RESUMEN
Aberrant aggregation of the RNA-binding protein TDP-43 in neurons is a hallmark of frontotemporal lobar degeneration caused by haploinsufficiency in the gene encoding progranulin1,2. However, the mechanism leading to TDP-43 proteinopathy remains unclear. Here we use single-nucleus RNA sequencing to show that progranulin deficiency promotes microglial transition from a homeostatic to a disease-specific state that causes endolysosomal dysfunction and neurodegeneration in mice. These defects persist even when Grn-/- microglia are cultured ex vivo. In addition, single-nucleus RNA sequencing reveals selective loss of excitatory neurons at disease end-stage, which is characterized by prominent nuclear and cytoplasmic TDP-43 granules and nuclear pore defects. Remarkably, conditioned media from Grn-/- microglia are sufficient to promote TDP-43 granule formation, nuclear pore defects and cell death in excitatory neurons via the complement activation pathway. Consistent with these results, deletion of the genes encoding C1qa and C3 mitigates microglial toxicity and rescues TDP-43 proteinopathy and neurodegeneration. These results uncover previously unappreciated contributions of chronic microglial toxicity to TDP-43 proteinopathy during neurodegeneration.
Asunto(s)
Microglía/metabolismo , Microglía/patología , Neuronas/metabolismo , Neuronas/patología , Progranulinas/deficiencia , Proteinopatías TDP-43/metabolismo , Proteinopatías TDP-43/patología , Envejecimiento/genética , Envejecimiento/patología , Animales , Núcleo Celular/genética , Núcleo Celular/patología , Activación de Complemento/efectos de los fármacos , Activación de Complemento/inmunología , Complemento C1q/antagonistas & inhibidores , Complemento C1q/inmunología , Complemento C3b/antagonistas & inhibidores , Complemento C3b/inmunología , Medios de Cultivo Condicionados/química , Medios de Cultivo Condicionados/farmacología , Proteínas de Unión al ADN/metabolismo , Modelos Animales de Enfermedad , Femenino , Masculino , Ratones , Poro Nuclear/metabolismo , Poro Nuclear/patología , Progranulinas/genética , RNA-Seq , Análisis de la Célula Individual , Proteinopatías TDP-43/tratamiento farmacológico , Proteinopatías TDP-43/genética , Tálamo/metabolismo , Tálamo/patología , TranscriptomaRESUMEN
Collagen type IV alpha 1 and alpha 2 chains form heterotrimers ([α1(IV)]2α2(IV)) that represent a fundamental basement membrane constituent. Dominant COL4A1 and COL4A2 mutations cause a multisystem disorder that is marked by clinical heterogeneity and variable expressivity and that is generally characterized by the presence of cerebrovascular disease with ocular, renal, and muscular involvement. Despite the fact that muscle pathology is reported in up to one-third of individuals with COL4A1 and COL4A2 mutations and in animal models with mutations in COL4A1 and COL4A2 orthologs, the pathophysiological mechanisms underlying COL4A1-related myopathy are unknown. In general, mutations are thought to impair [α1(IV)]2α2(IV) secretion. Whether pathogenesis results from intracellular retention, extracellular deficiency, or the presence of mutant proteins in basement membranes represents an important gap in knowledge and a major obstacle for developing targeted interventions. We report that Col4a1 mutant mice develop progressive neuromuscular pathology that models human disease. We demonstrate that independent muscular, neural, and vascular insults contribute to neuromyopathy and that there is mechanistic heterogeneity among tissues. Importantly, we provide evidence of a COL4A1 functional subdomain with disproportionate significance for tissue-specific pathology and demonstrate that a potential therapeutic strategy aimed at promoting [α1(IV)]2α2(IV) secretion can ameliorate or exacerbate myopathy in a mutation-dependent manner. These data have important translational implications for prediction of clinical outcomes based on genotype, development of mechanism-based interventions, and genetic stratification for clinical trials. Collectively, our data underscore the importance of the [α1(IV)]2α2(IV) network as a multifunctional signaling platform and show that allelic and tissue-specific mechanistic heterogeneities contribute to the variable expressivity of COL4A1 and COL4A2 mutations.
Asunto(s)
Colágeno Tipo IV/genética , Enfermedades Musculares/etiología , Mutación , Enfermedades Neuromusculares/etiología , Animales , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Enfermedades Musculares/patología , Enfermedades Neuromusculares/patología , Especificidad de Órganos , FenotipoRESUMEN
Human astrocytes provide trophic as well as structural support to the surrounding brain cells. Furthermore, they have been implicated in many physiological processes important for central nervous system function. Traditionally astrocytes have been considered to be a homogeneous class of cells, however, it has increasingly become more evident that astrocytes can have very different characteristics in different regions of the brain, or even within the same region. In this review we will discuss the features of human astrocytes, their heterogeneity, and their generation during neurodevelopment and the extraordinary progress that has been made to model these fascinating cells in vitro, mainly from induced pluripotent stem cells. Astrocytes' role in disease will also be discussed with a particular focus on their role in neurodegenerative disorders. As outlined here, astrocytes are important for the homeostasis of the central nervous system and understanding their regional specificity is a priority to elucidate the complexity of the human brain.
Asunto(s)
Astrocitos/fisiología , Encéfalo/fisiología , Diferenciación Celular/fisiología , Enfermedades Neurodegenerativas/patología , Astrocitos/citología , Encéfalo/citología , HumanosRESUMEN
Astrocytes, the most abundant cells in the central nervous system, promote synapse formation and help to refine neural connectivity. Although they are allocated to spatially distinct regional domains during development, it is unknown whether region-restricted astrocytes are functionally heterogeneous. Here we show that postnatal spinal cord astrocytes express several region-specific genes, and that ventral astrocyte-encoded semaphorin 3a (Sema3a) is required for proper motor neuron and sensory neuron circuit organization. Loss of astrocyte-encoded Sema3a leads to dysregulated α-motor neuron axon initial segment orientation, markedly abnormal synaptic inputs, and selective death of α- but not of adjacent γ-motor neurons. In addition, a subset of TrkA(+) sensory afferents projects to ectopic ventral positions. These findings demonstrate that stable maintenance of a positional cue by developing astrocytes influences multiple aspects of sensorimotor circuit formation. More generally, they suggest that regional astrocyte heterogeneity may help to coordinate postnatal neural circuit refinement.
Asunto(s)
Astrocitos/fisiología , Neuronas Motoras/fisiología , Vías Nerviosas/fisiología , Células Receptoras Sensoriales/fisiología , Animales , Astrocitos/citología , Axones/fisiología , Polaridad Celular , Supervivencia Celular/efectos de los fármacos , Humanos , Ratones , Neuronas Motoras/citología , Neuronas Motoras/efectos de los fármacos , Semaforina-3A/deficiencia , Semaforina-3A/genética , Semaforina-3A/metabolismo , Semaforina-3A/farmacología , Células Receptoras Sensoriales/citología , Médula Espinal/citología , Sinapsis/metabolismoRESUMEN
Astrocytes are no longer seen as a homogenous population of cells. In fact, recent studies indicate that astrocytes are morphologically and functionally diverse and play critical roles in neurodevelopmental diseases such as Rett syndrome and fragile X mental retardation. This review summarizes recent advances in astrocyte development, including the role of neural tube patterning in specification and developmental functions of astrocytes during synaptogenesis. We propose here that a precise understanding of astrocyte development is critical to defining heterogeneity and could lead advances in understanding and treating a variety of neuropsychiatric diseases.
Asunto(s)
Astrocitos/fisiología , Trastornos Heredodegenerativos del Sistema Nervioso/etiología , Trastornos Mentales/etiología , Neurogénesis , Astrocitos/patología , Trastornos Heredodegenerativos del Sistema Nervioso/patología , Humanos , Trastornos Mentales/patología , Células-Madre Neurales/patología , Células-Madre Neurales/fisiologíaRESUMEN
Hypoxia can injure brain white matter tracts, comprised of axons and myelinating oligodendrocytes, leading to cerebral palsy in neonates and delayed post-hypoxic leukoencephalopathy (DPHL) in adults. In these conditions, white matter injury can be followed by myelin regeneration, but myelination often fails and is a significant contributor to fixed demyelinated lesions, with ensuing permanent neurological injury. Non-myelinating oligodendrocyte precursor cells are often found in lesions in plentiful numbers, but fail to mature, suggesting oligodendrocyte precursor cell differentiation arrest as a critical contributor to failed myelination in hypoxia. We report a case of an adult patient who developed the rare condition DPHL and made a nearly complete recovery in the setting of treatment with clemastine, a widely available antihistamine that in preclinical models promotes oligodendrocyte precursor cell differentiation. This suggested possible therapeutic benefit in the more clinically prevalent hypoxic injury of newborns, and we demonstrate in murine neonatal hypoxic injury that clemastine dramatically promotes oligodendrocyte precursor cell differentiation, myelination, and improves functional recovery. We show that its effect in hypoxia is oligodendroglial specific via an effect on the M1 muscarinic receptor on oligodendrocyte precursor cells. We propose clemastine as a potential therapy for hypoxic brain injuries associated with white matter injury and oligodendrocyte precursor cell maturation arrest.
Asunto(s)
Clemastina/uso terapéutico , Enfermedades Desmielinizantes/tratamiento farmacológico , Enfermedades Desmielinizantes/etiología , Antagonistas de los Receptores Histamínicos H1/uso terapéutico , Hipoxia Encefálica/complicaciones , Recuperación de la Función/efectos de los fármacos , Potenciales de Acción/efectos de los fármacos , Animales , Animales Recién Nacidos , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Cerebelo/efectos de los fármacos , Cerebelo/metabolismo , Cerebelo/ultraestructura , Enfermedades Desmielinizantes/diagnóstico por imagen , Enfermedades Desmielinizantes/patología , Modelos Animales de Enfermedad , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Humanos , Hipoxia Encefálica/diagnóstico por imagen , Masculino , Ratones , Ratones Noqueados , Persona de Mediana Edad , Vaina de Mielina/efectos de los fármacos , Vaina de Mielina/ultraestructura , Células Precursoras de Oligodendrocitos/efectos de los fármacos , Nervio Óptico/fisiopatología , Oxígeno/farmacología , Receptor Muscarínico M1/genética , Receptor Muscarínico M1/metabolismoRESUMEN
The rapid spread of Zika virus (ZIKV) and its association with abnormal brain development constitute a global health emergency. Congenital ZIKV infection produces a range of mild to severe pathologies, including microcephaly. To understand the pathophysiology of ZIKV infection, we used models of the developing brain that faithfully recapitulate the tissue architecture in early to midgestation. We identify the brain cell populations that are most susceptible to ZIKV infection in primary human tissue, provide evidence for a mechanism of viral entry, and show that a commonly used antibiotic protects cultured brain cells by reducing viral proliferation. In the brain, ZIKV preferentially infected neural stem cells, astrocytes, oligodendrocyte precursor cells, and microglia, whereas neurons were less susceptible to infection. These findings suggest mechanisms for microcephaly and other pathologic features of infants with congenital ZIKV infection that are not explained by neural stem cell infection alone, such as calcifications in the cortical plate. Furthermore, we find that blocking the glia-enriched putative viral entry receptor AXL reduced ZIKV infection of astrocytes in vitro, and genetic knockdown of AXL in a glial cell line nearly abolished infection. Finally, we evaluate 2,177 compounds, focusing on drugs safe in pregnancy. We show that the macrolide antibiotic azithromycin reduced viral proliferation and virus-induced cytopathic effects in glial cell lines and human astrocytes. Our characterization of infection in the developing human brain clarifies the pathogenesis of congenital ZIKV infection and provides the basis for investigating possible therapeutic strategies to safely alleviate or prevent the most severe consequences of the epidemic.
Asunto(s)
Azitromicina/farmacología , Encéfalo/embriología , Encéfalo/virología , Tropismo Viral/efectos de los fármacos , Infección por el Virus Zika/tratamiento farmacológico , Virus Zika/efectos de los fármacos , Virus Zika/fisiología , Encéfalo/patología , Línea Celular , Efecto Citopatogénico Viral/efectos de los fármacos , Femenino , Humanos , Recién Nacido , Pruebas de Sensibilidad Microbiana , Microcefalia/tratamiento farmacológico , Microcefalia/embriología , Microcefalia/patología , Neuroglía/efectos de los fármacos , Neuroglía/patología , Neuroglía/virología , Embarazo , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/fisiología , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Proteínas Tirosina Quinasas Receptoras/fisiología , Tropismo Viral/fisiología , Internalización del Virus/efectos de los fármacos , Replicación Viral/efectos de los fármacos , Virus Zika/patogenicidad , Infección por el Virus Zika/embriología , Infección por el Virus Zika/patología , Tirosina Quinasa del Receptor AxlRESUMEN
This report summarizes and highlights the fifth International RASopathies Symposium: When Development and Cancer Intersect, held in Orlando, Florida in July 2017. The RASopathies comprise a recognizable pattern of malformation syndromes that are caused by germ line mutations in genes that encode components of the RAS/mitogen-activated protein kinase (MAPK) pathway. Because of their common underlying pathogenetic etiology, there is significant overlap in their phenotypic features, which includes craniofacial dysmorphology, cardiac, cutaneous, musculoskeletal, gastrointestinal and ocular abnormalities, neurological and neurocognitive issues, and a predisposition to cancer. The RAS pathway is a well-known oncogenic pathway that is commonly found to be activated in somatic malignancies. As in somatic cancers, the RASopathies can be caused by various pathogenetic mechanisms that ultimately impact or alter the normal function and regulation of the MAPK pathway. As such, the RASopathies represent an excellent model of study to explore the intersection of the effects of dysregulation and its consequence in both development and oncogenesis.
Asunto(s)
Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Proteínas ras/genética , Animales , Regulación de la Expresión Génica , Estudios de Asociación Genética/métodos , Desarrollo Humano , Humanos , Modelos Biológicos , Terapia Molecular Dirigida , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patología , Organogénesis/genética , Transducción de Señal , Síndrome , Proteínas ras/metabolismoRESUMEN
UNLABELLED: Key issues concerning ganglion cell type-specific loss and synaptic changes in animal models of experimental glaucoma remain highly debated. Importantly, changes in the structure and function of various RGC types that occur early, within 14 d after acute, transient intraocular pressure elevation, have not been previously assessed. Using biolistic transfection of individual RGCs and multielectrode array recordings to measure light responses in mice, we examined the effects of laser-induced ocular hypertension on the structure and function of a subset of RGCs. Among the α-like RGCs studied, αOFF-transient RGCs exhibited higher rates of cell death, with corresponding reductions in dendritic area, dendritic complexity, and synapse density. Functionally, OFF-transient RGCs displayed decreases in spontaneous activity and receptive field size. In contrast, neither αOFF-sustained nor αON-sustained RGCs displayed decreases in light responses, although they did exhibit a decrease in excitatory postsynaptic sites, suggesting that synapse loss may be one of the earliest signs of degeneration. Interestingly, presynaptic ribbon density decreased to a greater degree in the OFF sublamina of the inner plexiform layer, corroborating the hypothesis that RGCs with dendrites stratifying in the OFF sublamina may be damaged early. Indeed, OFF arbors of ON-OFF RGCs lose complexity more rapidly than ON arbors. Our results reveal type-specific differences in RGC responses to injury with a selective vulnerability of αOFF-transient RGCs, and furthermore, an increased susceptibility of synapses in the OFF sublamina. The selective vulnerability of specific RGC types offers new avenues for the design of more sensitive functional tests and targeted neuroprotection. SIGNIFICANCE STATEMENT: Conflicting reports regarding the selective vulnerability of specific retinal ganglion cell (RGC) types in glaucoma exist. We examine, for the first time, the effects of transient intraocular pressure elevation on the structure and function of various RGC types. Among the α-like RGCs studied, αOFF-transient RGCs are the most vulnerable to transient transient intraocular pressure elevation as measured by rates of cell death, morphologic alterations in dendrites and synapses, and physiological dysfunction. Specifically, we found that presynaptic ribbon density decreased to a greater degree in the OFF sublamina of the inner plexiform layer. Our results suggest selective vulnerability both of specific types of RGCs and of specific inner plexiform layer sublaminae, opening new avenues for identifying novel diagnostic and treatment targets in glaucoma.
Asunto(s)
Presión Intraocular/fisiología , Hipertensión Ocular/patología , Células Ganglionares de la Retina/patología , Sinapsis/patología , Oxidorreductasas de Alcohol/metabolismo , Animales , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Colina O-Acetiltransferasa/metabolismo , Proteínas de Unión al ADN/metabolismo , Modelos Animales de Enfermedad , Homólogo 4 de la Proteína Discs Large , Potenciales Evocados/fisiología , Femenino , Guanilato-Quinasas/metabolismo , Presión Intraocular/genética , Rayos Láser/efectos adversos , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Proteínas de la Membrana/metabolismo , Ratones , Proteínas de Neurofilamentos , Hipertensión Ocular/etiología , Estimulación Luminosa , Células Ganglionares de la Retina/fisiología , Estadísticas no Paramétricas , Sinapsis/fisiología , Factores de Tiempo , Transducción GenéticaRESUMEN
UNLABELLED: Myelin controls the time required for an action potential to travel from the neuronal soma to the axon terminal, defining the temporal manner in which information is processed within the CNS. The presence of myelin, the internodal length, and the thickness of the myelin sheath are powerful structural factors that control the velocity and fidelity of action potential transmission. Emerging evidence indicates that myelination is sensitive to environmental experience and neuronal activity. Activity-dependent modulation of myelination can dynamically alter action potential conduction properties but direct functional in vivo evidence and characterization of the underlying myelin changes is lacking. We demonstrate that in mice long-term monocular deprivation increases oligodendrogenesis in the retinogeniculate pathway but shortens myelin internode lengths without affecting other structural properties of myelinated fibers. We also demonstrate that genetically attenuating synaptic glutamate neurotransmission from retinal ganglion cells phenocopies the changes observed after monocular deprivation, suggesting that glutamate may constitute a signal for myelin length regulation. Importantly, we demonstrate that visual deprivation and shortened internodes are associated with a significant reduction in nerve conduction velocity in the optic nerve. Our results reveal the importance of sensory input in the building of myelinated fibers and suggest that this activity-dependent alteration of myelination is important for modifying the conductive properties of brain circuits in response to environmental experience. SIGNIFICANCE STATEMENT: Oligodendrocyte precursor cells differentiate into mature oligodendrocytes and are capable of ensheathing axons with myelin without molecular cues from neurons. However, this default myelination process can be modulated by changes in neuronal activity. Here, we show, for the first time, that experience-dependent activity modifies the length of myelin internodes along axons altering action potential conduction velocity. Such a mechanism would allow for variations in conduction velocities that provide a degree of plasticity in accordance to environmental needs. It will be important in future work to investigate how these changes in myelination and conduction velocity contribute to signal integration in postsynaptic neurons and circuit function.
Asunto(s)
Fibras Nerviosas Mielínicas/fisiología , Conducción Nerviosa/fisiología , Nervio Óptico/fisiología , Visión Monocular/fisiología , Vías Visuales/fisiología , Potenciales de Acción/fisiología , Factores de Edad , Animales , Animales Recién Nacidos , Antígenos/genética , Antígenos/metabolismo , Toxina del Cólera/metabolismo , Cuerpos Geniculados/citología , Cuerpos Geniculados/fisiología , Cuerpos Geniculados/ultraestructura , Proteínas con Homeodominio LIM/genética , Proteínas con Homeodominio LIM/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Vaina de Mielina/metabolismo , Vaina de Mielina/ultraestructura , Fibras Nerviosas Mielínicas/ultraestructura , Conducción Nerviosa/genética , Nervio Óptico/ultraestructura , Organogénesis/genética , Organogénesis/fisiología , Estimulación Luminosa , Proteoglicanos/genética , Proteoglicanos/metabolismo , Células Ganglionares de la Retina/metabolismo , Transmisión Sináptica/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Proteína 2 de Transporte Vesicular de Glutamato/genética , Proteína 2 de Transporte Vesicular de Glutamato/metabolismo , Vías Visuales/ultraestructuraRESUMEN
Increasing evidence implicates abnormal Ras signaling as a major contributor in neurodevelopmental disorders, yet how such signaling causes cortical pathogenesis is unknown. We examined the consequences of aberrant Ras signaling in the developing mouse brain and uncovered several critical phenotypes, including increased production of cortical neurons and morphological deficits. To determine whether these phenotypes are recapitulated in humans, we generated induced pluripotent stem (iPS) cell lines from patients with Costello syndrome (CS), a developmental disorder caused by abnormal Ras signaling and characterized by neurodevelopmental abnormalities, such as cognitive impairment and autism. Directed differentiation toward a neuroectodermal fate revealed an extended progenitor phase and subsequent increased production of cortical neurons. Morphological analysis of mature neurons revealed significantly altered neurite length and soma size in CS patients. This study demonstrates the synergy between mouse and human models and validates the use of iPS cells as a platform to study the underlying cellular pathologies resulting from signaling deficits. SIGNIFICANCE STATEMENT: Increasing evidence implicates Ras signaling dysfunction as a major contributor in psychiatric and neurodevelopmental disorders, such as cognitive impairment and autism, but the underlying cortical cellular pathogenesis remains unclear. This study is the first to reveal human neuronal pathogenesis resulting from abnormal Ras signaling and provides insights into how these phenotypic abnormalities likely contribute to neurodevelopmental disorders. We also demonstrate the synergy between mouse and human models, thereby validating the use of iPS cells as a platform to study underlying cellular pathologies resulting from signaling deficits. Recapitulating human cellular pathologies in vitro facilitates the future high throughput screening of potential therapeutic agents that may reverse phenotypic and behavioral deficits.
Asunto(s)
Síndrome de Costello/metabolismo , Síndrome de Costello/patología , Células Madre Pluripotentes Inducidas/metabolismo , Células-Madre Neurales/metabolismo , Células-Madre Neurales/patología , Proteínas ras/metabolismo , Adolescente , Adulto , Diferenciación Celular , Células Cultivadas , Niño , Preescolar , Femenino , Humanos , Células Madre Pluripotentes Inducidas/patología , Lactante , Masculino , Persona de Mediana Edad , Regulación hacia ArribaRESUMEN
Impaired cerebral glucose metabolism is a pathologic feature of Alzheimer Disease (AD), and recent proteomic studies highlight a disruption of glial carbohydrate metabolism with disease progression. Here, we report that inhibition of indoleamine-2,3-dioxygenase 1 (IDO1), which metabolizes tryptophan to kynurenine (KYN) in the first step of the kynurenine pathway, rescues hippocampal memory function and plasticity in preclinical models of amyloid and tau pathology by restoring astrocytic metabolic support of neurons. Activation of IDO1 in astrocytes by amyloid-beta 42 and tau oligomers, two major pathological effectors in AD, increases KYN and suppresses glycolysis in an AhR-dependent manner. Conversely, pharmacological IDO1 inhibition restores glycolysis and lactate production. In amyloid-producing APP Swe -PS1 ΔE9 and 5XFAD mice and in tau-producing P301S mice, IDO1 inhibition restores spatial memory and improves hippocampal glucose metabolism by metabolomic and MALDI-MS analyses. IDO1 blockade also rescues hippocampal long-term potentiation (LTP) in a monocarboxylate transporter (MCT)-dependent manner, suggesting that IDO1 activity disrupts astrocytic metabolic support of neurons. Indeed, in vitro mass-labeling of human astrocytes demonstrates that IDO1 regulates astrocyte generation of lactate that is then taken up by human neurons. In co-cultures of astrocytes and neurons derived from AD subjects, deficient astrocyte lactate transfer to neurons was corrected by IDO1 inhibition, resulting in improved neuronal glucose metabolism. Thus, IDO1 activity disrupts astrocytic metabolic support of neurons across both amyloid and tau pathologies and in a model of AD iPSC-derived neurons. These findings also suggest that IDO1 inhibitors developed for adjunctive therapy in cancer could be repurposed for treatment of amyloid- and tau-mediated neurodegenerative diseases.
RESUMEN
Impaired cerebral glucose metabolism is a pathologic feature of Alzheimer's disease (AD), with recent proteomic studies highlighting disrupted glial metabolism in AD. We report that inhibition of indoleamine-2,3-dioxygenase 1 (IDO1), which metabolizes tryptophan to kynurenine (KYN), rescues hippocampal memory function in mouse preclinical models of AD by restoring astrocyte metabolism. Activation of astrocytic IDO1 by amyloid ß and tau oligomers increases KYN and suppresses glycolysis in an aryl hydrocarbon receptor-dependent manner. In amyloid and tau models, IDO1 inhibition improves hippocampal glucose metabolism and rescues hippocampal long-term potentiation in a monocarboxylate transporter-dependent manner. In astrocytic and neuronal cocultures from AD subjects, IDO1 inhibition improved astrocytic production of lactate and uptake by neurons. Thus, IDO1 inhibitors presently developed for cancer might be repurposed for treatment of AD.
Asunto(s)
Enfermedad de Alzheimer , Péptidos beta-Amiloides , Astrocitos , Glucosa , Glucólisis , Hipocampo , Indolamina-Pirrol 2,3,-Dioxigenasa , Quinurenina , Potenciación a Largo Plazo , Neuronas , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/tratamiento farmacológico , Animales , Hipocampo/metabolismo , Glucosa/metabolismo , Ratones , Humanos , Astrocitos/metabolismo , Péptidos beta-Amiloides/metabolismo , Quinurenina/metabolismo , Neuronas/metabolismo , Indolamina-Pirrol 2,3,-Dioxigenasa/metabolismo , Cognición/efectos de los fármacos , Modelos Animales de Enfermedad , Proteínas tau/metabolismo , Transportadores de Ácidos Monocarboxílicos/metabolismo , Masculino , Receptores de Hidrocarburo de Aril/metabolismo , Ácido Láctico/metabolismo , Triptófano/metabolismo , Memoria/efectos de los fármacosRESUMEN
Many molecules regulate synaptogenesis, but intracellular signaling pathways required for their functions are poorly understood. Afadin is a Rap-regulated, actin-binding protein that promotes cadherin complex assembly as well as binding many other cell adhesion molecules and receptors. To examine its role in mediating synaptogenesis, we deleted afadin (mllt1), using a conditional allele, in postmitotic hippocampal neurons. Consistent with its role in promoting cadherin recruitment, afadin deletion resulted in 70% fewer and less intense N-cadherin puncta with similar reductions of ß-catenin and αN-catenin puncta densities and 35% reduction in EphB2 puncta density. Its absence also resulted in 40% decreases in spine and excitatory synapse densities in the stratum radiatum of CA1, as determined by morphology, apposition of presynaptic and postsynaptic markers, and synaptic transmission. The remaining synapses appeared to function normally. Thus, afadin is a key intracellular signaling molecule for cadherin recruitment and is necessary for spine and synapse formation in vivo.
Asunto(s)
Región CA1 Hipocampal/metabolismo , Cadherinas/fisiología , Espinas Dendríticas/metabolismo , Potenciales Postsinápticos Excitadores/fisiología , Proteínas de Microfilamentos/genética , Membranas Sinápticas/metabolismo , Animales , Región CA1 Hipocampal/crecimiento & desarrollo , Región CA1 Hipocampal/ultraestructura , Línea Celular , Espinas Dendríticas/ultraestructura , Femenino , Técnicas de Sustitución del Gen/métodos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Mutantes , Proteínas de Microfilamentos/deficiencia , Técnicas de Cultivo de Órganos , Membranas Sinápticas/ultraestructuraRESUMEN
Noncanonical microRNAs (miRNAs) and endogenous small interfering RNAs (endo-siRNAs) are distinct subclasses of small RNAs that bypass the DGCR8/DROSHA Microprocessor but still require DICER1 for their biogenesis. What role, if any, they have in mammals remains unknown. To identify potential functional properties for these subclasses, we compared the phenotypes resulting from conditional deletion of Dgcr8 versus Dicer1 in post-mitotic neurons. The loss of Dicer1 resulted in an earlier lethality, more severe structural abnormalities, and increased apoptosis relative to that from Dgcr8 loss. Deep sequencing of small RNAs from the hippocampus and cortex of the conditional knockouts and control littermates identified multiple noncanonical microRNAs that were expressed at high levels in the brain relative to other tissues, including mirtrons and H/ACA snoRNA-derived small RNAs. In contrast, we found no evidence for endo-siRNAs in the brain. Taken together, our findings provide evidence for a diverse population of highly expressed noncanonical miRNAs that together are likely to play important functional roles in post-mitotic neurons.
Asunto(s)
Encéfalo/metabolismo , ARN Helicasas DEAD-box/metabolismo , MicroARNs/genética , Proteínas/metabolismo , Ribonucleasa III/metabolismo , Animales , Encéfalo/citología , ARN Helicasas DEAD-box/deficiencia , Ratones , Ratones Noqueados , Mitosis , Fenotipo , Proteínas de Unión al ARN , Ribonucleasa III/deficiencia , Análisis de Secuencia de ARNRESUMEN
MicroRNAs (miRNAs) are critical regulators of nervous system function, and in vivo knockout studies have demonstrated that miRNAs are necessary for multiple aspects of neuronal development and survival. However, the role of miRNA biogenesis in the formation and function of synapses in the cerebral cortex is only minimally understood. Here, we have generated and characterized a mouse line with a conditional neuronal deletion of Dgcr8, a miRNA biogenesis protein predicted to process miRNAs exclusively. Loss of Dgcr8 in pyramidal neurons of the cortex results in a non-cell-autonomous reduction in parvalbumin interneurons in the prefrontal cortex, accompanied by a severe deficit in inhibitory synaptic transmission and a corresponding reduction of inhibitory synapses. Together, these results suggest a vital role for miRNAs in governing essential aspects of inhibitory transmission and interneuron development in the mammalian nervous system. These results may be relevant to human diseases such as schizophrenia, where both altered Dgcr8 levels as well as aberrant inhibitory transmission in the prefrontal cortex have been postulated to contribute to the pathophysiology of the disease.
Asunto(s)
Potenciales Postsinápticos Inhibidores/genética , MicroARNs/metabolismo , Corteza Prefrontal/fisiología , Proteínas/genética , Células Piramidales/fisiología , Animales , Encéfalo/anomalías , Tamaño de la Célula , Eliminación de Gen , Interneuronas/metabolismo , Ratones , Ratones Noqueados , MicroARNs/genética , Pilocarpina/farmacología , Corteza Prefrontal/citología , Corteza Prefrontal/metabolismo , Proteínas/metabolismo , Células Piramidales/metabolismo , Proteínas de Unión al ARN , Convulsiones/inducido químicamenteRESUMEN
The sheer complexity of the brain has complicated our ability to understand its cellular mechanisms in health and disease. Genome-wide association studies have uncovered genetic variants associated with specific neurological phenotypes and diseases. In addition, single-cell transcriptomics have provided molecular descriptions of specific brain cell types and the changes they undergo during disease. Although these approaches provide a giant leap forward towards understanding how genetic variation can lead to functional changes in the brain, they do not establish molecular mechanisms. To address this need, we developed a 3D co-culture system termed iAssembloids (induced multi-lineage assembloids) that enables the rapid generation of homogenous neuron-glia spheroids. We characterize these iAssembloids with immunohistochemistry and single-cell transcriptomics and combine them with large-scale CRISPRi-based screens. In our first application, we ask how glial and neuronal cells interact to control neuronal death and survival. Our CRISPRi-based screens identified that GSK3ß inhibits the protective NRF2-mediated oxidative stress response in the presence of reactive oxygen species elicited by high neuronal activity, which was not previously found in 2D monoculture neuron screens. We also apply the platform to investigate the role of APOE-ε4, a risk variant for Alzheimer's Disease, in its effect on neuronal survival. This platform expands the toolbox for the unbiased identification of mechanisms of cell-cell interactions in brain health and disease.
RESUMEN
Loss of function (LoF) of TAR-DNA binding protein 43 (TDP-43) and mis-localization, together with TDP-43-positive and hyperphosphorylated inclusions, are found in post-mortem tissue of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) patients, including those carrying LoF variants in the progranulin gene (GRN). Modeling TDP-43 pathology has been challenging in vivo and in vitro. We present a three-dimensional induced pluripotent stem cell (iPSC)-derived paradigm-mature brain organoids (mbOrg)-composed of cortical-like-astrocytes (iA) and neurons. When devoid of GRN, mbOrgs spontaneously recapitulate TDP-43 mis-localization, hyperphosphorylation, and LoF phenotypes. Mixing and matching genotypes in mbOrgs showed that GRN-/- iA are drivers for TDP-43 pathology. Finally, we rescued TDP-43 LoF by adding exogenous progranulin, demonstrating a link between TDP-43 LoF and progranulin expression. In conclusion, we present an iPSC-derived platform that shows striking features of human TDP-43 proteinopathy and provides a tool for the mechanistic modeling of TDP-43 pathology and patient-tailored therapeutic screening for FTD and ALS.
Asunto(s)
Esclerosis Amiotrófica Lateral , Demencia Frontotemporal , Humanos , Esclerosis Amiotrófica Lateral/patología , Demencia Frontotemporal/genética , Granulinas/genética , Granulinas/metabolismo , Progranulinas/genética , Progranulinas/metabolismo , Astrocitos/metabolismo , Mutación , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Encéfalo/metabolismoRESUMEN
Mutations in the human progranulin (GRN) gene are a leading cause of frontotemporal lobar degeneration (FTLD). While previous studies implicate aberrant microglial activation as a disease-driving factor in neurodegeneration in the thalamocortical circuit in Grn-/- mice, the exact mechanism for neurodegeneration in FTLD-GRN remains unclear. By performing comparative single-cell transcriptomics in the thalamus and frontal cortex of Grn-/- mice and patients with FTLD-GRN, we have uncovered a highly conserved astroglial pathology characterized by upregulation of gap junction protein GJA1, water channel AQP4, and lipid-binding protein APOE, and downregulation of glutamate transporter SLC1A2 that promoted profound synaptic degeneration across the two species. This astroglial toxicity could be recapitulated in mouse astrocyte-neuron cocultures and by transplanting induced pluripotent stem cell-derived astrocytes to cortical organoids, where progranulin-deficient astrocytes promoted synaptic degeneration, neuronal stress, and TDP-43 proteinopathy. Together, these results reveal a previously unappreciated astroglial pathology as a potential key mechanism in neurodegeneration in FTLD-GRN.
Asunto(s)
Demencia Frontotemporal , Degeneración Lobar Frontotemporal , Humanos , Animales , Ratones , Progranulinas/genética , Demencia Frontotemporal/genética , Astrocitos/metabolismo , Péptidos y Proteínas de Señalización Intercelular/genética , Mutación , Degeneración Lobar Frontotemporal/genética , Degeneración Lobar Frontotemporal/metabolismo , Degeneración Lobar Frontotemporal/patologíaRESUMEN
Using machine learning (ML), we interrogated the function of all human-chimpanzee variants in 2,645 human accelerated regions (HARs), finding 43% of HARs have variants with large opposing effects on chromatin state and 14% on neurodevelopmental enhancer activity. This pattern, consistent with compensatory evolution, was confirmed using massively parallel reporter assays in chimpanzee and human neural progenitor cells. The species-specific enhancer activity of HARs was accurately predicted from the presence and absence of transcription factor footprints in each species. Despite these striking cis effects, activity of a given HAR sequence was nearly identical in human and chimpanzee cells. This suggests that HARs did not evolve to compensate for changes in the trans environment but instead altered their ability to bind factors present in both species. Thus, ML prioritized variants with functional effects on human neurodevelopment and revealed an unexpected reason why HARs may have evolved so rapidly.