RESUMO
Elovl5 elongates fatty acids with 18 carbon atoms and in cooperation with other enzymes guarantees the normal levels of very long-chain fatty acids, which are necessary for a proper membrane structure. Action potential conduction along myelinated axons depends on structural integrity of myelin, which is maintained by a correct amount of fatty acids and a proper interaction between fatty acids and myelin proteins. We hypothesized that in Elovl5-/- mice, the lack of elongation of Elovl5 substrates might cause alterations of myelin structure. The analysis of myelin ultrastructure showed an enlarged periodicity with reduced G-ratio across all axonal diameters. We hypothesized that the structural alteration of myelin might affect the conduction of action potentials. The sciatic nerve conduction velocity was significantly reduced without change in the amplitude of the nerve compound potential, suggesting a myelin defect without a concomitant axonal degeneration. Since Elovl5 is important in attaining normal amounts of polyunsaturated fatty acids, which are the principal component of myelin, we performed a lipidomic analysis of peripheral nerves of Elovl5-deficient mice. The results revealed an unbalance, with reduction of fatty acids longer than 18 carbon atoms relative to shorter ones. In addition, the ratio of saturated to unsaturated fatty acids was strongly increased. These findings point out the essential role of Elovl5 in the peripheral nervous system in supporting the normal structure of myelin, which is the key element for a proper conduction of electrical signals along myelinated nerves.
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Axônios , Bainha de Mielina , Potenciais de Ação/genética , Animais , Axônios/fisiologia , Elongases de Ácidos Graxos/genética , Ácidos Graxos/metabolismo , Camundongos , Bainha de Mielina/metabolismo , Condução Nervosa/genética , Nervos PeriféricosRESUMO
In the last decade, microRNAs have been increasingly recognized as key modulators of glial development. Recently, we identified miR-125a-3p as a new player in oligodendrocyte physiology, regulating in vitro differentiation of oligodendrocyte precursor cells (OPCs). Here, we show that miR-125a-3p is upregulated in active lesions of multiple sclerosis (MS) patients and in OPCs isolated from the spinal cord of chronic experimental autoimmune encephalomyelitis (EAE) mice, but not in those isolated from the spontaneously remyelinating corpus callosum of lysolecithin-treated mice. To test whether a sustained expression of miR-125a-3p in OPCs contribute to defective remyelination, we modulated miR-125a-3p expression in vivo and ex vivo after lysolecithin-induced demyelination. We found that lentiviral over-expression of miR-125a-3p impaired OPC maturation, whereas its downregulation accelerated remyelination. Transcriptome analysis and luciferase reporter assay revealed that these effects are partly mediated by the direct interaction of miR-125a-3p with Slc8a3, a sodium-calcium membrane transporter, and identified novel candidate targets, such as Gas7, that we demonstrated necessary to correctly address oligodendrocytes to terminal maturation. These findings show that miR-125a-3p upregulation negatively affects OPC maturation in vivo, suggest its role in the pathogenesis of demyelinating diseases and unveil new targets for future promyelinating protective interventions.
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Encefalomielite Autoimune Experimental/metabolismo , Inativação Gênica/fisiologia , MicroRNAs/biossíntese , Bainha de Mielina/metabolismo , Remielinização/fisiologia , Substância Branca/metabolismo , Animais , Animais Recém-Nascidos , Células Cultivadas , Doenças Desmielinizantes/genética , Doenças Desmielinizantes/metabolismo , Doenças Desmielinizantes/patologia , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/patologia , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Bainha de Mielina/genética , Bainha de Mielina/patologia , Técnicas de Cultura de Órgãos , Ratos , Ratos Sprague-Dawley , Substância Branca/patologiaRESUMO
A newly proposed form of brain structural plasticity consists of non-newly generated, "immature" neurons of the adult cerebral cortex. Similar to newly generated neurons, these cells express the cytoskeletal protein Doublecortin (DCX), yet they are generated prenatally and then remain in a state of immaturity for long periods. In rodents, the immature neurons are restricted to the paleocortex, whereas in other mammals, they are also found in neocortex. Here, we analyzed the DCX-expressing cells in the whole sheep brain of both sexes to search for an indicator of structural plasticity at a cellular level in a relatively large-brained, long-living mammal. Brains from adult and newborn sheep (injected with BrdU and analyzed at different survival times) were processed for DCX, cell proliferation markers (Ki-67, BrdU), pallial/subpallial developmental origin (Tbr1, Sp8), and neuronal/glial antigens for phenotype characterization. We found immature-like neurons in the whole sheep cortex and in large populations of DCX-expressing cells within the external capsule and the surrounding gray matter (claustrum and amygdala). BrdU and Ki-67 detection at neonatal and adult ages showed that all of these DCX+ cells were generated during embryogenesis, not after birth. These results show that the adult sheep, unlike rodents, is largely endowed with non-newly generated neurons retaining immature features, suggesting that such plasticity might be particularly important in large-brained, long-living mammals.SIGNIFICANCE STATEMENT Brain plasticity is important in adaptation and brain repair. Structural changes span from synaptic plasticity to adult neurogenesis, the latter being highly reduced in large-brained, long-living mammals (e.g., humans). The cerebral cortex contains "immature" neurons, which are generated prenatally and then remain in an undifferentiated state for long periods, being detectable with markers of immaturity. We studied the distribution and developmental origin of these cells in the whole brain of sheep, relatively large-brained, long-living mammals. In addition to the expected cortical location, we also found populations of non-newly generated neurons in several subcortical regions (external capsule, claustrum, and amygdala). These results suggests that non-neurogenic, parenchymal structural plasticity might be more important in large mammals with respect to adult neurogenesis.
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Encéfalo/citologia , Neurogênese/fisiologia , Plasticidade Neuronal/fisiologia , Neurônios/citologia , Animais , Feminino , Masculino , Células-Tronco Neurais/citologia , OvinosRESUMO
Spinocerebellar ataxia 28 is an autosomal dominant neurodegenerative disorder caused by missense mutations affecting the proteolytic domain of AFG3L2, a major component of the mitochondrial m-AAA protease. However, little is known of the underlying pathogenetic mechanisms or how to treat patients with SCA28. Currently available Afg3l2 mutant mice harbour deletions that lead to severe, early-onset neurological phenotypes that do not faithfully reproduce the late-onset and slowly progressing SCA28 phenotype. Here we describe production and detailed analysis of a new knock-in murine model harbouring an Afg3l2 allele carrying the p.Met665Arg patient-derived mutation. Heterozygous mutant mice developed normally but adult mice showed signs of cerebellar ataxia detectable by beam test. Although cerebellar pathology was negative, electrophysiological analysis showed a trend towards increased spontaneous firing in Purkinje cells from heterozygous mutants with respect to wild-type controls. As homozygous mutants died perinatally with evidence of cardiac atrophy, for each genotype we generated mouse embryonic fibroblasts (MEFs) to investigate mitochondrial function. MEFs from mutant mice showed altered mitochondrial bioenergetics, with decreased basal oxygen consumption rate, ATP synthesis and mitochondrial membrane potential. Mitochondrial network formation and morphology was altered, with greatly reduced expression of fusogenic Opa1 isoforms. Mitochondrial alterations were also detected in cerebella of 18-month-old heterozygous mutants and may be a hallmark of disease. Pharmacological inhibition of de novo mitochondrial protein translation with chloramphenicol caused reversal of mitochondrial morphology in homozygous mutant MEFs, supporting the relevance of mitochondrial proteotoxicity for SCA28 pathogenesis and therapy development.
Assuntos
Proteases Dependentes de ATP/genética , ATPases Associadas a Diversas Atividades Celulares/genética , Modelos Animais de Doenças , Mitocôndrias/metabolismo , Ataxias Espinocerebelares/congênito , Animais , Feminino , Técnicas de Introdução de Genes , Potencial da Membrana Mitocondrial , Camundongos Endogâmicos C57BL , Proteínas Mitocondriais/metabolismo , Mutação de Sentido Incorreto , Células de Purkinje/fisiologia , Células de Purkinje/ultraestrutura , Ataxias Espinocerebelares/genética , Ataxias Espinocerebelares/metabolismo , Ataxias Espinocerebelares/patologiaRESUMO
Microglia are highly plastic immune cells which exist in a continuum of activation states. By shaping the function of oligodendrocyte precursor cells (OPCs), the brain cells which differentiate to myelin-forming cells, microglia participate in both myelin injury and remyelination during multiple sclerosis. However, the mode(s) of action of microglia in supporting or inhibiting myelin repair is still largely unclear. Here, we analysed the effects of extracellular vesicles (EVs) produced in vitro by either pro-inflammatory or pro-regenerative microglia on OPCs at demyelinated lesions caused by lysolecithin injection in the mouse corpus callosum. Immunolabelling for myelin proteins and electron microscopy showed that EVs released by pro-inflammatory microglia blocked remyelination, whereas EVs produced by microglia co-cultured with immunosuppressive mesenchymal stem cells promoted OPC recruitment and myelin repair. The molecular mechanisms responsible for the harmful and beneficial EV actions were dissected in primary OPC cultures. By exposing OPCs, cultured either alone or with astrocytes, to inflammatory EVs, we observed a blockade of OPC maturation only in the presence of astrocytes, implicating these cells in remyelination failure. Biochemical fractionation revealed that astrocytes may be converted into harmful cells by the inflammatory EV cargo, as indicated by immunohistochemical and qPCR analyses, whereas surface lipid components of EVs promote OPC migration and/or differentiation, linking EV lipids to myelin repair. Although the mechanisms through which the lipid species enhance OPC maturation still remain to be fully defined, we provide the first demonstration that vesicular sphingosine 1 phosphate stimulates OPC migration, the first fundamental step in myelin repair. From this study, microglial EVs emerge as multimodal and multitarget signalling mediators able to influence both OPCs and astrocytes around myelin lesions, which may be exploited to develop novel approaches for myelin repair not only in multiple sclerosis, but also in neurological and neuropsychiatric diseases characterized by demyelination.
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Astrócitos/fisiologia , Doenças Desmielinizantes/fisiopatologia , Vesículas Extracelulares/fisiologia , Microglia/fisiologia , Bainha de Mielina/fisiologia , Remielinização/fisiologia , Animais , Astrócitos/patologia , Diferenciação Celular/fisiologia , Movimento Celular/fisiologia , Técnicas de Cocultura , Corpo Caloso/patologia , Corpo Caloso/fisiopatologia , Doenças Desmielinizantes/patologia , Modelos Animais de Doenças , Vesículas Extracelulares/patologia , Inflamação/patologia , Inflamação/fisiopatologia , Lisofosfatidilcolinas , Masculino , Células-Tronco Mesenquimais/fisiologia , Camundongos Endogâmicos C57BL , Microglia/patologia , Bainha de Mielina/patologia , Neuroproteção/fisiologia , Células Precursoras de Oligodendrócitos/patologia , Células Precursoras de Oligodendrócitos/fisiologia , Ratos Sprague-DawleyRESUMO
Human genetic studies are rapidly identifying variants that increase risk for neurodevelopmental disorders. However, it remains unclear how specific mutations impact brain function and contribute to neuropsychiatric risk. Chromosome 16p11.2 deletion is one of the most common copy number variations in autism and related neurodevelopmental disorders. Using resting state functional MRI data from the Simons Variation in Individuals Project (VIP) database, we show that 16p11.2 deletion carriers exhibit impaired prefrontal connectivity, resulting in weaker long-range functional coupling with temporal-parietal regions. These functional changes are associated with socio-cognitive impairments. We also document that a mouse with the same genetic deficiency exhibits similarly diminished prefrontal connectivity, together with thalamo-prefrontal miswiring and reduced long-range functional synchronization. These results reveal a mechanistic link between specific genetic risk for neurodevelopmental disorders and long-range functional coupling, and suggest that deletion in 16p11.2 may lead to impaired socio-cognitive function via dysregulation of prefrontal connectivity.
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Transtorno Autístico/genética , Transtornos Cromossômicos/genética , Deficiência Intelectual/genética , Rede Nervosa/fisiologia , Adolescente , Animais , Transtorno Autístico/fisiopatologia , Transtorno Autístico/psicologia , Criança , Deleção Cromossômica , Transtornos Cromossômicos/fisiopatologia , Cromossomos Humanos Par 16/genética , Cognição/fisiologia , Disfunção Cognitiva/complicações , Variações do Número de Cópias de DNA , Modelos Animais de Doenças , Feminino , Humanos , Deficiência Intelectual/fisiopatologia , Imageamento por Ressonância Magnética/métodos , Masculino , Potenciais da Membrana/genética , Potenciais da Membrana/fisiologia , Camundongos , Camundongos Knockout , Transtornos do Neurodesenvolvimento/genética , Córtex Pré-Frontal/fisiologia , Lobo Temporal/fisiopatologiaRESUMO
Recent evidence indicates synaptic dysfunction as an early mechanism affected in neuroinflammatory diseases, such as multiple sclerosis, which are characterized by chronic microglia activation. However, the mode(s) of action of reactive microglia in causing synaptic defects are not fully understood. In this study, we show that inflammatory microglia produce extracellular vesicles (EVs) which are enriched in a set of miRNAs that regulate the expression of key synaptic proteins. Among them, miR-146a-5p, a microglia-specific miRNA not present in hippocampal neurons, controls the expression of presynaptic synaptotagmin1 (Syt1) and postsynaptic neuroligin1 (Nlg1), an adhesion protein which play a crucial role in dendritic spine formation and synaptic stability. Using a Renilla-based sensor, we provide formal proof that inflammatory EVs transfer their miR-146a-5p cargo to neuron. By western blot and immunofluorescence analysis we show that vesicular miR-146a-5p suppresses Syt1 and Nlg1 expression in receiving neurons. Microglia-to-neuron miR-146a-5p transfer and Syt1 and Nlg1 downregulation do not occur when EV-neuron contact is inhibited by cloaking vesicular phosphatidylserine residues and when neurons are exposed to EVs either depleted of miR-146a-5p, produced by pro-regenerative microglia, or storing inactive miR-146a-5p, produced by cells transfected with an anti-miR-146a-5p. Morphological analysis reveals that prolonged exposure to inflammatory EVs leads to significant decrease in dendritic spine density in hippocampal neurons in vivo and in primary culture, which is rescued in vitro by transfection of a miR-insensitive Nlg1 form. Dendritic spine loss is accompanied by a decrease in the density and strength of excitatory synapses, as indicated by reduced mEPSC frequency and amplitude. These findings link inflammatory microglia and enhanced EV production to loss of excitatory synapses, uncovering a previously unrecognized role for microglia-enriched miRNAs, released in association to EVs, in silencing of key synaptic genes.
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Vesículas Extracelulares/imunologia , Inflamação/metabolismo , MicroRNAs/metabolismo , Neuroglia/imunologia , Neurônios/imunologia , Sinapses/imunologia , Animais , Células Cultivadas , Líquido Cefalorraquidiano/metabolismo , Técnicas de Cocultura , Vesículas Extracelulares/patologia , Feminino , Hipocampo/imunologia , Hipocampo/patologia , Humanos , Inflamação/patologia , Masculino , Camundongos Endogâmicos C57BL , Neuroglia/patologia , Plasticidade Neuronal/fisiologia , Neurônios/patologia , Cultura Primária de Células , Ratos Sprague-Dawley , Sinapses/patologiaRESUMO
In the adult mammalian subventricular zone (SVZ), GFAP-positive neural stem cells (NSCs) generate neuroblasts that migrate tangentially along the rostral migratory stream (RMS) toward the olfactory bulb (OB). In the mouse brain, we found that the plasticity inhibitors Nogo-A and Nogo receptor 1 (NgR1) are differentially expressed in the SVZ-OB system, in which Nogo-A identifies immature neuroblasts and NgR1 germinal astrocytes. We therefore examined the role of Nogo-A and NgR1 in the regulation of neurogenesis. Pharmacological experiments show that Nogo-66/NgR1 interaction reduces the proliferation of NSCs. This is consistent with a negative-feedback loop, in which newly generated neurons modulate cell division of SVZ stem cells. Moreover, the Nogo-A-Δ20 domain promotes neuroblast migration toward the OB through activation of the Rho/ROCK (Rho-associated, coiled-coil containing protein kinase) pathway, without the participation of NgR1. Our findings reveal a new unprecedented function for Nogo-A and NgR1 in the homeostatic regulation of the pace of neurogenesis in the adult mouse SVZ and in the migration of neuroblasts along the RMS.
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Movimento Celular/fisiologia , Homeostase/fisiologia , Proteínas da Mielina/fisiologia , Células-Tronco Neurais/fisiologia , Neurogênese/fisiologia , Receptores de Superfície Celular/fisiologia , Animais , Astrócitos/metabolismo , Encéfalo/metabolismo , Encéfalo/fisiologia , Proteínas Ligadas por GPI/antagonistas & inibidores , Proteínas Ligadas por GPI/biossíntese , Proteínas Ligadas por GPI/fisiologia , Camundongos , Proteínas da Mielina/antagonistas & inibidores , Proteínas da Mielina/biossíntese , Células-Tronco Neurais/metabolismo , Proteínas Nogo , Receptor Nogo 1 , Receptores de Superfície Celular/antagonistas & inibidores , Receptores de Superfície Celular/biossíntese , Quinases Associadas a rho/metabolismoRESUMO
Introduction: Medulloblastoma (MB), the most common malignant pediatric brain tumor, is currently treated with surgery followed by radiation and chemotherapy, which is accompanied by severe side effects, raising the need for innovative therapies. Disruption of the microcephaly-related gene Citron kinase (CITK) impairs the expansion of xenograft models as well as spontaneous MB arising in transgenic mice. No specific CITK inhibitors are available. Methods: Lestaurtinib, a Staurosporine derivative also known as CEP-701, inhibits CITK with IC50 of 90 nM. We therefore tested the biological effects of this molecule on different MB cell lines, as well as in vivo, injecting the drug in MBs arising in SmoA1 transgenic mice. Results: Similar to CITK knockdown, treatment of MB cells with 100 nM Lestaurtinib reduces phospho-INCENP levels at the midbody and leads to late cytokinesis failure. Moreover, Lestaurtinib impairs cell proliferation through CITK-sensitive mechanisms. These phenotypes are accompanied by accumulation of DNA double strand breaks, cell cycle block and TP53 superfamily activation in vitro and in vivo. Lestaurtinib treatment reduces tumor growth and increases mice survival. Discussion: Our data indicate that Lestaurtinib produces in MB cells poly-pharmacological effects extending beyond the inhibition of its validated targets, supporting the possibility of repositioning this drug for MB treatment.
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BACKGROUND: Huntington's disease (HD) is a motor and cognitive neurodegenerative disorder due to prominent loss of striatal medium spiny neurons (MSNs). Cell replacement using human embryonic stem cells (hESCs) derivatives may offer new therapeutic opportunities to replace degenerated neurons and repair damaged circuits. METHODS: With the aim to develop effective cell replacement for HD, we assessed the long-term therapeutic value of hESC-derived striatal progenitors by grafting the cells into the striatum of a preclinical model of HD [i.e., adult immunodeficient rats in which the striatum was lesioned by monolateral injection of quinolinic acid (QA)]. We examined the survival, maturation, self-organization and integration of the graft as well as its impact on lesion-dependent motor alterations up to 6 months post-graft. Moreover, we tested whether exposing a cohort of QA-lesioned animals to environmental enrichment (EE) could improve graft integration and function. RESULTS: Human striatal progenitors survived up to 6 months after transplantation and showed morphological and neurochemical features typical of human MSNs. Donor-derived interneurons were also detected. Grafts wired in both local and long-range striatal circuits, formed domains suggestive of distinct ganglionic eminence territories and displayed emerging striosome features. Moreover, over time grafts improved complex motor performances affected by QA. EE selectively increased cell differentiation into MSN phenotype and promoted host-to-graft connectivity. However, when combined to the graft, the EE paradigm used in this study was insufficient to produce an additive effect on task execution. CONCLUSIONS: The data support the long-term therapeutic potential of ESC-derived human striatal progenitor grafts for the replacement of degenerated striatal neurons in HD and suggest that EE can effectively accelerate the maturation and promote the integration of human striatal cells.
Assuntos
Transplante de Tecido Encefálico , Células-Tronco Embrionárias Humanas , Doença de Huntington , Ratos , Animais , Humanos , Doença de Huntington/terapia , Corpo Estriado/fisiologia , Neurônios , Modelos Animais de DoençasRESUMO
In the developing mouse forebrain, temporally distinct waves of oligodendrocyte progenitor cells (OPCs) arise from different germinal zones and eventually populate either dorsal or ventral regions, where they present as transcriptionally and functionally equivalent cells. Despite that, developmental heterogeneity influences adult OPC responses upon demyelination. Here we show that accumulation of DNA damage due to ablation of citron-kinase or cisplatin treatment cell-autonomously disrupts OPC fate, resulting in cell death and senescence in the dorsal and ventral subsets, respectively. Such alternative fates are associated with distinct developmental origins of OPCs, and with a different activation of NRF2-mediated anti-oxidant responses. These data indicate that, upon injury, dorsal and ventral OPC subsets show functional and molecular diversity that can make them differentially vulnerable to pathological conditions associated with DNA damage.
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Células Precursoras de Oligodendrócitos , Animais , Dano ao DNA , Camundongos , Células Precursoras de Oligodendrócitos/fisiologia , Oligodendroglia/metabolismo , ProsencéfaloRESUMO
OBJECTIVES: The aim of this study is to quantitatively investigate, at the preclinical level, the extent of Gd retention in the CNS, and peripheral organs, of immune-mediated murine models (Experimental Autoimmune Encephalomyelitis -EAE) of Multiple Sclerosis, compared to control animals, upon the injection of gadodiamide. The influence of the Gadolinium Based Contrast Agent administration timing during the course of EAE development is also monitored. METHODS: EAE mice were injected with three doses (1.2 mmol/kg each) of gadodiamide at three different time points during the EAE development and sacrificed after 21 or 39 days. Organs were collected and the amount of Gd was quantified through Inductively Coupled Plasma-Mass Spectrometry. Transmission electron microscopy (TEM) and MRI techniques were applied to add spatial and qualitative information to the obtained results. RESULTS: In the spinal cord of EAE group, 21 days after gadodiamide administration, a significantly higher accumulation of Gd occurred. Conversely, in the encephalon, a lower amount of Gd retention was reached, even if differences emerged between EAE and controls mice. After 39 days, the amounts of retained Gd markedly decreased. TEM validated the presence of Gd in CNS. MRI of the encephalon at 7.1T did not highlight any hyper intense region. CONCLUSION: In the spinal cord of EAE mice, which is the mostly damaged region in this specific animal model, a preferential but transient accumulation of Gd is observed. In the encephalon, the Gd retention could be mostly related to inflammation occurring upon immunization rather than to demyelination.
Assuntos
Encefalomielite Autoimune Experimental , Esclerose Múltipla , Compostos Organometálicos , Animais , Modelos Animais de Doenças , Gadolínio , Camundongos , Camundongos Endogâmicos C57BL , Esclerose Múltipla/diagnóstico por imagemRESUMO
Epidemiological studies show a strong association between exposure to air pollution - and particularly to particulate matter (PM) -, increased prevalence of Multiple Sclerosis (MS) and higher rates of hospital admissions for MS and MS relapses. Besides having immunomodulatory effects and sustaining a systemic oxidative-inflammatory response, PM may participate in MS pathogenesis by targeting also Central Nervous System (CNS)-specific processes, such as myelin repair. Here we show that, in a mouse model of lysolecithin-induced demyelination of the subcortical white matter, post-injury exposure to fine PM hampers remyelination, disturbs oligodendroglia differentiation dynamics and promotes astroglia and microglia reactivity. These findings support the view that exposure to fine PM can contribute to demyelinating pathologies by targeting the endogenous regenerative capability of the CNS tissue.
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Doenças Desmielinizantes/induzido quimicamente , Doenças Desmielinizantes/patologia , Modelos Animais de Doenças , Bainha de Mielina/patologia , Material Particulado/toxicidade , Substância Branca/patologia , Animais , Diferenciação Celular/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Material Particulado/administração & dosagem , Traqueia/patologiaRESUMO
During Central Nervous System ontogenesis, myelinating oligodendrocytes (OLs) arise from highly ramified and proliferative precursors called oligodendrocyte progenitor cells (OPCs). OPC architecture, proliferation and oligodendro-/myelino-genesis are finely regulated by the interplay of cell-intrinsic and extrinsic factors. A variety of extrinsic cues converge on the extracellular signal-regulated kinase/mitogen activated protein kinase (ERK/MAPK) pathway. Here we found that the germinal ablation of the MAPK c-Jun N-Terminal Kinase isoform 1 (JNK1) results in a significant reduction of myelin in the cerebral cortex and corpus callosum at both postnatal and adult stages. Myelin alterations are accompanied by higher OPC density and proliferation during the first weeks of life, consistent with a transient alteration of mechanisms regulating OPC self-renewal and differentiation. JNK1 KO OPCs also show smaller occupancy territories and a less complex branching architecture in vivo. Notably, these latter phenotypes are recapitulated in pure cultures of JNK1 KO OPCs and of WT OPCs treated with the JNK inhibitor D-JNKI-1. Moreover, JNK1 KO and WT D-JNKI-1 treated OLs, while not showing overt alterations of differentiation in vitro, display a reduced surface compared to controls. Our results unveil a novel player in the complex regulation of OPC biology, on the one hand showing that JNK1 ablation cell-autonomously determines alterations of OPC proliferation and branching architecture and, on the other hand, suggesting that JNK1 signaling in OLs participates in myelination in vivo.
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Proliferação de Células , Sistema de Sinalização das MAP Quinases , Proteína Quinase 8 Ativada por Mitógeno/metabolismo , Bainha de Mielina/metabolismo , Células Precursoras de Oligodendrócitos/enzimologia , Oligodendroglia/enzimologia , Animais , Camundongos , Camundongos Knockout , Proteína Quinase 8 Ativada por Mitógeno/genética , Bainha de Mielina/genéticaRESUMO
Brain structural plasticity is an extraordinary tool that allows the mature brain to adapt to environmental changes, to learn, to repair itself after lesions or disease, and to slow aging. A long history of neuroscience research led to fascinating discoveries of different types of plasticity, involving changes in the genetically determined structure of nervous tissue, up to the ultimate dream of neuronal replacement: a stem cell-driven "adult neurogenesis" (AN). Yet, this road does not seem a straight one, since mutable dogmas, conflicting results and conflicting interpretations continue to warm the field. As a result, after more than 10,000 papers published on AN, we still do not know its time course, rate or features with respect to other kinds of structural plasticity in our brain. The solution does not appear to be behind the next curve, as differences among mammals reveal a very complex landscape that cannot be easily understood from rodents models alone. By considering evolutionary aspects, some pitfalls in the interpretation of cell markers, and a novel population of undifferentiated cells that are not newly generated [immature neurons (INs)], we address some conflicting results and controversies in order to find the right road forward. We suggest that considering plasticity in a comparative framework might help assemble the evolutionary, anatomical and functional pieces of a very complex biological process with extraordinary translational potential.
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Huntington disease (HD) is an inherited late-onset neurological disorder characterized by progressive neuronal loss and disruption of cortical and basal ganglia circuits. Cell replacement using human embryonic stem cells may offer the opportunity to repair the damaged circuits and significantly ameliorate disease conditions. Here, we showed that in-vitro-differentiated human striatal progenitors undergo maturation and integrate into host circuits upon intra-striatal transplantation in a rat model of HD. By combining graft-specific immunohistochemistry, rabies virus-mediated synaptic tracing, and ex vivo electrophysiology, we showed that grafts can extend projections to the appropriate target structures, including the globus pallidus, the subthalamic nucleus, and the substantia nigra, and receive synaptic contact from both host and graft cells with 6.6 ± 1.6 inputs cell per transplanted neuron. We have also shown that transplants elicited a significant improvement in sensory-motor tasks up to 2 months post-transplant further supporting the therapeutic potential of this approach.
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Corpo Estriado/citologia , Células-Tronco Embrionárias Humanas/transplante , Doença de Huntington/terapia , Células-Tronco Neurais/transplante , Transplante de Células-Tronco/métodos , Animais , Células Cultivadas , Corpo Estriado/fisiologia , Células-Tronco Embrionárias Humanas/citologia , Humanos , Locomoção , Masculino , Células-Tronco Neurais/citologia , Neurogênese , Ratos , Regeneração , Sensação , Substância Negra/citologia , Substância Negra/fisiologia , Núcleo Subtalâmico/citologia , Núcleo Subtalâmico/fisiologia , Sinapses/metabolismo , Sinapses/fisiologiaRESUMO
Since a growing number of studies based on the real-time reverse transcriptase polymerase chain reaction (RT-PCR) continue to be published in order to highlight genes specifically involved in brain development, maturation, and function, the identification of reference genes suitable for this kind of experiments is now an urgent need in the neuroscience field. The aim of this work was to verify the suitability of some very common housekeeping genes (such as Gapdh, 18s, and B2m) and of some relatively new control genes (such as Pgk1, Tfrc, and Gusb) during mouse brain maturation. We tested the candidate reference genes in mouse whole brain, cerebellum, brain stem, hippocampus, medial septum, frontal neocortex, and olfactory bulb. Moreover, we reported the first complete study of Pgk1 expression throughout the development and the aging of mouse brain. Although no tested gene showed to be the optimal reference for all mouse brain regions, in general, the new housekeeping genes were highly stable in most of the analyzed regions. Above all, with few exceptions, Pgk1 showed to be a reliable control for the analyzed mouse brain regions during development, maturation, and aging.
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Encéfalo/fisiologia , Expressão Gênica , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Animais , Encéfalo/anatomia & histologia , Feminino , Perfilação da Expressão Gênica , Camundongos , Dados de Sequência Molecular , Gravidez , Padrões de Referência , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/normasRESUMO
Pre-clinical research is carried out on animal models, mostly laboratory rodents, with the ultimate aim of translating the acquired knowledge to humans. In the last decades, adult neurogenesis (AN) has been intensively studied since it is viewed as a tool for fostering brain plasticity, possibly repair. Yet, occurrence, location, and rate of AN vary among mammals: the capability for constitutive neuronal production is substantially reduced when comparing small-brained, short living (laboratory rodents) and large-brained, long-living species (humans, dolphins). Several difficulties concerning scarce availability of fresh tissues, technical limits and ethical concerns did contribute in delaying and diverting the achievement of the picture of neurogenic plasticity in large-brained mammals. Some reports appeared in the last few years, starting to shed more light on this issue. Despite technical limits, data from recent studies mostly converge to indicate that neurogenesis is vestigial, or possibly absent, in regions of the adult human brain where in rodents neuronal addition continues into adult life. Analyses carried out in dolphins, mammals devoid of olfaction, but descendant of ancestors provided with olfaction, has shown disappearance of neurogenesis in both neonatal and adult individuals. Heterogeneity in mammalian structural plasticity remains largely underestimated by scientists focusing their research in rodents. Comparative studies are the key to understand the function of AN and the possible translational significance of neuronal replacement in humans. Here, we summarize comparative studies on AN and discuss the evolutionary implications of variations on the recruitment of new neurons in different regions and different species.
RESUMO
The cytoskeletal protein doublecortin (DCX) is a marker for neuronal cells retaining high potential for structural plasticity, originating from both embryonic and adult neurogenic processes. Some of these cells have been described in the subcortical white matter of neonatal and postnatal mammals. In mice and humans it has been shown they are young neurons migrating through the white matter after birth, reaching the cortex in a sort of protracted neurogenesis. Here we show that DCX+ cells in the white matter of neonatal and young Cetartiodactyla (dolphin and sheep) form large clusters which are not newly generated (in sheep, and likely neither in dolphins) and do not reach the cortical layers, rather appearing "trapped" in the white matter tissue. No direct contact or continuity can be observed between the subventricular zone region and the DCX+ clusters, thus indicating their independence from any neurogenic source (in dolphins further confirmed by the recent demonstration that periventricular neurogenesis is inactive since birth). Cetartiodactyla include two orders of large-brained, relatively long-living mammals (cetaceans and artiodactyls) which were recognized as two separate monophyletic clades until recently, yet, despite the evident morphological distinctions, they are monophyletic in origin. The brain of Cetartiodactyla is characterized by an advanced stage of development at birth, a feature that might explain the occurrence of "static" cell clusters confined within their white matter. These results further confirm the existence of high heterogeneity in the occurrence, distribution and types of structural plasticity among mammals, supporting the emerging view that multiple populations of DCX+, non-newly generated cells can be abundant in large-brained, long-living species.
Assuntos
Encéfalo/citologia , Encéfalo/crescimento & desenvolvimento , Proteínas Associadas aos Microtúbulos/metabolismo , Neuropeptídeos/metabolismo , Substância Branca/citologia , Substância Branca/crescimento & desenvolvimento , Anatomia Comparada , Animais , Golfinho Nariz-de-Garrafa , Encéfalo/metabolismo , Movimento Celular , Proliferação de Células , Proteínas do Domínio Duplacortina , Proteína Duplacortina , Feminino , Masculino , Neurogênese , Carneiro Doméstico , Especificidade da Espécie , Stenella , Substância Branca/metabolismoRESUMO
Adult neurogenesis has been implicated in brain plasticity and brain repair. In mammals, it is mostly restricted to specific brain regions and specific physiological functions. The function and evolutionary history of mammalian adult neurogenesis has been elusive so far. The largest neurogenic site in mammals (subventricular zone, SVZ) generates neurons destined to populate the olfactory bulb. The SVZ neurogenic activity appears to be related to the dependence of the species on olfaction since it occurs at high rates throughout life in animals strongly dependent on this function for their survival. Indeed, it dramatically decreases in humans, who do not depend so much on it. This study investigates whether the SVZ neurogenic site exists in mammals devoid of olfaction and olfactory brain structures, such as dolphins. Our results demonstate that a small SVZ-like region persists in these aquatic mammals. However, this region seems to have lost its neurogenic capabilities since neonatal stages. In addition, instead of the typical newly generated neuroblasts, some mature neurons were observed in the dolphin SVZ. Since cetaceans evolved from terrestrial ancestors, non-neurogenic SVZ may indicate extinction of adult neurogenesis in the absence of olfactory function, with the retention of an SVZ-like anatomical region either vestigial or of still unknown role.