RESUMEN
We previously reported that in the absence of Prostaglandin D2 synthase (L-PGDS) peripheral nerves are hypomyelinated in development and that with aging they present aberrant myelin sheaths. We now demonstrate that L-PGDS expressed in Schwann cells is part of a coordinated program aiming at preserving myelin integrity. In vivo and in vitro lipidomic, metabolomic and transcriptomic analyses confirmed that myelin lipids composition, Schwann cells energetic metabolism and key enzymes controlling these processes are altered in the absence of L-PGDS. Moreover, Schwann cells undergo a metabolic rewiring and turn to acetate as the main energetic source. Further, they produce ketone bodies to ensure glial cell and neuronal survival. Importantly, we demonstrate that all these changes correlate with morphological myelin alterations and describe the first physiological pathway implicated in preserving PNS myelin. Collectively, we posit that myelin lipids serve as a reservoir to provide ketone bodies, which together with acetate represent the adaptive substrates Schwann cells can rely on to sustain the axo-glial unit and preserve the integrity of the PNS.
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Although inflammation appears to play a role in neurolymphomatosis (NL), the mechanisms leading to degeneration in the peripheral nervous system are poorly understood. The purpose of this exploratory study was to identify molecular pathways underlying NL pathogenesis, combining clinical and neuropathological investigation with gene expression (GE) studies. We characterized the clinical and pathological features of eight patients with NL. We further analysed GE changes in sural nerve biopsies obtained from a subgroup of NL patients (n=3) and thirteen patients with inflammatory neuropathies as neuropathic controls. Based on the neuropathic symptoms and signs, NL patients were classified into three forms of neuropathy: chronic symmetrical sensorimotor polyneuropathy (SMPN, n=3), multiple mononeuropathy (MN, n=4) and acute motor-sensory axonal neuropathy (AMSAN, n=1). Predominantly diffuse malignant cells infiltration of epineurium was present in chronic SMPN, whereas endoneurial perivascular cells invasion was observed in MN. In contrast, diffuse endoneurium malignant cells localization occurred in AMSAN. We identified alterations in the expression of 1266 genes, with 115 up-regulated and 1151 down-regulated genes, which were mainly associated with ribosomal proteins (RP) and olfactory receptors (OR) signaling pathways, respectively. Among the top up-regulated genes were actin alpha 1 skeletal muscle (ACTA1) and desmin (DES). Similarly, in NL nerves ACTA1, DES and several RPs were highly expressed, associated with endothelial cells and pericytes abnormalities. Peripheral nerve involvement may be due to conversion towards a more aggressive phenotype, potentially explaining the poor prognosis. The candidate genes reported in this study may be a source of clinical biomarkers for NL.
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Phosphorylated TDP-43 (pTDP-43) aggregates in the cytoplasm of motor neurons and neuroglia in the brain are one of the pathological hallmarks of amyotrophic lateral sclerosis. Although the axons exceed the total volume of motor neuron soma by several orders of magnitude, systematic studies investigating the presence and distribution of pTDP-43 aggregates within motor nerves are still lacking. The aim of this study is to define the TDP-43/pTDP-43 pathology in diagnostic motor nerve biopsies performed on a large cohort of patients presenting with a lower motor neuron syndrome and to assess whether this might be a discriminating tissue biomarker for amyotrophic lateral sclerosis and non-amyotrophic lateral sclerosis cases. We retrospectively evaluated 102 lower motor neuron syndrome patients referred to our centre for a diagnostic motor nerve biopsy. Histopathological criteria of motor neuron disease and motor neuropathy were applied by two independent evaluators, who were blind to clinical data. TDP-43 and pTDP-43 were evaluated by immunohistochemistry, and results compared to final clinical diagnosis. We detected significant differences between amyotrophic lateral sclerosis and non-amyotrophic lateral sclerosis cases in pTDP-43 expression in myelinated fibres: axonal accumulation was detected in 98.2% of patients with amyotrophic lateral sclerosis versus 30.4% of non-amyotrophic lateral sclerosis samples (P < 0.0001), while concomitant positive staining in Schwan cell cytoplasm was found in 70.2% of patients with amyotrophic lateral sclerosis versus 17.4% of patients who did not have amyotrophic lateral sclerosis (P < 0.001). Importantly, we were also able to detect pTDP-43 aggregates in amyotrophic lateral sclerosis cases displaying normal features at standard histopathological analysis. Our findings demonstrated that a specific pTDP-43 signature is present in the peripheral nervous system of patients with amyotrophic lateral sclerosis, and could be exploited as a specific, accessible tissue biomarker. The detection of pTDP-43 aggregates within motor nerves of living patients with amyotrophic lateral sclerosis, occurring before axonal degeneration, suggests that this is an early event that may contribute to amyotrophic lateral sclerosis pathogenesis.
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Esclerosis Amiotrófica Lateral , Proteínas de Unión al ADN/metabolismo , Esclerosis Amiotrófica Lateral/metabolismo , Humanos , Neuronas Motoras/metabolismo , Sistema Nervioso Periférico , Estudios RetrospectivosRESUMEN
Oligodendrocytes are the primary target of demyelinating disorders, and progressive neurodegenerative changes may evolve in the CNS. DNA damage and oxidative stress are considered key pathogenic events, but the underlying molecular mechanisms remain unclear. Moreover, animal models do not fully recapitulate human diseases, complicating the path to effective treatments. Here we report that mice with cell-autonomous deletion of the nuclear COP9 signalosome component CSN5 (JAB1) in oligodendrocytes develop DNA damage and defective DNA repair in myelinating glial cells. Interestingly, oligodendrocytes lacking JAB1 expression underwent a senescence-like phenotype that fostered chronic inflammation and oxidative stress. These mutants developed progressive CNS demyelination, microglia inflammation, and neurodegeneration, with severe motor deficits and premature death. Notably, blocking microglia inflammation did not prevent neurodegeneration, whereas the deletion of p21CIP1 but not p16INK4a pathway ameliorated the disease. We suggest that senescence is key to sustaining neurodegeneration in demyelinating disorders and may be considered a potential therapeutic target.
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Envejecimiento/metabolismo , Complejo del Señalosoma COP9/deficiencia , Eliminación de Gen , Enfermedades Neurodegenerativas/metabolismo , Oligodendroglía/metabolismo , Péptido Hidrolasas/deficiencia , Envejecimiento/genética , Envejecimiento/patología , Animales , Complejo del Señalosoma COP9/metabolismo , Inflamación/genética , Inflamación/metabolismo , Inflamación/patología , Ratones , Ratones Noqueados , Enfermedades Neurodegenerativas/genética , Enfermedades Neurodegenerativas/patología , Oligodendroglía/patología , Péptido Hidrolasas/metabolismoRESUMEN
Demyelination is a key pathogenic feature of multiple sclerosis (MS). Here, we evaluated the astrocyte contribution to myelin loss and focused on the neurotrophin receptor TrkB, whose up-regulation on the astrocyte finely demarcated chronic demyelinated areas in MS and was paralleled by neurotrophin loss. Mice lacking astrocyte TrkB were resistant to demyelination induced by autoimmune or toxic insults, demonstrating that TrkB signaling in astrocytes fostered oligodendrocyte damage. In vitro and ex vivo approaches highlighted that astrocyte TrkB supported scar formation and glia proliferation even in the absence of neurotrophin binding, indicating TrkB transactivation in response to inflammatory or toxic mediators. Notably, our neuropathological studies demonstrated copper dysregulation in MS and model lesions and TrkB-dependent expression of copper transporter (CTR1) on glia cells during neuroinflammation. In vitro experiments evidenced that TrkB was critical for the generation of glial intracellular calcium flux and CTR1 up-regulation induced by stimuli distinct from neurotrophins. These events led to copper uptake and release by the astrocyte, and in turn resulted in oligodendrocyte loss. Collectively, these data demonstrate a pathogenic demyelination mechanism via the astrocyte release of copper and open up the possibility of restoring copper homeostasis in the white matter as a therapeutic target in MS.
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Astrocitos/metabolismo , Astrocitos/patología , Cobre/metabolismo , Esclerosis Múltiple/metabolismo , Animales , Transporte Biológico , Enfermedad Crónica , Cicatriz/patología , Cuprizona , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental , Humanos , Inflamación/patología , Ligandos , Proteínas de Transporte de Membrana/metabolismo , Ratones Noqueados , Vaina de Mielina/metabolismo , Factores de Crecimiento Nervioso/metabolismo , Receptor trkB/metabolismo , Regulación hacia Arriba , Sustancia Blanca/patologíaRESUMEN
BACKGROUND: Mutations of the mitochondrial protein paraplegin cause hereditary spastic paraplegia type 7 (SPG7), a so-far untreatable degenerative disease of the upper motoneuron with still undefined pathomechanism. The intermittent mitochondrial permeability transition pore (mPTP) opening, called flickering, is an essential process that operates to maintain mitochondrial homeostasis by reducing intra-matrix Ca2+ and reactive oxygen species (ROS) concentration, and is critical for efficient synaptic function. METHODS: We use a fluorescence-based approach to measure mPTP flickering in living cells and biochemical and molecular biology techniques to dissect the pathogenic mechanism of SPG7. In the SPG7 animal model we evaluate the potential improvement of the motor defect, neuroinflammation and neurodegeneration by means of an mPTP inducer, the benzodiazepine Bz-423. FINDINGS: We demonstrate that paraplegin is required for efficient transient opening of the mPTP, that is impaired in both SPG7 patients-derived fibroblasts and primary neurons from Spg7-/- mice. We show that dysregulation of mPTP opening at the pre-synaptic terminal impairs neurotransmitter release leading to ineffective synaptic transmission. Lack of paraplegin impairs mPTP flickering by a mechanism involving increased expression and activity of sirtuin3, which promotes deacetylation of cyclophilin D, thus hampering mPTP opening. Pharmacological treatment with Bz-423, which bypasses the activity of CypD, normalizes synaptic transmission and rescues the motor impairment of the SPG7 mouse model. INTERPRETATION: mPTP targeting opens a new avenue for the potential therapy of this form of spastic paraplegia. FUNDING: Telethon Foundation grant (TGMGCSBX16TT); Dept. of Defense, US Army, grant W81XWH-18-1-0001.
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ATPasas Asociadas con Actividades Celulares Diversas/genética , Metaloendopeptidasas/genética , Poro de Transición de la Permeabilidad Mitocondrial/metabolismo , Paraplejía Espástica Hereditaria/genética , Paraplejía Espástica Hereditaria/metabolismo , ATPasas Asociadas con Actividades Celulares Diversas/metabolismo , Animales , Apoptosis/genética , Transporte Biológico , Calcio/metabolismo , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades , Edición Génica , Células HEK293 , Humanos , Potencial de la Membrana Mitocondrial , Metaloendopeptidasas/metabolismo , Ratones , Ratones Noqueados , Mitocondrias/genética , Mitocondrias/metabolismo , Mutación , Neuronas/metabolismo , Fenotipo , Especies Reactivas de Oxígeno/metabolismo , Sirtuina 3/metabolismo , Médula Espinal/metabolismo , Médula Espinal/patología , Vesículas Sinápticas/metabolismoRESUMEN
Mucopolysaccharidosis type I (MPS-I), a lysosomal storage disorder caused by a deficiency of alpha-L-iduronidase enzyme, results in the progressive accumulation of glycosaminoglycans and consequent multiorgan dysfunction. Despite the effectiveness of hematopoietic stem cell transplantation (HSCT) and enzyme replacement therapy (ERT) in correcting clinical manifestations related to visceral organs, complete improvement of musculoskeletal and neurocognitive defects remains an unmet challenge and provides an impact on patients' quality of life. We tested the therapeutic efficacy of combining HSCT and ERT in the neonatal period. Using a mouse model of MPS-I, we demonstrated that the combination therapy improved clinical manifestations in organs usually refractory to current treatment. Moreover, combination with HSCT prevented the production of anti-IDUA antibodies that negatively impact ERT efficacy. The added benefits of combining both treatments also resulted in a reduction of skeletal anomalies and a trend towards decreased neuroinflammation and metabolic abnormalities. As currently there are limited therapeutic options for MPS-I patients, our findings suggest that the combination of HSCT and ERT during the neonatal period may provide a further step forward in the treatment of this rare disease.
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Remodelación Ósea , Modelos Animales de Enfermedad , Terapia de Reemplazo Enzimático/métodos , Trasplante de Células Madre Hematopoyéticas/métodos , Iduronidasa/fisiología , Mucopolisacaridosis I/terapia , Animales , Animales Recién Nacidos , Terapia Combinada , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mucopolisacaridosis I/enzimología , Mucopolisacaridosis I/patologíaRESUMEN
Myelin, one of the most important adaptations of vertebrates, is essential to ensure efficient propagation of the electric impulse in the nervous system and to maintain neuronal integrity. In the central nervous system (CNS), the development of oligodendrocytes and the process of myelination are regulated by the coordinated action of several positive and negative cell-extrinsic factors. We and others previously showed that secretases regulate the activity of proteins essential for myelination. We now report that the neuronal α-secretase ADAM17 controls oligodendrocyte differentiation and myelin formation in the CNS. Ablation of Adam17 in neurons impairs in vivo and in vitro oligodendrocyte differentiation, delays myelin formation throughout development and results in hypomyelination. Furthermore, we show that this developmental defect is, in part, the result of altered Notch/Jagged 1 signaling. Surprisingly, in vivo conditional loss of Adam17 in immature oligodendrocytes has no effect on myelin formation. Collectively, our data indicate that the neuronal α-secretase ADAM17 is required for proper CNS myelination. Further, our studies confirm that secretases are important post-translational regulators of myelination although the mechanisms controlling CNS and peripheral nervous system (PNS) myelination are distinct.
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Proteína ADAM17/metabolismo , Sistema Nervioso Central/metabolismo , Vaina de Mielina/metabolismo , Neuronas/metabolismo , Oligodendroglía/metabolismo , 2',3'-Nucleótido Cíclico 3'-Fosfodiesterasa/metabolismo , Proteína ADAM17/genética , Animales , Diferenciación Celular/fisiología , Sistema Nervioso Central/citología , Ratones Transgénicos , Neurogénesis/fisiologíaRESUMEN
BACKGROUND: The major histocompatibility complex I (MHCI) is a key molecule for the interaction of mononucleated cells with CD8+T lymphocytes. We previously showed that MHCI is upregulated in the spinal cord microglia and motor axons of transgenic SOD1G93A mice. METHODS: To assess the role of MHCI in the disease, we examined transgenic SOD1G93A mice crossbred with ß2 microglobulin-deficient mice, which express little if any MHCI on the cell surface and are defective for CD8+ T cells. RESULTS: The lack of MHCI and CD8+ T cells in the sciatic nerve affects the motor axon stability, anticipating the muscle atrophy and the disease onset. In contrast, MHCI depletion in resident microglia and the lack of CD8+ T cell infiltration in the spinal cord protect the cervical motor neurons delaying the paralysis of forelimbs and prolonging the survival of SOD1G93A mice. CONCLUSIONS: We provided straightforward evidence for a dual role of MHCI in the peripheral nervous system (PNS) compared to the CNS, pointing out regional and temporal differences in the clinical responses of ALS mice. These findings offer a possible explanation for the failure of systemic immunomodulatory treatments and suggest new potential strategies to prevent the progression of ALS.
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Esclerosis Amiotrófica Lateral/inmunología , Linfocitos T CD8-positivos/inmunología , Antígenos de Histocompatibilidad Clase I/inmunología , Sistema Nervioso Periférico/inmunología , Médula Espinal/inmunología , Esclerosis Amiotrófica Lateral/patología , Animales , Linfocitos T CD8-positivos/patología , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Sistema Nervioso Periférico/patología , Médula Espinal/patologíaRESUMEN
BACKGROUND: Increasing evidence suggests that the immune system has a beneficial role in the progression of amyotrophic lateral sclerosis (ALS) although the mechanism remains unclear. Recently, we demonstrated that motor neurons (MNs) of C57SOD1G93A mice with slow disease progression activate molecules classically involved in the cross-talk with the immune system. This happens a lot less in 129SvSOD1G93A mice which, while expressing the same amount of transgene, had faster disease progression and earlier axonal damage. The present study investigated whether and how the immune response is involved in the preservation of motor axons in the mouse model of familial ALS with a more benign disease course. METHODS: First, the extent of axonal damage, Schwann cell proliferation, and neuromuscular junction (NMJ) denervation were compared between the two ALS mouse models at the disease onset. Then, we compared the expression levels of different immune molecules, the morphology of myelin sheaths, and the presence of blood-derived immune cell infiltrates in the sciatic nerve of the two SOD1G93A mouse strains using immunohistochemical, immunoblot, quantitative reverse transcription PCR, and rotating-polarization Coherent Anti-Stokes Raman Scattering techniques. RESULTS: Muscle denervation, axonal dysregulation, and myelin disruption together with reduced Schwann cell proliferation are prominent in 129SvSOD1G93A compared to C57SOD1G93A mice at the disease onset, and this correlates with a faster disease progression in the first strain. On the contrary, a striking increase of immune molecules such as CCL2, MHCI, and C3 was seen in sciatic nerves of slow progressor C57SOD1G93A mice and this was accompanied by heavy infiltration of CD8+ T lymphocytes and macrophages. These phenomena were not detectable in the peripheral nervous system of fast-progressing mice. CONCLUSIONS: These data show for the first time that damaged MNs in SOD1-related ALS actively recruit immune cells in the peripheral nervous system to delay muscle denervation and prolong the lifespan. On the contrary, the lack of this response has a negative impact on the disease course.
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Esclerosis Amiotrófica Lateral/complicaciones , Citocinas/metabolismo , Mutación/genética , Enfermedades del Sistema Nervioso Periférico , Superóxido Dismutasa/genética , Esclerosis Amiotrófica Lateral/genética , Animales , Citocinas/genética , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Femenino , Regulación de la Expresión Génica/genética , Regulación de la Expresión Génica/fisiología , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Desnervación Muscular , Proteínas del Tejido Nervioso/metabolismo , Nervio Obturador/metabolismo , Nervio Obturador/patología , Enfermedades del Sistema Nervioso Periférico/etiología , Enfermedades del Sistema Nervioso Periférico/inmunología , Enfermedades del Sistema Nervioso Periférico/patología , Complejo de la Endopetidasa Proteasomal/metabolismo , Nervio Ciático/metabolismo , Nervio Ciático/patología , Transducción de Señal/genéticaRESUMEN
Neuregulin 1 type III is processed following regulated intramembrane proteolysis, which allows communication from the plasma membrane to the nucleus. We found that the intracellular domain of neuregulin 1 type III upregulated the prostaglandin D2 synthase (L-pgds, also known as Ptgds) gene, which, together with the G protein-coupled receptor Gpr44, forms a previously unknown pathway in PNS myelination. Neuronal L-PGDS is secreted and produces the PGD2 prostanoid, a ligand of Gpr44. We found that mice lacking L-PGDS were hypomyelinated. Consistent with this, specific inhibition of L-PGDS activity impaired in vitro myelination and caused myelin damage. Furthermore, in vivo ablation and in vitro knockdown of glial Gpr44 impaired myelination. Finally, we identified Nfatc4, a key transcription factor for myelination, as one of the downstream effectors of PGD2 activity in Schwann cells. Thus, L-PGDS and Gpr44 are previously unknown components of an axo-glial interaction that controls PNS myelination and possibly myelin maintenance.
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Oxidorreductasas Intramoleculares/biosíntesis , Lipocalinas/biosíntesis , Vaina de Mielina/metabolismo , Fibras Nerviosas Mielínicas/metabolismo , Sistema Nervioso Periférico/metabolismo , Receptores Inmunológicos/biosíntesis , Receptores de Prostaglandina/biosíntesis , Transducción de Señal/fisiología , Animales , Células Cultivadas , Femenino , Ganglios Espinales/metabolismo , Ganglios Espinales/ultraestructura , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Vaina de Mielina/ultraestructura , Fibras Nerviosas Mielínicas/ultraestructura , Sistema Nervioso Periférico/ultraestructura , RatasRESUMEN
Growing evidence indicates that alterations within the peripheral nervous system (PNS) are involved at an early stage in the amyotrophic lateral sclerosis (ALS) pathogenetic cascade. In this study, magnetic resonance imaging (MRI), neurophysiologic analyses, and histologic analyses were used to monitor the extent of PNS damage in the hSOD-1 ALS rat model. The imaging signature of the disease was defined using in vivo MRI of the sciatic nerve. Initial abnormalities were detected in the nerves by an increase in T2 relaxation time before the onset of clinical disease; diffusion MRI showed a progressive increase in mean and radial diffusivity and reduction of fractional anisotropy at advanced stages of disease. Histologic analysis demonstrated early impairment of the blood-nerve barrier followed by acute axonal degeneration associated with endoneurial edema and macrophage response in motor nerve compartments. Progressive axonal degeneration and motor nerve fiber loss correlated with MRI and neurophysiologic changes. These functional and morphologic investigations of the PNS might be applied in following disease progression in preclinical therapeutic studies. This study establishes the PNS signature in this rat ALS model (shedding new light into pathogenesis) and provides a rationale for translating into future systematic MRI studies of PNS involvement in patients with ALS.
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Esclerosis Amiotrófica Lateral/genética , Enfermedades del Sistema Nervioso Periférico/genética , Superóxido Dismutasa/genética , Esclerosis Amiotrófica Lateral/patología , Animales , Axones/patología , Fenómenos Electrofisiológicos/genética , Fenómenos Electrofisiológicos/fisiología , Femenino , Humanos , Imagen por Resonancia Magnética , Microscopía Electrónica , Degeneración Nerviosa/patología , Enfermedades del Sistema Nervioso Periférico/patología , Ratas , Ratas Sprague-Dawley , Ratas Transgénicas , Nervio Ciático/patología , Superóxido Dismutasa/fisiología , Superóxido Dismutasa-1RESUMEN
Axonal sorting is a crucial event in nerve formation and requires proper Schwann cell proliferation, differentiation, and contact with axons. Any defect in axonal sorting results in dysmyelinating peripheral neuropathies. Evidence from mouse models shows that axonal sorting is regulated by laminin211- and, possibly, neuregulin 1 (Nrg1)-derived signals. However, how these signals are integrated in Schwann cells is largely unknown. We now report that the nuclear Jun activation domain-binding protein 1 (Jab1) may transduce laminin211 signals to regulate Schwann cell number and differentiation during axonal sorting. Mice with inactivation of Jab1 in Schwann cells develop a dysmyelinating neuropathy with axonal sorting defects. Loss of Jab1 increases p27 levels in Schwann cells, which causes defective cell cycle progression and aberrant differentiation. Genetic down-regulation of p27 levels in Jab1-null mice restores Schwann cell number, differentiation, and axonal sorting and rescues the dysmyelinating neuropathy. Thus, Jab1 constitutes a regulatory molecule that integrates laminin211 signals in Schwann cells to govern cell cycle, cell number, and differentiation. Finally, Jab1 may constitute a key molecule in the pathogenesis of dysmyelinating neuropathies.
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Axones/fisiología , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Neurogénesis/fisiología , Péptido Hidrolasas/metabolismo , Células de Schwann/fisiología , Animales , Western Blotting , Pesos y Medidas Corporales , Bromodesoxiuridina , Complejo del Señalosoma COP9 , Diferenciación Celular/fisiología , Proliferación Celular , Inmunohistoquímica , Etiquetado Corte-Fin in Situ , Péptidos y Proteínas de Señalización Intracelular/genética , Laminina/metabolismo , Ratones , Ratones Noqueados , Péptido Hidrolasas/genética , Prueba de Desempeño de Rotación con Aceleración ConstanteRESUMEN
Animal models provide an important tool to investigate the pathogenesis of neuromuscular disorders. In the present study, we analyze fiber composition of the brachial plexus branches to the pectoral muscles: the medial anterior thoracic nerve (MATN) and the lateral anterior thoracic nerve (LATN). The morphological and morphometric characteristics and the percentage of motor fibers within each nerve are here reported, adding information to microscopic anatomy knowledge of the rat brachial plexus. As control, we employed the quadriceps nerve, commonly used for the evaluation of motor fibers at hindlimbs. We demonstrated that the MATN and the LATN are predominantly composed of large motor fibers and therefore could be employed to evaluate the peripheral nervous system (PNS) involvement at forelimbs in neurological diseases models, predominantly affecting the motor fiber compartment.
RESUMEN
Remodeling of extracellular matrix (ECM) is a critical step in peripheral nerve regeneration. In fact, in human neuropathies, endoneurial ECM enriched in fibrin and vitronectin associates with poor regeneration and worse clinical prognosis. Accordingly in animal models, modification of the fibrinolytic complex activity has profound effects on nerve regeneration: high fibrinolytic activity and low levels of fibrin correlate with better nerve regeneration. The urokinase plasminogen receptor (uPAR) is a major component of the fibrinolytic complex, and binding to urokinase plasminogen activator (uPA) promotes fibrinolysis and cell movement. uPAR is expressed in peripheral nerves, however, little is known on its potential function on nerve development and regeneration. Thus, we investigated uPAR null mice and observed that uPAR is dispensable for nerve development, whereas, loss of uPAR affects nerve regeneration. uPAR null mice showed reduced nerve repair after sciatic nerve crush. This was a consequence of reduced fibrinolytic activity and increased deposition of endoneurial fibrin and vitronectin. Exogenous fibrinolysis in uPAR null mice rescued nerve repair after sciatic nerve crush. Finally, we measured the fibrinolytic activity in sural nerve biopsies from patients with peripheral neuropathies. We showed that neuropathies with defective regeneration had reduced fibrinolytic activity. On the contrary, neuropathies with signs of active regeneration displayed higher fibrinolytic activity. Overall, our results suggest that enforced fibrinolysis may facilitate regeneration and outcome of peripheral neuropathies.
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Fibrinólisis , Compresión Nerviosa , Regeneración Nerviosa/fisiología , Enfermedades del Sistema Nervioso Periférico/metabolismo , Enfermedades del Sistema Nervioso Periférico/fisiopatología , Receptores del Activador de Plasminógeno Tipo Uroquinasa/metabolismo , Adulto , Anciano , Animales , Matriz Extracelular/efectos de los fármacos , Matriz Extracelular/metabolismo , Femenino , Fibrina/metabolismo , Fibrinólisis/efectos de los fármacos , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Vaina de Mielina/efectos de los fármacos , Vaina de Mielina/metabolismo , Vaina de Mielina/patología , Regeneración Nerviosa/efectos de los fármacos , Enfermedades del Sistema Nervioso Periférico/tratamiento farmacológico , Enfermedades del Sistema Nervioso Periférico/patología , Receptores del Activador de Plasminógeno Tipo Uroquinasa/deficiencia , Proteínas Recombinantes/farmacología , Proteínas Recombinantes/uso terapéutico , Nervio Ciático/efectos de los fármacos , Nervio Ciático/metabolismo , Nervio Ciático/patología , Nervio Ciático/fisiopatología , Activador de Plasminógeno de Tipo Uroquinasa/farmacología , Activador de Plasminógeno de Tipo Uroquinasa/uso terapéuticoRESUMEN
Myelination is a complex process that requires coordinated Schwann cell-axon interactions during development and regeneration. Positive and negative regulators of myelination have been recently described, and can belong either to Schwann cells or neurons. Vimentin is a fibrous component present in both Schwann cell and neuron cytoskeleton, the expression of which is timely and spatially regulated during development and regeneration. We now report that vimentin negatively regulates myelination, as loss of vimentin results in peripheral nerve hypermyelination, owing to increased myelin thickness in vivo, in transgenic mice and in vitro in a myelinating co-culture system. We also show that this is due to a neuron-autonomous increase in the levels of axonal neuregulin 1 (NRG1) type III. Accordingly, genetic reduction of NRG1 type III in vimentin-null mice rescues hypermyelination. Finally, we demonstrate that vimentin acts synergistically with TACE, a negative regulator of NRG1 type III activity, as shown by hypermyelination of double Vim/Tace heterozygous mice. Our results reveal a novel role for the intermediate filament vimentin in myelination, and indicate vimentin as a regulator of NRG1 type III function.
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Regulación del Desarrollo de la Expresión Génica , Vaina de Mielina/metabolismo , Nervios Periféricos/metabolismo , Vimentina/fisiología , Proteínas ADAM/metabolismo , Proteína ADAM17 , Animales , Axones/metabolismo , Técnicas de Cocultivo , Citoesqueleto/metabolismo , Heterocigoto , Humanos , Ratones , Ratones Endogámicos C57BL , Neurregulina-1/metabolismo , Ratas , Células de Schwann/citologíaRESUMEN
We previously reported that autosomal recessive demyelinating Charcot-Marie-Tooth (CMT) type 4B1 neuropathy with myelin outfoldings is caused by loss of MTMR2 (Myotubularin-related 2) in humans, and we created a faithful mouse model of the disease. MTMR2 dephosphorylates both PtdIns3P and PtdIns(3,5)P(2), thereby regulating membrane trafficking. However, the function of MTMR2 and the role of the MTMR2 phospholipid phosphatase activity in vivo in the nerve still remain to be assessed. Mutations in FIG4 are associated with CMT4J neuropathy characterized by both axonal and myelin damage in peripheral nerve. Loss of Fig4 function in the plt (pale tremor) mouse produces spongiform degeneration of the brain and peripheral neuropathy. Since FIG4 has a role in generation of PtdIns(3,5)P(2) and MTMR2 catalyzes its dephosphorylation, these two phosphatases might be expected to have opposite effects in the control of PtdIns(3,5)P(2) homeostasis and their mutations might have compensatory effects in vivo. To explore the role of the MTMR2 phospholipid phosphatase activity in vivo, we generated and characterized the Mtmr2/Fig4 double null mutant mice. Here we provide strong evidence that Mtmr2 and Fig4 functionally interact in both Schwann cells and neurons, and we reveal for the first time a role of Mtmr2 in neurons in vivo. Our results also suggest that imbalance of PtdIns(3,5)P(2) is at the basis of altered longitudinal myelin growth and of myelin outfolding formation. Reduction of Fig4 by null heterozygosity and downregulation of PIKfyve both rescue Mtmr2-null myelin outfoldings in vivo and in vitro.
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Enfermedad de Charcot-Marie-Tooth/genética , Flavoproteínas/genética , Proteínas Tirosina Fosfatasas no Receptoras/genética , Células de Schwann/enzimología , Aminopiridinas/farmacología , Animales , Axones/enzimología , Axones/metabolismo , Enfermedad de Charcot-Marie-Tooth/enzimología , Enfermedad de Charcot-Marie-Tooth/metabolismo , Flavoproteínas/metabolismo , Compuestos Heterocíclicos con 3 Anillos/farmacología , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mutación , Vaina de Mielina/genética , Vaina de Mielina/metabolismo , Neuronas/enzimología , Neuronas/metabolismo , Nervios Periféricos/enzimología , Nervios Periféricos/crecimiento & desarrollo , Nervios Periféricos/metabolismo , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfatos de Fosfatidilinositol/genética , Fosfatos de Fosfatidilinositol/metabolismo , Fosfoinosítido Fosfatasas , Fosfolípidos/genética , Fosfolípidos/metabolismo , Proteínas Tirosina Fosfatasas no Receptoras/metabolismo , Ratas , Células de Schwann/metabolismoRESUMEN
Numerous transgenic and knockout mouse models of human hereditary neuropathies have become available over the past decade. We describe a simple, reproducible, and safe biopsy of mouse skin for histopathological evaluation of the peripheral nervous system (PNS) in models of hereditary neuropathies. We compared the diagnostic outcome between sciatic nerve and dermal nerves found in skin biopsy (SB) from the hind foot. A total of five animal models of different Charcot-Marie-Tooth neuropathies, and one model of congenital muscular dystrophy associated neuropathy were examined. In wild type mice, dermal nerve fibers were readily identified by immunohistochemistry, light, and electron microscopy and they appeared similar to myelinated fibers in sciatic nerve. In mutant mice, SB manifested myelin abnormalities similar to those observed in sciatic nerves, including hypomyelination, onion bulbs, myelin outfolding, redundant loops, and tomacula. In many strains, however, SB showed additional abnormalities--fiber loss, dense neurofilament packing with lower phosphorylation status, and axonal degeneration-undetected in sciatic nerve, possibly because SB samples distal nerves. SB, a reliable technique to investigate peripheral neuropathies in human beings, is also useful to investigate animal models of hereditary neuropathies. Our data indicate that SB may reveal distal axonal pathology in mouse models and permits sequential follow-up of the neuropathy in an individual mouse, thereby reducing the number of mice necessary to document pathology of the PNS.
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Axones/patología , Biopsia/métodos , Enfermedad de Charcot-Marie-Tooth/patología , Pie/inervación , Pie/patología , Animales , Dermis/inervación , Dermis/patología , Modelos Animales de Enfermedad , Epidermis/inervación , Epidermis/patología , Humanos , Ratones , Ratones Mutantes Neurológicos , Vaina de Mielina/patología , Fibras Nerviosas Mielínicas/patología , Nervio Ciático/patología , Nervio Sural/patologíaRESUMEN
The peri-ventricular area of the forebrain constitutes a preferential site of inflammation in multiple sclerosis, and the sub-ventricular zone (SvZ) is functionally altered in its animal model experimental autoimmune encephalomyelitis (EAE). The reasons for this preferential localization are still poorly understood. We show here that, in EAE mice, blood-derived macrophages, T and B cells and microglia (Mg) from the surrounding parenchyma preferentially accumulate within the SvZ, deranging its cytoarchitecture. We found that the chemokine Cxcl10 is constitutively expressed by a subset of cells within the SvZ, constituting a primary chemo-attractant signal for activated T cells. During EAE, T cells and macrophages infiltrating the SvZ in turn secrete pro-inflammatory cytokines such as TNFalpha and IFNgamma capable to induce Mg cells accumulation and SvZ derangement. Accordingly, lentiviral-mediated over-expression of IFNgamma or TNFalpha in the healthy SvZ mimics Mg/microglia recruitment occurring during EAE, while Cxcl10 over-expression in the SvZ is able to increase the frequency of peri-ventricular inflammatory lesions only in EAE mice. Finally, we show, by RT-PCR and in situ hybridization, that Cxcl10 is expressed also in the healthy human SvZ, suggesting a possible molecular parallelism between multiple sclerosis and EAE.
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Movimiento Celular/fisiología , Quimiocina CXCL10/inmunología , Encefalomielitis Autoinmune Experimental/inmunología , Prosencéfalo/anatomía & histología , Animales , Trasplante de Médula Ósea , Quimiocina CXCL10/genética , Quimiocina CXCL10/metabolismo , Quimiocinas/inmunología , Encefalomielitis Autoinmune Experimental/patología , Femenino , Humanos , Interferón gamma/inmunología , Interferón gamma/metabolismo , Activación de Linfocitos/inmunología , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones , Ratones Endogámicos C57BL , Microglía/inmunología , Esclerosis Múltiple/inmunología , Linfocitos T/inmunología , Linfocitos T/metabolismo , Quimera por Trasplante , Factor de Necrosis Tumoral alfa/inmunología , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Schwann cells integrate signals deriving from the axon and the basal lamina to myelinate peripheral nerves. Integrin alpha6beta4 is a laminin receptor synthesized by Schwann cells and displayed apposed to the basal lamina. alpha6beta4 integrin expression in Schwann cells is induced by axons at the onset of myelination, and rises in adulthood. The beta4 chain has a uniquely long cytoplasmic domain that interacts with intermediate filaments such as dystonin, important in peripheral myelination. Furthermore, alpha6beta4 integrin binds peripheral myelin protein 22, whose alteration causes the most common demyelinating hereditary neuropathy. All these data suggest a role for alpha6beta4 integrin in peripheral nerve myelination. Here we show that ablating alpha6beta4 integrin specifically in Schwann cells of transgenic mice does not affect peripheral nerve development, myelin formation, maturation, or regeneration. However, consistent with maximal expression in adult nerves, alpha6beta4 integrin-null myelin is more prone to abnormal folding with aging. When the laminin receptor dystroglycan is also ablated, major folding abnormalities occur, associated with acute demyelination in some peripheral nervous system districts. These data indicate that, similar to its role in skin, alpha6beta4 integrin confers stability to myelin in peripheral nerves.