RÉSUMÉ
Introduction: Enteric glial cells are important players in the control of motility, intestinal barrier integrity and inflammation. During inflammation, they switch into a reactive phenotype enabling them to release inflammatory mediators, thereby shaping the inflammatory environment. While a plethora of well-established in vivo models exist, cell culture models necessary to decipher the mechanistic pathways of enteric glial reactivity are less well standardized. In particular, the composition of extracellular matrices (ECM) can massively affect the experimental outcome. Considering the growing number of studies involving primary enteric glial cells, a better understanding of their homeostatic and inflammatory in vitro culture conditions is needed. Methods: We examined the impact of different ECMs on enteric glial culture purity, network morphology and immune responsiveness. Therefore, we used immunofluorescence and brightfield microscopy, as well as 3' bulk mRNA sequencing. Additionally, we compared cultured cells with in vivo enteric glial transcriptomes isolated from Sox10iCreERT2Rpl22HA/+ mice. Results: We identified Matrigel and laminin as superior over other coatings, including poly-L-ornithine, different lysines, collagens, and fibronectin, gaining the highest enteric glial purity and most extended glial networks expressing connexin-43 hemichannels allowing intercellular communication. Transcriptional analysis revealed strong similarities between enteric glia on Matrigel and laminin with enrichment of gene sets supporting neuronal differentiation, while cells on poly-L-ornithine showed enrichment related to cell proliferation. Comparing cultured and in vivo enteric glial transcriptomes revealed a 50% overlap independent of the used coating substrates. Inflammatory activation of enteric glia by IL-1ß treatment showed distinct coating-dependent gene expression signatures, with an enrichment of genes related to myeloid and epithelial cell differentiation on Matrigel and laminin coatings, while poly-L-ornithine induced more gene sets related to lymphocyte differentiation. Discussion: Together, changes in morphology, differentiation and immune activation of primary enteric glial cells proved a strong effect of the ECM. We identified Matrigel and laminin as pre-eminent substrates for murine enteric glial cultures. These new insights will help to standardize and improve enteric glial culture quality and reproducibility between in vitro studies in the future, allowing a better comparison of their functional role in enteric neuroinflammation.
Sujet(s)
Matrice extracellulaire , Homéostasie , Laminine , Névroglie , Animaux , Matrice extracellulaire/métabolisme , Névroglie/métabolisme , Névroglie/immunologie , Souris , Laminine/métabolisme , Système nerveux entérique/métabolisme , Système nerveux entérique/immunologie , Cellules cultivées , Association médicamenteuse , Collagène/métabolisme , Souris de lignée C57BL , Protéoglycanes/métabolismeRÉSUMÉ
BACKGROUND: To explore whether nobiletin has a protective effect on high-fat diet (HFD)-induced enteric nerve injury and its underlying mechanism. METHODS: An obesity model was induced by a HFD. Nobiletin (100 mg/kg and 200 mg/kg) and vehicle were administered by gastric gavage for 4 weeks. Lee's index, body weight, OGTT and intestinal propulsion assays were performed before sacrifice. After sampling, lipids were detected using Bodipy 493/503; lipid peroxidation was detected using MDA and SOD kits and the expression of PGP 9.5, Trem2, GFAP, ß-tubulin 3, Bax, Bcl2, Nestin, P75 NTR, SOX10 and EDU was detected using immunofluorescence. The GDNF, p-AKT, AKT, p-FOXO3a, FOXO3a and P21 proteins were detected using western blotting. The relative mRNA expression levels of NOS2 were detected via qPCR. Primary enteric neural stem cells (ENSCs) were cultured. After ENSCs were treated with palmitic acid (PA) and nobiletin, CCK-8 and caspase-3/7 activity assays were performed to evaluate proliferation and apoptosis. RESULTS: HFD consumption caused colon lipid accumulation and peroxidation, induced enteric nerve damage and caused intestinal motor dysfunction. However, nobiletin reduced lipid accumulation and peroxidation in the colon; promoted Trem2, ß-tubulin 3, Nestin, P75NTR, SOX10 and Bcl2 expression; inhibited Bax and GFAP expression; reduced NOS2 mRNA transcription; and regulated the GDNF/AKT/FOXO3a/P21 pathway. Nobiletin also promoted PA-induced impairment of ENSCs. CONCLUSIONS: Nobiletin restored HFD-induced enteric nerve injury, which may be associated with inhibiting enteric nerve apoptosis, promoting enteric nerve survival and regulating the GDNF/AKT/FOXO3a/P21 pathway.
Sujet(s)
Alimentation riche en graisse , Système nerveux entérique , Flavones , Protéine O3 à motif en tête de fourche , Facteur neurotrophique dérivé des cellules gliales , Protéines proto-oncogènes c-akt , Transduction du signal , Animaux , Protéine O3 à motif en tête de fourche/métabolisme , Facteur neurotrophique dérivé des cellules gliales/métabolisme , Protéines proto-oncogènes c-akt/métabolisme , Alimentation riche en graisse/effets indésirables , Transduction du signal/effets des médicaments et des substances chimiques , Mâle , Flavones/pharmacologie , Flavones/usage thérapeutique , Système nerveux entérique/métabolisme , Système nerveux entérique/effets des médicaments et des substances chimiques , Névroglie/métabolisme , Névroglie/effets des médicaments et des substances chimiques , Souris , Modèles animaux de maladie humaine , Rats , Obésité/métabolisme , Obésité/traitement médicamenteux , Apoptose/effets des médicaments et des substances chimiquesRÉSUMÉ
Due to their extensive use, the release of zinc oxide nanoparticles (ZnO NP) into the environment is increasing and may lead to unintended risk to both human health and ecosystems. Access of ZnO NP to the brain has been demonstrated, so their potential toxicity on the nervous system is a matter of particular concern. Although evaluation of ZnO NP toxicity has been reported in several previous studies, the specific effects on the nervous system are not completely understood and, particularly, effects on genetic material and on organism behaviour are poorly addressed. We evaluated the potential toxic effects of ZnO NP in vitro and in vivo, and the role of zinc ions (Zn2+) in these effects. In vitro, the ability of ZnO NP to be internalized by A172 glial cells was verified, and the cytotoxic and genotoxic effects of ZnO NP or the released Zn2+ ions were addressed by means of vital dye exclusion and comet assay, respectively. In vivo, behavioural alterations were evaluated in zebrafish embryos using a total locomotion assay. ZnO NP induced decreases in viability of A172 cells after 24 h of exposure and genetic damage after 3 and 24 h. The involvement of the Zn2+ ions released from the NP in genotoxicity was confirmed. ZnO NP exposure also resulted in decreased locomotor activity of zebrafish embryos, with a clear role of released Zn2+ ions in this effect. These findings support the toxic potential of ZnO NP showing, for the first time, genetic effects on glial cells and proving the intervention of Zn2+ ions.
Sujet(s)
Danio zébré , Oxyde de zinc , Oxyde de zinc/toxicité , Animaux , Humains , Nanoparticules métalliques/toxicité , Altération de l'ADN , Survie cellulaire/effets des médicaments et des substances chimiques , Comportement animal/effets des médicaments et des substances chimiques , Test des comètes , Névroglie/effets des médicaments et des substances chimiques , Nanoparticules/toxicitéRÉSUMÉ
Alzheimer's disease (AD) is the leading cause of dementia in older adults, and the need for effective, sustainable therapeutic targets is imperative. The complement pathway has been proposed as a therapeutic target. C5aR1 inhibition reduces plaque load, gliosis, and memory deficits in animal models, however, the cellular bases underlying this neuroprotection were unclear. Here, we show that the C5aR1 antagonist PMX205 improves outcomes in the Arctic48 mouse model of AD. A combination of single cell and single nucleus RNA-seq analysis of hippocampi derived from males and females identified neurotoxic disease-associated microglia clusters in Arctic mice that are C5aR1-dependent, while microglial genes associated with synapse organization and transmission and learning were overrepresented in PMX205-treated mice. PMX205 also reduced neurotoxic astrocyte gene expression, but clusters associated with protective responses to injury were unchanged. C5aR1 inhibition promoted mRNA-predicted signaling pathways between brain cell types associated with cell growth and repair, while suppressing inflammatory pathways. Finally, although hippocampal plaque load was unaffected, PMX205 prevented deficits in short-term memory in female Arctic mice. In conclusion, C5aR1 inhibition prevents cognitive loss, limits detrimental glial polarization while permitting neuroprotective responses, as well as leaving most protective functions of complement intact, making C5aR1 antagonism an attractive therapeutic strategy for AD.
Sujet(s)
Maladie d'Alzheimer , Modèles animaux de maladie humaine , Hippocampe , Microglie , Récepteur à l'anaphylatoxine C5a , Transduction du signal , Animaux , Récepteur à l'anaphylatoxine C5a/antagonistes et inhibiteurs , Récepteur à l'anaphylatoxine C5a/métabolisme , Récepteur à l'anaphylatoxine C5a/génétique , Maladie d'Alzheimer/métabolisme , Maladie d'Alzheimer/traitement médicamenteux , Maladie d'Alzheimer/anatomopathologie , Maladie d'Alzheimer/génétique , Souris , Femelle , Mâle , Transduction du signal/effets des médicaments et des substances chimiques , Microglie/effets des médicaments et des substances chimiques , Microglie/métabolisme , Hippocampe/métabolisme , Hippocampe/anatomopathologie , Hippocampe/effets des médicaments et des substances chimiques , Névroglie/effets des médicaments et des substances chimiques , Névroglie/métabolisme , Astrocytes/effets des médicaments et des substances chimiques , Astrocytes/métabolisme , Souris transgéniques , Humains , Souris de lignée C57BLRÉSUMÉ
Mosaic Analysis with Double Markers (MADM) is a powerful genetic method typically used for lineage tracing and to disentangle cell autonomous and tissue-wide roles of candidate genes with single cell resolution. Given the relatively sparse labeling, depending on which of the 19 MADM chromosomes one chooses, the MADM approach represents the perfect opportunity for cell morphology analysis. Various MADM studies include reports of morphological anomalies and phenotypes in the central nervous system (CNS). MADM for any candidate gene can easily incorporate morphological analysis within the experimental workflow. Here, we describe the methods of morphological cell analysis which we developed in the course of diverse recent MADM studies. This chapter will specifically focus on methods to quantify aspects of the morphology of neurons and astrocytes within the CNS, but these methods can broadly be applied to any MADM-labeled cells throughout the entire organism. We will cover two analyses-soma volume and dendrite characterization-of physical characteristics of pyramidal neurons in the somatosensory cortex, and two analyses-volume and Sholl analysis-of astrocyte morphology.
Sujet(s)
Astrocytes , Névroglie , Neurones , Animaux , Neurones/cytologie , Neurones/métabolisme , Astrocytes/cytologie , Astrocytes/métabolisme , Névroglie/cytologie , Névroglie/métabolisme , Souris , Mosaïcisme , Marqueurs biologiques , Dendrites/métabolisme , Cortex somatosensoriel/cytologieRÉSUMÉ
Chondroitin sulfate proteoglycans (CSPGs) are fundamental components of the extracellular matrix in the central nervous system (CNS). Among these, the Nerve-Glial antigen 2 (NG2) stands out as a transmembrane CSPG exclusively expressed in a different population of cells collectively termed NG2-expressing cells. These enigmatic cells, found throughout the developing and adult CNS, have been indicated with various names, including NG2 progenitor cells, polydendrocytes, synantocytes, NG2 cells, and NG2-Glia, but are more commonly referred to as oligodendrocyte progenitor cells. Characterized by high proliferation rates and unique morphology, NG2-expressing cells stand apart from neurons, astrocytes, and oligodendrocytes. Intriguingly, some NG2-expressing cells form functional glutamatergic synapses with neurons, challenging the long-held belief that only neurons possess the intricate machinery required for neurotransmission. In the CNS, the complexity surrounding NG2-expressing cells extends to their classification. Additionally, NG2 expression has been documented in pericytes and immune cells, suggesting a role in regulating brain innate immunity and neuro-immune crosstalk in homeostasis. Ongoing debates revolve around their heterogeneity, potential as progenitors for various cell types, responses to neuroinflammation, and the role of NG2. Therefore, this review aims to shed light on the enigma of NG2-expressing cells by delving into their structure, functions, and signaling pathways. We will critically evaluate the literature on NG2 expression across the CNS, and address the contentious issues surrounding their classification and roles in neuroinflammation and neurodegeneration. By unraveling the intricacies of NG2-expressing cells, we hope to pave the way for a more comprehensive understanding of their contributions to CNS health and during neurological disorders.
Sujet(s)
Antigènes , Système nerveux central , Humains , Animaux , Système nerveux central/immunologie , Système nerveux central/métabolisme , Antigènes/immunologie , Antigènes/métabolisme , Névroglie/métabolisme , Névroglie/immunologie , Névroglie/physiologie , Neurones/métabolisme , Protéoglycanes à chondroïtine sulfate/métabolisme , ProtéoglycanesRÉSUMÉ
BACKGROUND: Satellite glial cells (SGCs) in the dorsal root ganglia (DRG) play a pivotal role in the formation of neuropathic pain (NP). Sciatic nerve stimulation (SNS) neuromodulation was reported to alleviate NP and reduce neuroinflammation. However, the mechanisms underlying SNS in the DRG remain unclear. This study aimed to elucidate the mechanism of electric stimulation in reducing NP, focusing on the DRG. METHODS: L5 nerve root ligation (NRL) NP rat model was studied. Ipsilateral SNS performed 1 day after NRL. Behavioral tests were performed to assess pain phenotypes. NanoString Ncounter technology was used to explore the differentially expressed genes and cellular pathways. Activated SGCs were characterized in vivo and in vitro. The histochemical alterations of SGCs, macrophages, and neurons in DRG were examined in vivo on post-injury day 8. RESULTS: NRL induced NP behaviors including decreased pain threshold and latency on von Frey and Hargreaves tests. We found that following nerve injury, SGCs were hyperactivated, neurotoxic and had increased expression of NP-related ion channels including TRPA1, Cx43, and SGC-neuron gap junctions. Mechanistically, nerve injury induced reciprocal activation of SGCs and M1 macrophages via cytokines including IL-6, CCL3, and TNF-α mediated by the HIF-1α-NF-κB pathways. SNS suppressed SGC hyperactivation, reduced the expression of NP-related ion channels, and induced M2 macrophage polarization, thereby alleviating NP and associated neuroinflammation in the DRG. CONCLUSIONS: NRL induced hyperactivation of SGCs, which had increased expression of NP-related ion channels. Reciprocal activation of SGCs and M1 macrophages surrounding the primary sensory neurons was mediated by the HIF-1α and NF-κB pathways. SNS suppressed SGC hyperactivation and skewed M1 macrophage towards M2. Our findings establish SGC activation as a crucial pathomechanism in the gliopathic alterations in NP, which can be modulated by SNS neuromodulation.
Sujet(s)
Modèles animaux de maladie humaine , Ganglions sensitifs des nerfs spinaux , Névralgie , Maladies neuro-inflammatoires , Rat Sprague-Dawley , Nerf ischiatique , Animaux , Ganglions sensitifs des nerfs spinaux/métabolisme , Névralgie/thérapie , Névralgie/métabolisme , Mâle , Maladies neuro-inflammatoires/métabolisme , Nerf ischiatique/anatomopathologie , Macrophages/métabolisme , Névroglie/métabolisme , Rats , Comportement animalRÉSUMÉ
The sympathetic nervous system controls bodily functions including vascular tone, cardiac rhythm, and the "fight-or-flight response". Sympathetic chain ganglia develop in parallel with preganglionic motor nerves extending from the neural tube, raising the question of whether axon targeting contributes to sympathetic chain formation. Using nerve-selective genetic ablations and lineage tracing in mouse, we reveal that motor nerve-associated Schwann cell precursors (SCPs) contribute sympathetic neurons and satellite glia after the initial seeding of sympathetic ganglia by neural crest. Motor nerve ablation causes mispositioning of SCP-derived sympathoblasts as well as sympathetic chain hypoplasia and fragmentation. Sympathetic neurons in motor-ablated embryos project precociously and abnormally towards dorsal root ganglia, eventually resulting in fusion of sympathetic and sensory ganglia. Cell interaction analysis identifies semaphorins as potential motor nerve-derived signaling molecules regulating sympathoblast positioning and outgrowth. Overall, central innervation functions both as infrastructure and regulatory niche to ensure the integrity of peripheral ganglia morphogenesis.
Sujet(s)
Ganglions sympathiques , Motoneurones , Crête neurale , Cellules de Schwann , Système nerveux sympathique , Animaux , Système nerveux sympathique/embryologie , Souris , Motoneurones/physiologie , Cellules de Schwann/métabolisme , Crête neurale/cytologie , Crête neurale/métabolisme , Ganglions sympathiques/cytologie , Ganglions sensitifs des nerfs spinaux , Sémaphorines/métabolisme , Sémaphorines/génétique , Souris transgéniques , Névroglie/métabolisme , FemelleRÉSUMÉ
Glial scar formation represents a fundamental response to central nervous system (CNS) injuries. It is mainly characterized by a well-defined spatial rearrangement of reactive astrocytes and microglia. The mechanisms underlying glial scar formation have been extensively studied, yet quantitative descriptors of the spatial arrangement of reactive glial cells remain limited. Here, we present a novel approach using point pattern analysis (PPA) and topological data analysis (TDA) to quantify spatial patterns of reactive glial cells after experimental ischemic stroke in mice. We provide open and reproducible tools using R and Julia to quantify spatial intensity, cell covariance and conditional distribution, cell-to-cell interactions, and short/long-scale arrangement, which collectively disentangle the arrangement patterns of the glial scar. This approach unravels a substantial divergence in the distribution of GFAP+ and IBA1+ cells after injury that conventional analysis methods cannot fully characterize. PPA and TDA are valuable tools for studying the complex spatial arrangement of reactive glia and other nervous cells following CNS injuries and have potential applications for evaluating glial-targeted restorative therapies.
Sujet(s)
Astrocytes , Cicatrice , Névroglie , Animaux , Souris , Cicatrice/anatomopathologie , Névroglie/anatomopathologie , Astrocytes/anatomopathologie , Microglie/anatomopathologie , Accident vasculaire cérébral ischémique/anatomopathologie , Analyse de données , Modèles animaux de maladie humaine , Mâle , Protéine gliofibrillaire acide/métabolisme , Souris de lignée C57BLRÉSUMÉ
Amyloid-peptide (Aß) monomeric forms (ABM) occurring in presymptomatic Alzheimer's disease (AD) brain are thought to be devoid of neurotoxicity while the transition/aggregation of ABM into oligomers is determinant for Aß-induced toxicity since Aß is predominantly monomeric up to 3 µM and aggregates over this concentration. However, recent imaging and/or histopathological investigations revealed alterations of myelin in prodromal AD brain in absence of aggregated Aß oligomers, suggesting that ABM may induce toxicity in myelin-producing cells in early AD-stages. To check this hypothesis, here we studied ABM effects on the viability of the Human oligodendrocyte cell line (HOG), a reliable oligodendrocyte model producing myelin proteins. Furthermore, to mimic closely interactions between oligodendrocytes and other glial cells regulating myelination, we investigated also ABM effects on mouse brain primary mixed-glial cell cultures. Various methods were combined to show that ABM concentrations (600 nM-1 µM), extremely lower than 3 µM, significantly decreased HOG cell and mouse brain primary mixed-glial cell survival. Interestingly, flow-cytometry studies using specific cell-type markers demonstrated that oligodendrocytes represent the most vulnerable glial cell population affected by ABM toxicity. Our work also shows that the neurosteroid 3α-O-allyl-allopregnanolone BR351 (250 and 500 nM) efficiently prevented ABM-induced HOG and brain primary glial cell toxicity. Bicuculline (50-100 nM), the GABA-A-receptor antagonist, was unable to block/reduce BR351 effect against ABM-induced HOG and primary glial cell toxicity, suggesting that BR351-evoked neuroprotection of these cells may not depend on GABA-A-receptor allosterically modulated by neurosteroids. Altogether, our results suggest that further exploration of BR351 therapeutic potential may offer interesting perspectives to develop effective neuroprotective strategies.
Sujet(s)
Peptides bêta-amyloïdes , Neuroprotecteurs , Oligodendroglie , Prégnanolone , Animaux , Oligodendroglie/effets des médicaments et des substances chimiques , Oligodendroglie/métabolisme , Humains , Peptides bêta-amyloïdes/toxicité , Neuroprotecteurs/pharmacologie , Prégnanolone/pharmacologie , Souris , Lignée cellulaire , Encéphale/effets des médicaments et des substances chimiques , Encéphale/métabolisme , Survie cellulaire/effets des médicaments et des substances chimiques , Survie cellulaire/physiologie , Névroglie/effets des médicaments et des substances chimiques , Névroglie/métabolisme , Souris de lignée C57BL , Fragments peptidiques/toxicité , Cellules cultivées , Relation dose-effet des médicamentsRÉSUMÉ
The role of central nervous system (CNS) glia in sustaining self-autonomous inflammation and driving clinical progression in multiple sclerosis (MS) is gaining scientific interest. We applied a single transcription factor (SOX10)-based protocol to accelerate oligodendrocyte differentiation from human induced pluripotent stem cell (hiPSC)-derived neural precursor cells, generating self-organizing forebrain organoids. These organoids include neurons, astrocytes, oligodendroglia, and hiPSC-derived microglia to achieve immunocompetence. Over 8 weeks, organoids reproducibly generated mature CNS cell types, exhibiting single-cell transcriptional profiles similar to the adult human brain. Exposed to inflamed cerebrospinal fluid (CSF) from patients with MS, organoids properly mimic macroglia-microglia neurodegenerative phenotypes and intercellular communication seen in chronic active MS. Oligodendrocyte vulnerability emerged by day 6 post-MS-CSF exposure, with nearly 50% reduction. Temporally resolved organoid data support and expand on the role of soluble CSF mediators in sustaining downstream events leading to oligodendrocyte death and inflammatory neurodegeneration. Such findings support the implementation of this organoid model for drug screening to halt inflammatory neurodegeneration.
Sujet(s)
Encéphale , Différenciation cellulaire , Cellules souches pluripotentes induites , Sclérose en plaques , Névroglie , Organoïdes , Phénotype , Humains , Sclérose en plaques/anatomopathologie , Sclérose en plaques/métabolisme , Cellules souches pluripotentes induites/métabolisme , Cellules souches pluripotentes induites/anatomopathologie , Organoïdes/anatomopathologie , Organoïdes/métabolisme , Névroglie/métabolisme , Névroglie/anatomopathologie , Encéphale/anatomopathologie , Encéphale/métabolisme , Oligodendroglie/métabolisme , Oligodendroglie/anatomopathologie , Cellules souches neurales/métabolisme , Cellules souches neurales/anatomopathologie , Microglie/métabolisme , Microglie/anatomopathologieRÉSUMÉ
Viral infection is frequently assayed by ongoing expression of viral genes. These assays fail to identify cells that have been exposed to the virus but limit or inhibit viral replication. To address this limitation, we used a dual-labeling vesicular stomatitis virus (DL-VSV), which has a deletion of the viral glycoprotein gene, to allow evaluation of primary infection outcomes. This virus encodes Cre, which can stably mark any cell with even a minimal level of viral gene expression. Additionally, the virus encodes GFP, which distinguishes cells with higher levels of viral gene expression, typically due to genome replication. Stereotactic injections of DL-VSV into the murine brain showed that different cell types had very different responses to the virus. Almost all neurons hosted high levels of viral gene expression, while glial cells varied in their responses. Astrocytes (Sox9+) were predominantly productively infected, while oligodendrocytes (Sox10+) were largely abortively infected. Microglial cells (Iba1+) were primarily uninfected. Furthermore, we monitored the early innate immune response to viral infection and identified unique patterns of interferon (IFN) induction. Shortly after infection, microglia were the main producers of IFNb, whereas later, oligodendrocytes were the main producers. IFNb+ cells were primarily abortively infected regardless of cell type. Last, we investigated whether IFN signaling had any impact on the outcome of primary infection and did not observe significant changes, suggesting that intrinsic factors are likely responsible for determining the outcome of primary infection.
Sujet(s)
Astrocytes , Animaux , Souris , Astrocytes/virologie , Astrocytes/métabolisme , Réplication virale , Microglie/virologie , Microglie/métabolisme , Microglie/immunologie , Neurones/virologie , Neurones/métabolisme , Facteur de transcription SOX-9/métabolisme , Facteur de transcription SOX-9/génétique , Vesiculovirus/physiologie , Vesiculovirus/immunologie , Vesiculovirus/génétique , Oligodendroglie/virologie , Oligodendroglie/métabolisme , Stomatite vésiculeuse/virologie , Stomatite vésiculeuse/immunologie , Immunité innée , Souris de lignée C57BL , Encéphale/virologie , Encéphale/métabolisme , Encéphale/immunologie , Névroglie/virologie , Névroglie/métabolismeRÉSUMÉ
Powassan virus (POWV) is an emergent tick-borne flavivirus that causes fatal encephalitis in the elderly and long-term neurologic sequelae in survivors. How age contributes to severe POWV encephalitis remains an enigma, and no animal models have assessed age-dependent POWV neuropathology. Inoculating C57BL/6 mice with a POWV strain (LI9) currently circulating in Ixodes ticks resulted in age-dependent POWV lethality 10-20 dpi. POWV infection of 50-week-old mice was 82% fatal with lethality sequentially reduced by age to 7.1% in 10-week-old mice. POWV LI9 was neuroinvasive in mice of all ages, causing acute spongiform CNS pathology and reactive gliosis 5-15 dpi that persisted in survivors 30 dpi. High CNS viral loads were found in all mice 10 dpi. However, by 15 dpi, viral loads decreased by 2-4 logs in 10- to 40-week-old mice, while remaining at high levels in 50-week-old mice. Age-dependent differences in CNS viral loads 15 dpi occurred concomitantly with striking changes in CNS cytokine responses. In the CNS of 50-week-old mice, POWV induced Th1-type cytokines (IFNγ, IL-2, IL-12, IL-4, TNFα, IL-6), suggesting a neurodegenerative pro-inflammatory M1 microglial program. By contrast, in 10-week-old mice, POWV-induced Th2-type cytokines (IL-10, TGFß, IL-4) were consistent with a neuroprotective M2 microglial phenotype. These findings correlate age-dependent CNS cytokine responses and viral loads with POWV lethality and suggest potential neuroinflammatory therapeutic targets. Our results establish the age-dependent lethality of POWV in a murine model that mirrors human POWV severity and long-term CNS pathology in the elderly. IMPORTANCE: Powassan virus is an emerging tick-borne flavivirus causing lethal encephalitis in aged individuals. We reveal an age-dependent POWV murine model that mirrors human POWV encephalitis and long-term CNS damage in the elderly. We found that POWV is neuroinvasive and directs reactive gliosis in all age mice, but at acute stages selectively induces pro-inflammatory Th1 cytokine responses in 50-week-old mice and neuroprotective Th2 cytokine responses in 10-week-old mice. Our findings associate CNS viral loads and divergent cytokine responses with age-dependent POWV lethality and survival outcomes. Responses of young mice suggest potential therapeutic targets and approaches for preventing severe POWV encephalitis that may be broadly applicable to other neurodegenerative diseases. Our age-dependent murine POWV model permits analysis of vaccines that prevent POWV lethality, and therapeutics that resolve severe POWV encephalitis.
Sujet(s)
Cytokines , Modèles animaux de maladie humaine , Virus de l'encéphalite à tiques (sous-groupe) , Encéphalites à tiques , Souris de lignée C57BL , Névroglie , Charge virale , Animaux , Souris , Virus de l'encéphalite à tiques (sous-groupe)/immunologie , Encéphalites à tiques/immunologie , Encéphalites à tiques/virologie , Encéphalites à tiques/mortalité , Encéphalites à tiques/anatomopathologie , Cytokines/métabolisme , Cytokines/immunologie , Névroglie/virologie , Névroglie/immunologie , Névroglie/anatomopathologie , Femelle , Facteurs âges , Ixodes/virologie , Ixodes/immunologie , Système nerveux central/virologie , Système nerveux central/immunologie , Système nerveux central/anatomopathologie , Encéphale/virologie , Encéphale/anatomopathologie , Encéphale/immunologieRÉSUMÉ
The central and peripheral nervous systems (CNS and PNS, respectively) exhibit remarkable diversity in the capacity to regenerate following neuronal injury with PNS injuries being much more likely to regenerate than those that occur in the CNS. Glial responses to damage greatly influence the likelihood of regeneration by either promoting or inhibiting axonal regrowth over time. However, despite our understanding of how some glial lineages participate in nerve degeneration and regeneration, less is known about the contributions of peripheral satellite glial cells (SGC) to regeneration failure following central axon branch injury of dorsal root ganglia (DRG) sensory neurons. Here, using in vivo, time-lapse imaging in larval zebrafish coupled with laser axotomy, we investigate the role of SGCs in axonal regeneration. In our studies we show that SGCs respond to injury by relocating their nuclei to the injury site during the same period that DRG neurons produce new central branch neurites. Laser ablation of SGCs prior to axon injury results in more neurite growth attempts and ultimately a higher rate of successful central axon regrowth, implicating SGCs as inhibitors of regeneration. We also demonstrate that this SGC response is mediated in part by ErbB signaling, as chemical inhibition of this receptor results in reduced SGC motility and enhanced central axon regrowth. These findings provide new insights into SGC-neuron interactions under injury conditions and how these interactions influence nervous system repair.
Sujet(s)
Axotomie , Ganglions sensitifs des nerfs spinaux , Régénération nerveuse , Danio zébré , Animaux , Régénération nerveuse/physiologie , Animal génétiquement modifié , Moelle spinale , Cellules satellites périneuronales/physiologie , Névroglie/physiologie , Protéines de poisson-zèbre/métabolisme , Axones/physiologieRÉSUMÉ
A recent approach to promote central nervous system (CNS) regeneration after injury or disease is direct conversion of somatic cells to neurons. This is achieved by transduction of viral vectors that express neurogenic transcription factors. In this work we propose adult human mucosal olfactory ensheathing glia (hmOEG) as a candidate for direct reprogramming to neurons due to its accessibility and to its well-characterized neuroregenerative capacity. After induction of hmOEG with the single neurogenic transcription factor NEUROD1, the cells under study exhibited morphological and immunolabeling neuronal features, fired action potentials and expressed glutamatergic and GABAergic markers. In addition, after engraftment of transduced hmOEG cells in the mouse hippocampus, these cells showed specific neuronal labeling. Thereby, if we add to the neuroregenerative capacity of hmOEG cultures the conversion to neurons of a fraction of their population through reprogramming techniques, the engraftment of hmOEG and hmOEG-induced neurons could be a procedure to enhance neural repair after central nervous system injury.
Sujet(s)
Névroglie , Neurones , Humains , Animaux , Névroglie/métabolisme , Névroglie/cytologie , Neurones/métabolisme , Neurones/cytologie , Souris , Adulte , Muqueuse olfactive/cytologie , Muqueuse olfactive/métabolisme , Facteurs de transcription à motif basique hélice-boucle-hélice/métabolisme , Facteurs de transcription à motif basique hélice-boucle-hélice/génétique , Lignage cellulaire , Hippocampe/cytologie , Hippocampe/métabolisme , Bulbe olfactif/cytologie , Bulbe olfactif/métabolisme , Cellules cultivéesRÉSUMÉ
During central nervous system (CNS) development, neural progenitor cells (NPCs) generate neurons and glia in two different ways. In direct neurogenesis, daughter cells differentiate directly into neurons or glia, whereas in indirect neurogenesis, neurons or glia are generated after one or more daughter cell divisions. Intriguingly, indirect neurogenesis is not stochastically deployed and plays instructive roles during CNS development: increased generation of cells from specific lineages; increased generation of early or late-born cell types within a lineage; and increased cell diversification. Increased indirect neurogenesis might contribute to the anterior CNS expansion evident throughout the Bilateria and help to modify brain-region size without requiring increased NPC numbers or extended neurogenesis. Increased indirect neurogenesis could be an evolutionary driver of the gyrencephalic (that is, folded) cortex that emerged during mammalian evolution and might even have increased during hominid evolution. Thus, selection of indirect versus direct neurogenesis provides a powerful developmental and evolutionary instrument that drives not only the evolution of CNS complexity but also brain expansion and modulation of brain-region size, and thereby the evolution of increasingly advanced cognitive abilities. This Review describes indirect neurogenesis in several model species and humans, and highlights some of the molecular genetic mechanisms that control this important process.
Sujet(s)
Neurogenèse , Neurogenèse/physiologie , Humains , Animaux , Évolution biologique , Cellules souches neurales/physiologie , Cellules souches neurales/cytologie , Neurones/physiologie , Différenciation cellulaire/physiologie , Système nerveux central/physiologie , Système nerveux central/croissance et développement , Système nerveux central/cytologie , Névroglie/physiologie , Encéphale/physiologie , Encéphale/croissance et développement , Encéphale/cytologieRÉSUMÉ
The polarization of cells often involves the transport of specific mRNAs and their localized translation in distal projections. Neurons and glia are both known to contain long cytoplasmic processes, while localized transcripts have only been studied extensively in neurons, not glia, especially in intact nervous systems. Here, we predict 1,740 localized Drosophila glial transcripts by extrapolating from our meta-analysis of seven existing studies characterizing the localized transcriptomes and translatomes of synaptically associated mammalian glia. We demonstrate that the localization of mRNAs in mammalian glial projections strongly predicts the localization of their high-confidence Drosophila homologs in larval motor neuron-associated glial projections and are highly statistically enriched for genes associated with neurological diseases. We further show that some of these localized glial transcripts are specifically required in glia for structural plasticity at the nearby neuromuscular junction synapses. We conclude that peripheral glial mRNA localization is a common and conserved phenomenon and propose that it is likely to be functionally important in disease.