RESUMO
BACKGROUND AND PURPOSE: While classic brain MR imaging features of Alexander disease have been well-documented, lesional patterns can overlap with other leukodystrophies, especially in the early stages of the disease or in milder phenotypes. We aimed to assess the utility of a new neuroimaging sign to help increase the diagnostic specificity of Alexander disease. MATERIALS AND METHODS: A peculiar bilateral symmetric hyperintense signal on T2-weighted images affecting the medulla oblongata was identified in an index patient with type I Alexander disease. Subsequently, 5 observers performed a systematic MR imaging review for this pattern by examining 55 subjects with Alexander disease and 74 subjects with other leukodystrophies. Interobserver agreement was assessed by the κ index. Sensitivity, specificity, and receiver operating characteristic curves were determined. RESULTS: The identified pattern was present in 87% of subjects with Alexander disease and 14% of those without Alexander disease leukodystrophy (P < .001), 3 with vanishing white matter, 4 with adult polyglucosan body disease, and 3 others. It was found equally in both type I and type II Alexander disease (28/32, 88% versus 18/21, 86%; P = .851) and in subjects with unusual disease features (2/2). Sensitivity (87.3%; 95% CI, 76.0%-93.7%), specificity (86.5%; 95% CI, 76.9%-92.5%), and interobserver agreement (κ index = 0.82) were high. CONCLUSIONS: The identified pattern in the medulla oblongata, called the chipmunk sign due to its resemblance to the face of this rodent, is extremely common in subjects with Alexander disease and represents a diagnostic tool that can aid in early diagnosis, especially in subjects with otherwise atypical MR imaging findings and/or clinical features.
Assuntos
Doença de Alexander , Imageamento por Ressonância Magnética , Sensibilidade e Especificidade , Humanos , Doença de Alexander/diagnóstico por imagem , Masculino , Feminino , Adulto , Imageamento por Ressonância Magnética/métodos , Pessoa de Meia-Idade , Adulto Jovem , Adolescente , Tronco Encefálico/diagnóstico por imagem , Tronco Encefálico/patologia , Criança , Idoso , Bulbo/diagnóstico por imagem , Bulbo/patologia , Pré-EscolarRESUMO
With concurrent global epidemics of chronic pain and opioid use disorders, there is a critical need to identify, target and manipulate specific cell populations expressing the mu-opioid receptor (MOR). However, available tools and transgenic models for gaining long-term genetic access to MOR+ neural cell types and circuits involved in modulating pain, analgesia and addiction across species are limited. To address this, we developed a catalog of MOR promoter (MORp) based constructs packaged into adeno-associated viral vectors that drive transgene expression in MOR+ cells. MORp constructs designed from promoter regions upstream of the mouse Oprm1 gene (mMORp) were validated for transduction efficiency and selectivity in endogenous MOR+ neurons in the brain, spinal cord, and periphery of mice, with additional studies revealing robust expression in rats, shrews, and human induced pluripotent stem cell (iPSC)-derived nociceptors. The use of mMORp for in vivo fiber photometry, behavioral chemogenetics, and intersectional genetic strategies is also demonstrated. Lastly, a human designed MORp (hMORp) efficiently transduced macaque cortical OPRM1+ cells. Together, our MORp toolkit provides researchers cell type specific genetic access to target and functionally manipulate mu-opioidergic neurons across a range of vertebrate species and translational models for pain, addiction, and neuropsychiatric disorders.
Assuntos
Analgesia , Dor Crônica , Células-Tronco Pluripotentes Induzidas , Animais , Humanos , Camundongos , Ratos , Macaca , Receptores Opioides , Receptores Opioides mu/genética , TransgenesRESUMO
Hematopoietic stem cell transplantation (HSCT) can replace endogenous microglia with circulation-derived macrophages but has high mortality. To mitigate the risks of HSCT and expand the potential for microglia replacement, we engineered an inhibitor-resistant CSF1R that enables robust microglia replacement. A glycine to alanine substitution at position 795 of human CSF1R (G795A) confers resistance to multiple CSF1R inhibitors, including PLX3397 and PLX5622. Biochemical and cell-based assays show no discernable gain or loss of function. G795A- but not wildtype-CSF1R expressing macrophages efficiently engraft the brain of PLX3397-treated mice and persist after cessation of inhibitor treatment. To gauge translational potential, we CRISPR engineered human-induced pluripotent stem cell-derived microglia (iMG) to express G795A. Xenotransplantation studies demonstrate that G795A-iMG exhibit nearly identical gene expression to wildtype iMG, respond to inflammatory stimuli, and progressively expand in the presence of PLX3397, replacing endogenous microglia to fully occupy the brain. In sum, we engineered a human CSF1R variant that enables nontoxic, cell type, and tissue-specific replacement of microglia.
Assuntos
Microglia , Engenharia de Proteínas , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos , Animais , Humanos , Camundongos , Aminopiridinas/farmacologia , Encéfalo/metabolismo , Microglia/metabolismo , Engenharia de Proteínas/métodos , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/genética , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Terapia Baseada em Transplante de Células e Tecidos/métodosRESUMO
SUMMARY: Quantitative analysis of continuous electroencephalography (QEEG) is increasingly being used to augment seizure detection in critically ill patients. Typically, seizures manifest on QEEG as abrupt increases in power and frequency, a visual pattern often called "flames." Here, we present a case of a 16-year-old patient with intractable Lennox-Gastaut syndrome secondary to a pathogenic variant in the SCN2A gene who had tonic seizures that manifest as abrupt decreases in power on QEEG, a visual pattern we term "icicles." Recognition of QEEG patterns representative of different seizure types is important as QEEG use becomes more widespread including in pediatric populations.
Assuntos
Síndrome de Lennox-Gastaut , Criança , Humanos , Adolescente , Síndrome de Lennox-Gastaut/diagnóstico , Síndrome de Lennox-Gastaut/complicações , Convulsões/diagnóstico , Convulsões/complicações , EletroencefalografiaRESUMO
The molecular diversity of glia in the human hippocampus and their temporal dynamics over the lifespan remain largely unknown. Here, we performed single-nucleus RNA sequencing to generate a transcriptome atlas of the human hippocampus across the postnatal lifespan. Detailed analyses of astrocytes, oligodendrocyte lineages, and microglia identified subpopulations with distinct molecular signatures and revealed their association with specific physiological functions, age-dependent changes in abundance, and disease relevance. We further characterized spatiotemporal heterogeneity of GFAP-enriched astrocyte subpopulations in the hippocampal formation using immunohistology. Leveraging glial subpopulation classifications as a reference map, we revealed the diversity of glia differentiated from human pluripotent stem cells and identified dysregulated genes and pathological processes in specific glial subpopulations in Alzheimer's disease (AD). Together, our study significantly extends our understanding of human glial diversity, population dynamics across the postnatal lifespan, and dysregulation in AD and provides a reference atlas for stem-cell-based glial differentiation.
Assuntos
Doença de Alzheimer , Transcriptoma , Humanos , Transcriptoma/genética , Longevidade/genética , Neuroglia/patologia , Hipocampo , Astrócitos/patologia , Doença de Alzheimer/patologiaRESUMO
Viral infections of the central nervous system (CNS) are a significant cause of neurological impairment and mortality worldwide. As tissue resident macrophages, microglia are critical initial responders to CNS viral infection. Microglia seem to coordinate brain-wide antiviral responses of both brain resident cells and infiltrating immune cells. This review discusses how microglia may promote this antiviral response at a molecular level, from potential mechanisms of virus recognition to downstream cytokine responses and interaction with antiviral T cells. Recent advancements in genetic tools to specifically target microglia in vivo promise to further our understanding about the precise mechanistic role of microglia in CNS infection.
Assuntos
Antivirais , Microglia , Humanos , Encéfalo , Medula Espinal , Sistema Nervoso CentralRESUMO
Dissecting contributions of microglia to human brain development and disease pathogenesis requires modeling interactions between these microglia and their local environment. In this issue of Cell Stem Cell, Popova et al. (2021) propose a transcriptomic "microglia report card" and create a neuroimmune organoid to model complex interactions involving human microglia.
Assuntos
Microglia , Organoides , Encéfalo , Humanos , Transcriptoma/genéticaRESUMO
In injury and disease, microglia and astrocytes - two major non-neuronal cell types in the central nervous system (CNS) - undergo morphological, transcriptional, and functional changes, which can underlie pathogenesis and dysfunction of the CNS. Microglia, the brain's tissue resident parenchymal macrophages, are described as becoming "activated" as they deftly change their production of different inflammatory mediators, alter the surveillance behavior of their cellular protrusions, and differentially influence the function of astrocytes. For their part, astrocytes - the most abundant glial cell type - are said to become "reactive", which implies (perhaps inappropriately) causality for the changes astrocytes undergo. Reactive astrocytes variably undergo process hypertrophy, decrease their normal homeostatic functions such as facilitating synapse formation, and in some cases act to form a tissue scar in response to insult. But what do these terms "activation" and "reactivity" mean, anyway? And how do these changed microglia and astrocytes contribute to neurodegenerative disease (ND)? Here, we describe our current understanding of the role of activated and reactive microglia and astrocytes in ND, as well as our current understanding about what these states are and might mean. We survey the earliest description of these cells by histopathologists, their transcriptomic identities, and finally our mechanistic understanding of their functions in ND.
Assuntos
Astrócitos/metabolismo , Microglia/metabolismo , Doenças Neurodegenerativas/genética , Astrócitos/patologia , Humanos , Microglia/patologia , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/patologia , Neuroglia/metabolismo , Neuroglia/patologiaRESUMO
Microglia, the brain's tissue-resident macrophages, contribute to the developmental elimination of extranumerary synapses and to pathologic synapse loss in mouse models of neurodegeneration. Two papers published in The EMBO Journal reveal that phosphatidylserine (PS) is a neuronal cue for microglial synapse elimination.
Assuntos
Microglia , Fosfatidilserinas , Animais , Apoptose , Sinais (Psicologia) , Camundongos , Isoformas de Proteínas , Receptores Acoplados a Proteínas G , SinapsesRESUMO
The tumor microenvironment (TME) is critical for tumor progression. However, the establishment and function of the TME remain obscure because of its complex cellular composition. Using a mouse genetic system called mosaic analysis with double markers (MADMs), we delineated TME evolution at single-cell resolution in sonic hedgehog (SHH)-activated medulloblastomas that originate from unipotent granule neuron progenitors in the brain. First, we found that astrocytes within the TME (TuAstrocytes) were trans-differentiated from tumor granule neuron precursors (GNPs), which normally never differentiate into astrocytes. Second, we identified that TME-derived IGF1 promotes tumor progression. Third, we uncovered that insulin-like growth factor 1 (IGF1) is produced by tumor-associated microglia in response to interleukin-4 (IL-4) stimulation. Finally, we found that IL-4 is secreted by TuAstrocytes. Collectively, our studies reveal an evolutionary process that produces a multi-lateral network within the TME of medulloblastoma: a fraction of tumor cells trans-differentiate into TuAstrocytes, which, in turn, produce IL-4 that stimulates microglia to produce IGF1 to promote tumor progression.
Assuntos
Astrócitos/metabolismo , Carcinogênese/metabolismo , Transdiferenciação Celular , Neoplasias Cerebelares/metabolismo , Meduloblastoma/metabolismo , Comunicação Parácrina , Animais , Linhagem da Célula , Neoplasias Cerebelares/patologia , Modelos Animais de Doenças , Feminino , Proteínas Hedgehog/metabolismo , Xenoenxertos , Humanos , Fator de Crescimento Insulin-Like I/genética , Fator de Crescimento Insulin-Like I/metabolismo , Interleucina-4/genética , Interleucina-4/metabolismo , Masculino , Meduloblastoma/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neurônios/metabolismo , Microambiente TumoralRESUMO
Microglia are the tissue-resident macrophages of the brain and spinal cord. They are critical players in the development, normal function, and decline of the CNS. Unlike traditional monocyte-derived macrophages, microglia originate from primitive hematopoiesis in the embryonic yolk sac and self-renew throughout life. Microglia also have a unique genetic signature among tissue resident macrophages. Recent studies identify the contributions of both brain environment and developmental history to the transcriptomic identity of microglia. Here we review this emerging literature and discuss the potential implications of origin on microglial function, with particular focus on existing and future therapies using bone-marrow- or stem-cell-derived cells for the treatment of neurological diseases.
Assuntos
Encéfalo/citologia , Microglia/citologia , Animais , Linhagem da Célula/fisiologia , Humanos , Macrófagos/citologiaRESUMO
Glioblastoma multiforme (GBM) is a highly aggressive malignant brain tumor with fatal outcome. Tumor-associated macrophages and microglia (TAMs) have been found to be major tumor-promoting immune cells in the tumor microenvironment. Hence, modulation and reeducation of tumor-associated macrophages and microglia in GBM is considered a promising antitumor strategy. Resident microglia and invading macrophages have been shown to have distinct origin and function. Whereas yolk sac-derived microglia reside in the brain, blood-derived monocytes invade the central nervous system only under pathological conditions like tumor formation. We recently showed that disruption of the SIRPα-CD47 signaling axis is efficacious against various brain tumors including GBM primarily by inducing tumor phagocytosis. However, most effects are attributed to macrophages recruited from the periphery but the role of the brain resident microglia is unknown. Here, we sought to utilize a model to distinguish resident microglia and peripheral macrophages within the GBM-TAM pool, using orthotopically xenografted, immunodeficient, and syngeneic mouse models with genetically color-coded macrophages (Ccr2RFP) and microglia (Cx3cr1GFP). We show that even in the absence of phagocytizing macrophages (Ccr2RFP/RFP), microglia are effector cells of tumor cell phagocytosis in response to anti-CD47 blockade. Additionally, macrophages and microglia show distinct morphological and transcriptional changes. Importantly, the transcriptional profile of microglia shows less of an inflammatory response which makes them a promising target for clinical applications.
Assuntos
Neoplasias Encefálicas/imunologia , Antígeno CD47/imunologia , Glioblastoma/imunologia , Microglia/imunologia , Proteínas de Neoplasias/imunologia , Neoplasias Experimentais/imunologia , Fagocitose , Receptores Imunológicos/imunologia , Transdução de Sinais/imunologia , Animais , Neoplasias Encefálicas/patologia , Antígeno CD47/genética , Glioblastoma/genética , Glioblastoma/patologia , Macrófagos/imunologia , Macrófagos/patologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos Transgênicos , Microglia/patologia , Monócitos/imunologia , Monócitos/patologia , Proteínas de Neoplasias/genética , Neoplasias Experimentais/genética , Neoplasias Experimentais/patologia , Receptores Imunológicos/genética , Transdução de Sinais/genéticaRESUMO
Microglia are increasingly recognized for their major contributions during brain development and neurodegenerative disease. It is currently unknown whether these functions are carried out by subsets of microglia during different stages of development and adulthood or within specific brain regions. Here, we performed deep single-cell RNA sequencing (scRNA-seq) of microglia and related myeloid cells sorted from various regions of embryonic, early postnatal, and adult mouse brains. We found that the majority of adult microglia expressing homeostatic genes are remarkably similar in transcriptomes, regardless of brain region. By contrast, early postnatal microglia are more heterogeneous. We discovered a proliferative-region-associated microglia (PAM) subset, mainly found in developing white matter, that shares a characteristic gene signature with degenerative disease-associated microglia (DAM). Such PAM have amoeboid morphology, are metabolically active, and phagocytose newly formed oligodendrocytes. This scRNA-seq atlas will be a valuable resource for dissecting innate immune functions in health and disease.
Assuntos
Encéfalo , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Microglia/fisiologia , Células Mieloides/fisiologia , Análise de Sequência de RNA , Transcriptoma/fisiologia , Algoritmos , Animais , Animais Recém-Nascidos , Antígenos CD/metabolismo , Encéfalo/citologia , Encéfalo/embriologia , Encéfalo/crescimento & desenvolvimento , Proliferação de Células/fisiologia , Plexo Corióideo/citologia , Análise por Conglomerados , Simulação por Computador , Embrião de Mamíferos , Redes Reguladoras de Genes/fisiologia , Sequenciamento de Nucleotídeos em Larga Escala , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Oligodendroglia/fisiologia , Fagocitose/fisiologiaRESUMO
Microglia represent 5-10% of cells in the central nervous system and contribute to the development, homeostasis, injury, and repair of neural tissues. As the tissue-resident macrophages of the central nervous system, microglia execute core innate immune functions such as detection of pathogens/damage, cytokine secretion, and phagocytosis. However, additional properties that are specific to microglia and their neural environment are beginning to be appreciated. This article describes approaches for purification of microglia by fluorescence-activated cell sorting using microglia-specific surface markers and for enrichment of microglia by magnetic sorting and immunopanning. Detailed information about culturing primary microglia at various developmental stages is also provided. Throughout, we focus on special considerations for handling microglia and compare the relative strengths or disadvantages of different protocols. © 2018 by John Wiley & Sons, Inc.
Assuntos
Técnicas de Cultura de Células , Citometria de Fluxo , Microglia , Animais , Camundongos , RatosRESUMO
Microglia, the brain's resident macrophages, are dynamic CNS custodians with surprising origins in the extra-embryonic yolk sac. The consequences of their distinct ontogeny are unknown but critical to understanding and treating brain diseases. We created a brain macrophage transplantation system to disentangle how environment and ontogeny specify microglial identity. We find that donor cells extensively engraft in the CNS of microglia-deficient mice, and even after exposure to a cell culture environment, microglia fully regain their identity when returned to the CNS. Though transplanted macrophages from multiple tissues can express microglial genes in the brain, only those of yolk-sac origin fully attain microglial identity. Transplanted macrophages of inappropriate origin, including primary human cells in a humanized host, express disease-associated genes and specific ontogeny markers. Through brain macrophage transplantation, we discover new principles of microglial identity that have broad applications to the study of disease and development of myeloid cell therapies.
Assuntos
Encéfalo/citologia , Linhagem da Célula , Células-Tronco Hematopoéticas/citologia , Macrófagos/citologia , Microglia/citologia , Animais , Encéfalo/metabolismo , Sistema Nervoso Central , Humanos , Macrófagos/metabolismo , Camundongos , Camundongos Knockout , Microglia/metabolismo , Receptores de Fator Estimulador das Colônias de Granulócitos e MacrófagosRESUMO
Stromal cells (SCs) establish the compartmentalization of lymphoid tissues critical to the immune response. However, the full diversity of lymph node (LN) SCs remains undefined. Using droplet-based single-cell RNA sequencing, we identified nine peripheral LN non-endothelial SC clusters. Included are the established subsets, Ccl19hi T-zone reticular cells (TRCs), marginal reticular cells, follicular dendritic cells (FDCs), and perivascular cells. We also identified Ccl19lo TRCs, likely including cholesterol-25-hydroxylase+ cells located at the T-zone perimeter, Cxcl9+ TRCs in the T-zone and interfollicular region, CD34+ SCs in the capsule and medullary vessel adventitia, indolethylamine N-methyltransferase+ SCs in the medullary cords, and Nr4a1+ SCs in several niches. These data help define how transcriptionally distinct LN SCs support niche-restricted immune functions and provide evidence that many SCs are in an activated state.