de Transcripción GATA4/genética , Proteínas Represoras/metabolismo , Animales , Secuencia de Bases , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/genética , Línea Celular , ADN Complementario/genética , Regulación de la Expresión Génica , Marcación de Gen , Secuencias Hélice-Asa-Hélice/genética , Histona Desacetilasas/metabolismo , Humanos , Ratones , Ratones Noqueados , Proteínas Musculares , Proteínas Nucleares/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Regiones Promotoras Genéticas , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas Represoras/genéticaRESUMEN
Leukemia inhibitory factor (LIF) receptor beta (LIFRbeta) is a receptor for a variety of neurotrophic cytokines, including LIF, ciliary neurotrophic factor (CNTF), and cardiotrophin-1 (CT-1). These cytokines play an essential role for the survival and maintenance of developing and postnatal somatic motoneurons. CNTF may also serve the maintenance of autonomic, preganglionic sympathetic neurons (PSNs) in the spinal cord, as suggested by its capacity to prevent their death after destruction of one of their major targets, the adrenal medulla. Although somatic motoneurons and PSNs share a common embryonic origin, they are distinct in several respects, including responses to lesions. We have studied PSNs in mice with targeted deletions of the LIFRbeta or CT-1 genes, respectively. We show that LIF, CNTF, and CT-1 are synthesized in embryonic adrenal gland and spinal cord and that PSNs express LIFRbeta. In embryonic day 18.5 LIFRbeta (-/-) and CT-1 (-/-) mice, PSNs were reduced by approximately 20%. PSNs projecting to the adrenal medulla were more severely affected (-55%). Although LIFRbeta (-/-) mice revealed normal numbers of adrenal chromaffin cells and axons terminating on chromaffin cells, levels of adrenaline and numbers of adrenaline-synthesizing cells were significantly reduced. We conclude that activation of LIFRbeta is required for normal development of PSNs and one of their prominent targets, the adrenal medulla. Thus, both somatic motoneurons and PSNs in the spinal cord depend on LIFRbeta signaling for their development and maintenance, although PSNs seem to be overall less affected than somatic motoneurons by LIFRbeta deprivation.
Asunto(s)
Médula Suprarrenal/fisiología , Citocinas/fisiología , Interleucina-6/fisiología , Fibras Nerviosas/fisiología , Neuronas/fisiología , Actinas/fisiología , Médula Suprarrenal/efectos de los fármacos , Médula Suprarrenal/inervación , Animales , Bencilaminas/farmacología , Citocinas/deficiencia , Citocinas/genética , Maleato de Dizocilpina/farmacología , Hipocampo/fisiología , Interleucina-6/deficiencia , Interleucina-6/genética , Factor Inhibidor de Leucemia , Ratones , Fibras Nerviosas/efectos de los fármacos , Neuronas/efectos de los fármacos , Oligopéptidos/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Sulfonamidas/farmacología , Sistema Nervioso Simpático/embriología , Sinapsis/efectos de los fármacos , Sinapsis/fisiología , Membranas Sinápticas/efectos de los fármacos , Membranas Sinápticas/fisiología , Sinaptosomas/efectos de los fármacos , Sinaptosomas/fisiologíaRESUMEN
Members of the ciliary neurotrophic factor (CNTF)-leukemia inhibitory factor (LIF) gene family play an essential role for survival of developing and postnatal motoneurons. When subunits of the shared receptor complex are inactivated by homologous recombination, the mice die at approximately birth and exhibit reduced numbers of motoneurons in the spinal cord and brainstem nuclei. However, mice in which cntf, lif, or cardiotrophin-1 (ct-1) are inactivated can survive and show less motoneuron cell loss. This suggests cooperative and redundant roles of these ligands. However, their cooperative functions are not well understood. We generated cntf/lif/ct-1 triple-knock-out and combinations of double-knock-out mice to study the individual and combined roles of CNTF, LIF and CT-1 on postnatal motoneuron survival and function. Triple-knock-out mice exhibit increased motoneuron cell loss in the lumbar spinal cord that correlates with muscle weakness during early postnatal development. LIF deficiency leads to pronounced loss of distal axons and motor endplate alterations, whereas CNTF-and/or CT-1-deficient mice do not show significant changes in morphology of these structures. In cntf/lif/ct-1 triple-knock-out mice, various degrees of muscle fiber type grouping are found, indicating that denervation and reinnervation had occurred. We conclude from these findings that CNTF, LIF, and CT-1 have distinct functions for motoneuron survival and function and that LIF plays a more important role for postnatal maintenance of distal axons and motor endplates than CNTF or CT-1.
Asunto(s)
Factor Neurotrófico Ciliar/fisiología , Citocinas/fisiología , Interleucina-6/fisiología , Neuronas Motoras/fisiología , Fibras Musculares Esqueléticas/patología , Debilidad Muscular/fisiopatología , Animales , Factor Neurotrófico Ciliar/deficiencia , Factor Neurotrófico Ciliar/genética , Citocinas/deficiencia , Citocinas/genética , Fuerza de la Mano , Interleucina-6/genética , Factor Inhibidor de Leucemia , Ratones , Ratones Noqueados , Placa Motora/ultraestructura , Desnervación Muscular , Debilidad Muscular/genética , Debilidad Muscular/patología , Degeneración Nerviosa , Nervios Periféricos/patología , Médula Espinal/patologíaRESUMEN
The microtubule associated tumor suppressor gene 1 (MTUS1) is a recently published tumor suppressor gene, which has also been shown to act as an early component in the growth inhibitory signaling cascade of the angiotensin II type 2 receptor (AT2R). In this study we report the generation of MTUS1 knock-out (KO) mice, which develop normally but reveal higher body weights and slightly decreased blood pressure levels. Twenty-eight percent of the studied MTUS1 KO mice also developed heart hypertrophy and 12% developed nephritis, independent of blood pressure levels. Forty-three percent of the MTUS1 KO mice revealed lymphoid hyperplasia affecting spleen (20%), kidney (37%), lung (23%), lymph nodes (17%), and liver (17%) accompanied with leukocytosis, lymphocytosis, and mild anemia. One animal (3%) developed a marginal zone B-cell lymphoma affecting submandibular salivary gland and regional lymph nodes. The symptoms of all mentioned animals are consistent with a B-cell lymphoproliferative disease with features of systemic lupus erythematosus. In addition, body weight of the MTUS1 KO mice was significantly increased and isolated skin fibroblasts showed increased cell proliferation and decreased cell size, compared to wild-type (WT) fibroblasts in response to depleted FCS concentration and lack of growth factors. In conclusion we herein report the first generation of a MTUS1 KO mouse, developing spontaneous heart hypertrophy and increased cell proliferation, confirming once more the anti-proliferative effect of MTUS1, and a SLE-like lymphoproliferative disease suggesting crucial role in regulation of inflammation. These MTUS1 KO mice can therefore serve as a model for further investigations in cardiovascular disease, autoimmune disease and carcinogenesis.
Asunto(s)
Cardiomegalia/patología , Linfoma de Células B de la Zona Marginal/patología , Trastornos Linfoproliferativos/patología , Proteínas Supresoras de Tumor/deficiencia , Animales , Cardiomegalia/sangre , Cardiomegalia/genética , Cardiomegalia/metabolismo , Proteínas Portadoras/genética , Procesos de Crecimiento Celular/fisiología , Fibroblastos/patología , Inmunohistoquímica , Linfoma de Células B de la Zona Marginal/sangre , Linfoma de Células B de la Zona Marginal/genética , Linfoma de Células B de la Zona Marginal/metabolismo , Trastornos Linfoproliferativos/sangre , Trastornos Linfoproliferativos/genética , Trastornos Linfoproliferativos/metabolismo , Ratones , Ratones Noqueados , Piel/patología , Proteínas Supresoras de Tumor/genéticaRESUMEN
Based on its potent neurotrophic and myotrophic activities, insulin-like growth factor I (IGF-I) has been proposed for treatment of neuromuscular disorders such as muscular dystrophies and amyotrophic lateral sclerosis (ALS). However, the short half life in the circulation limits its use in vivo. At least in mouse models, beneficial effects are generally only observed by dosing regimens such as minipumps or gene therapy that are difficult to translate to patients. We have developed a polyethylene glycol coupled IGF-I (PEG-IGF-I) that could circumvent these problems by longer half-life, showing all features of a therapeutic agent supporting muscular and neuronal function. Here we investigated its effects in the pmn mutant mouse, a model with typical dying-back motoneuron degeneration. In vitro, PEG-IGF-I and rhIGF-I profoundly promoted survival axonal growth of wild-type as well as pmn mutant embryonic motoneurons, suggesting that PEG-IGF-I had a fully conserved neurotrophic activity via its receptor. In vivo, treatment of pmn mutant mice with PEG-IGF-I prolonged survival, protected against late stage weight loss and significantly maintained muscle force and motor coordination. Consistently, PEG-IGF-I treatment rescued facial and lumbar motoneurons from cell death and partially preserved phrenic nerve myelinated axons. The data support that PEG-IGF-I could be used for treatment of neuromuscular diseases in a clinically feasible manner.
Asunto(s)
Diseño de Fármacos , Factor I del Crecimiento Similar a la Insulina/análogos & derivados , Factor I del Crecimiento Similar a la Insulina/farmacocinética , Enfermedad de la Neurona Motora/tratamiento farmacológico , Neuronas Motoras/efectos de los fármacos , Polietilenglicoles/farmacocinética , Animales , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Modelos Animales de Enfermedad , Femenino , Feto/citología , Técnicas In Vitro , Ratones , Ratones Endogámicos , Ratones Mutantes , Neuronas Motoras/citología , Fibras Nerviosas Mielínicas/efectos de los fármacos , Nervio Frénico/citología , Embarazo , Proteínas Recombinantes/farmacología , Médula Espinal/citologíaRESUMEN
The relative contribution of the two tryptophan hydroxylase (TPH) isoforms, TPH1 and TPH2, to brain serotonergic system function is controversial. To investigate the respective role of TPH2 in neuron serotonin (5-HT) synthesis and the role of 5-HT in brain development, mice with a targeted disruption of Tph2 were generated. The preliminary results indicate that in Tph2 knockout mice raphe neurons are completely devoid of 5-HT, whereas no obvious alteration in morphology and fiber distribution are observed. The findings confirm the exclusive specificity of Tph2 in brain 5-HT synthesis and suggest that Tph2-synthesized 5-HT is not required for serotonergic neuron formation.
Asunto(s)
Química Encefálica/genética , Encéfalo/citología , Neuronas/fisiología , Serotonina/deficiencia , Triptófano Hidroxilasa/fisiología , Animales , Eliminación de Gen , Inmunohistoquímica , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Núcleos del Rafe/citología , Núcleos del Rafe/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Serotonina/biosíntesis , Proteínas de Transporte de Serotonina en la Membrana Plasmática/genética , Proteínas de Transporte de Serotonina en la Membrana Plasmática/metabolismo , Triptófano Hidroxilasa/genéticaRESUMEN
The neurotrophic cytokines ciliary neurotrophic factor and leukemia inhibitory factor (LIF) play a key role in neuronal and oligodendrocyte survival and as protective factors in neuroinflammation. To further elucidate the potential of endogenous LIF in modulating neuroinflammation, we studied myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis in LIF knockout mice (LIF(-/-) mice). In the late phase of active myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis, LIF(-/-) mice exhibited a markedly milder disease course. The inflammatory infiltrate in LIF(-/-) mice was characterized by an increase in neutrophilic granulocytes early and fewer infiltrating macrophages associated with less demyelination later in the disease. In good correlation with an effect of endogenous LIF on the immune response, we found an Ag-specific T cell-priming defect with impaired IFN-gamma production in LIF(-/-) mice. On the molecular level, the altered recruitment of inflammatory cells is associated with distinct patterns of chemokine production in LIF(-/-) mice with an increase of CXCL1 early and a decrease of CCL2, CCL3, and CXCL10 later in the disease. These data reveal that endogenous LIF is an immunologically active molecule in neuroinflammation. This establishes a link between LIF and the immune system which was not observed in the ciliary neurotrophic factor knockout mouse.
Asunto(s)
Quimiocinas/fisiología , Quimiotaxis de Leucocito/inmunología , Encefalomielitis Autoinmune Experimental/inmunología , Encefalomielitis Autoinmune Experimental/patología , Factor Inhibidor de Leucemia/deficiencia , Factor Inhibidor de Leucemia/fisiología , Animales , Línea Celular , Células Cultivadas , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Células Dendríticas/patología , Encefalomielitis Autoinmune Experimental/genética , Encefalomielitis Autoinmune Experimental/metabolismo , Femenino , Predisposición Genética a la Enfermedad , Glicoproteínas/inmunología , Glicoproteínas/toxicidad , Factor Inhibidor de Leucemia/genética , Macrófagos/inmunología , Macrófagos/metabolismo , Macrófagos/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Glicoproteína Mielina-Oligodendrócito , Fragmentos de Péptidos/inmunología , Fragmentos de Péptidos/toxicidad , Receptores OSM-LIF/biosíntesis , Receptores OSM-LIF/genética , Linfocitos T/inmunología , Linfocitos T/metabolismo , Linfocitos T/patologíaRESUMEN
Neurotrophins are potent survival factors for developing and injured neurons. However, they are not being used to treat neurodegenerative diseases because of difficulties in administration and numerous side effects that have been encountered in previous clinical trials. Their biological activities use Trk (tropomyosin-related kinase) transmembrane tyrosine kinases. Therefore, one alternative approach is to use transactivation pathways such as adenosine 2A receptor agonists, which can activate Trk receptor signaling independent of neurotrophin binding. However, the relevance in vivo and applicability of these transactivation events during neurodegenerative and injury conditions have never been extensively studied. Here we demonstrate that motoneuron survival after facial nerve lesioning is significantly enhanced by transactivation of Trk receptor tyrosine kinases by adenosine agonists. Moreover, survival of motoneurons directly required the activation of the BDNF receptor TrkB and an increase in Akt (AKT8 virus oncogene cellular homolog) activity. The ability of small molecules to activate a trophic response by using Trk signaling provides a unique mechanism to promote survival signals in motoneurons and suggests new strategies for using transactivation in neurodegenerative diseases.
Asunto(s)
Neuronas Motoras/citología , Neuronas Motoras/enzimología , Receptor trkB/genética , Receptores de Adenosina A2/metabolismo , Activación Transcripcional/genética , Adenosina/análogos & derivados , Adenosina/farmacología , Agonistas del Receptor de Adenosina A2 , Animales , Axotomía , Separación Celular , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Embrión de Mamíferos/citología , Embrión de Mamíferos/efectos de los fármacos , Embrión de Mamíferos/enzimología , Activación Enzimática/efectos de los fármacos , Nervio Facial/efectos de los fármacos , Nervio Facial/patología , Ratones , Neuronas Motoras/efectos de los fármacos , Fenetilaminas/farmacología , Activación Transcripcional/efectos de los fármacosRESUMEN
UNLABELLED: We previously reported that exogenous cardiotrophin-1 (CT-1), a member of the IL-6 family of cytokines, exerts hepatoprotective effects. Because CT-1 is expressed in the normal liver, we hypothesized that this cytokine may constitute an endogenous defense of the liver against proapoptotic stimuli. Here, we found that CT-1-/- mice died faster than wild-type animals after challenge with a lethal dose of the Fas agonist Jo-2. At sublethal doses of Jo-2, all wild-type mice survived whereas CT-1-/- animals developed extensive hepatocyte apoptosis with 50% mortality at 24 hours. Pretreatment with CT-1 improved survival and reduced injury in both CT-1-/- and wild-type animals. Upon Fas ligation the activation of STAT-3, a molecule that defends the liver against apoptosis, was lower in CT-1-/- mice than in wild-type animals despite similar IL-6 up-regulation in the 2 groups. Analysis of liver transcriptome in CT-1-/- and wild-type mice showed that 9 genes reported to be associated with cell survival/death functions were differentially expressed in the 2 groups. Four of these genes [IGFBP1, peroxiredoxin3, TNFR1, and calpastatin (endogenous inhibitor of calpain)] could be validated by real-time PCR. All of them were down-regulated in CT-1-/- mice and were modulated by CT-1 administration. Treatment of CT-1-/- animals with the calpain inhibitor MDL28170 afforded significant protection against Fas-induced liver injury. CONCLUSION: CT-1-/- mice are highly sensitive to Fas-mediated apoptosis due in part to deficient STAT-3 activation and inadequate control of calpain activity during the apoptotic process. Our data show that CT-1 is a natural defense of the liver against apoptosis. This cytokine may have therapeutic potential.
Asunto(s)
Apoptosis/fisiología , Citocinas/fisiología , Hepatocitos/fisiología , Hígado/patología , Animales , Anticuerpos/farmacología , Apoptosis/efectos de los fármacos , Proteínas de Unión al Calcio/genética , Proteínas de Unión al Calcio/fisiología , Citocinas/genética , Regulación de la Expresión Génica , Hepatocitos/patología , Hígado/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Peroxidasas/genética , Peroxidasas/fisiología , Peroxiredoxina III , Peroxirredoxinas , Receptores Tipo I de Factores de Necrosis Tumoral/genética , Receptores Tipo I de Factores de Necrosis Tumoral/fisiología , Factor de Transcripción STAT3/genética , Factor de Transcripción STAT3/fisiología , Receptor fas/inmunologíaRESUMEN
The transcription factor Sox10 regulates early neural crest development, specification of neural crest-derived lineages and terminal differentiation of oligodendrocytes in the central nervous system. Here, we generated two novel hypomorphic Sox10 alleles in the mouse. Mutant mice either expressed a Sox10 protein with a triple alanine substitution in the dimerization domain, or a Sox10 protein with a deletion in the central portion that we define as a cell-specific transactivation domain. Phenotypic analysis revealed important roles for a functional dimerization domain and the newly defined novel transactivation domain in melanocyte and enteric nervous system development, whereas early neural crest development and oligodendrocyte differentiation were surprisingly little disturbed in both mutants. Unique requirements were additionally detected for the novel transactivation domain in satellite glia differentiation and during Schwann cell myelination, whereas DNA-dependent dimerization was needed for immature Schwann cells to enter the promyelinating stage. These two hypomorphic alleles thus uncover novel functions of Sox10 in satellite glia and Schwann cells during late developmental stages and reveal important developmental differences between these two types of peripheral glia and oligodendrocytes regarding their reliance on Sox10.
Asunto(s)
Linaje de la Célula , Proteínas del Grupo de Alta Movilidad/fisiología , Neuroglía/fisiología , Sistema Nervioso Periférico/embriología , Factores de Transcripción/fisiología , Alelos , Animales , Linaje de la Célula/genética , Dimerización , Proteínas del Grupo de Alta Movilidad/genética , Proteínas del Grupo de Alta Movilidad/metabolismo , Melanocitos/fisiología , Ratones , Ratones Mutantes , Mutación , Vaina de Mielina , Neuroglía/citología , Oligodendroglía/citología , Oligodendroglía/fisiología , Factores de Transcripción SOXE , Células de Schwann , Eliminación de Secuencia , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Recent evidence has shown that the activation of receptor tyrosine kinases is not only dependent on binding of their ligands but in addition requires adhesion molecules as coreceptors. We have identified CD44v6 as a coreceptor for c-Met in several tumor and primary cells. The CD44v6 ectodomain is required for c-Met activation, whereas the cytoplasmic tail recruits ERM proteins and the cytoskeleton into a signalosome complex. Here we demonstrate that c-Met (and hepatocyte growth factor and Gab1) is haploinsufficient in a cd44-/- background, as the cd44-/-; met+/- (and cd44-/-; hgf+/- and cd44-/-; gab1+/-) mice die at birth. They have impaired synaptic transmission in the respiratory rhythm-generating network and alterations in the phrenic nerve. These results are the first genetic data showing that CD44 and c-Met collaborate in vivo and that they are involved in synaptogenesis and axon myelination in the central and peripheral nervous systems.
Asunto(s)
Haploidia , Receptores de Hialuranos/metabolismo , Proteínas Proto-Oncogénicas c-met/metabolismo , Animales , Animales Recién Nacidos , Encéfalo/patología , Factor de Crecimiento de Hepatocito/metabolismo , Pulmón/anomalías , Pulmón/patología , Ratones , Ratones Endogámicos C57BL , Neuronas Motoras/patología , Fibras Nerviosas/patología , Nervio Frénico/patología , Nervio Frénico/ultraestructura , Transmisión SinápticaRESUMEN
Transgenic mice expressing the superoxide dismutase G93A mutation (SOD1(G93A)) were used to investigate the role of glial inwardly rectifying K(+) (Kir)4.1 channels, which buffer extracellular K(+) increases in response to neuronal excitation. A progressive decrease in Kir4.1 immunoreactivity was observed predominantly in the ventral horn of SOD1(G93A) mutants. Immunoblotting of spinal cord extracts mirrored these changes by showing a loss of Kir4.1 channels from presymptomatic stages onwards. Kir4.1 channels were found to be expressed in the spinal cord grey matter, targetting astrocytes and clustering around capillaries, supporting their role in clearance of extracellular K(+). To understand the functional implications of extracellular K(+) increases, we challenged the NSC34 motor neurone cell line with increasing extracellular K(+) concentrations. Exposure to high extracellular K(+) induced progressive motor neurone cell death. We suggest that loss of Kir4.1 impairs perineural K(+) homeostasis and may contribute to motor neurone degeneration in SOD1(G93A) mutants by K(+) excitotoxic mechanisms.
Asunto(s)
Esclerosis Amiotrófica Lateral/metabolismo , Neuroglía/metabolismo , Canales de Potasio de Rectificación Interna/metabolismo , Médula Espinal/metabolismo , Superóxido Dismutasa/metabolismo , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/patología , Animales , Acuaporina 4/metabolismo , Astrocitos/metabolismo , Western Blotting , Capilares/citología , Capilares/metabolismo , Supervivencia Celular/fisiología , Células Cultivadas , Espacio Extracelular/metabolismo , Técnica del Anticuerpo Fluorescente , Proteína Ácida Fibrilar de la Glía/metabolismo , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Inmunohistoquímica , Ratones , Ratones Transgénicos , Neuronas Motoras/metabolismo , Neuronas Motoras/fisiología , Potasio/metabolismo , Médula Espinal/citología , Médula Espinal/patología , Superóxido Dismutasa/genética , Superóxido Dismutasa-1RESUMEN
Neuronal degeneration in spinal muscular atrophy is caused by reduced expression of the survival motor neuron (SMN) protein. SMN and the tightly interacting Gemin2 form part of a macromolecular complex (SMN complex) that mediates assembly of spliceosomal small nuclear ribonucleoproteins (U snRNPs). We used mouse genetics to investigate the function of this complex in motoneuron maintenance. Reduced Smn/Gemin2 protein levels lead to disturbed U snRNP assembly as indicated by reduced nuclear accumulation of Sm proteins. This finding correlates with enhanced motoneuron degeneration in Gemin2(+/-)/Smn(+/-) mice. Our data provide in vivo evidence that impaired production of U snRNPs contributes to motoneuron degeneration.
Asunto(s)
Neuronas Motoras/patología , Proteínas del Tejido Nervioso/genética , Animales , Muerte Celular , Proteína de Unión a Elemento de Respuesta al AMP Cíclico , Modelos Animales de Enfermedad , Marcación de Gen , Ratones , Ratones Noqueados , Atrofia Muscular Espinal/genética , Atrofia Muscular Espinal/patología , Proteínas de Unión al ARN , Mapeo Restrictivo , Ribonucleoproteínas Nucleares Pequeñas/genética , Proteínas del Complejo SMNRESUMEN
Selenoprotein P (SePP), the major selenoprotein in plasma, has been implicated in selenium transport, selenium detoxification or antioxidant defence. We generated SePP-knockout mice that were viable, but exhibited reduced growth and developed ataxia. Selenium content was elevated in liver, but low in plasma and other tissues, and selenoenzyme activities changed accordingly. Our data reveal that SePP plays a pivotal role in delivering hepatic selenium to target tissues.