RESUMEN
Microglia are specialized macrophages in the brain parenchyma that exist in multiple transcriptional states and reside within a wide range of neuronal environments1-4. However, how and where these states are generated remains poorly understood. Here, using the mouse somatosensory cortex, we demonstrate that microglia density and molecular state acquisition are determined by the local composition of pyramidal neuron classes. Using single-cell and spatial transcriptomic profiling, we unveil the molecular signatures and spatial distributions of diverse microglia populations and show that certain states are enriched in specific cortical layers, whereas others are broadly distributed throughout the cortex. Notably, conversion of deep-layer pyramidal neurons to an alternate class identity reconfigures the distribution of local, layer-enriched homeostatic microglia to match the new neuronal niche. Leveraging the transcriptional diversity of pyramidal neurons in the neocortex, we construct a ligand-receptor atlas describing interactions between individual pyramidal neuron subtypes and microglia states, revealing rules of neuron-microglia communication. Our findings uncover a fundamental role for neuronal diversity in instructing the acquisition of microglia states as a potential mechanism for fine-tuning neuroimmune interactions within the cortical local circuitry.
Asunto(s)
Microglía , Neocórtex , Células Piramidales , Corteza Somatosensorial , Animales , Recuento de Células , Ratones , Microglía/clasificación , Microglía/fisiología , Neocórtex/citología , Neocórtex/fisiología , Células Piramidales/clasificación , Células Piramidales/fisiología , Análisis de la Célula Individual , Corteza Somatosensorial/citología , Corteza Somatosensorial/fisiología , TranscriptomaRESUMEN
In utero infection and maternal inflammation can adversely impact fetal brain development. Maternal systemic illness, even in the absence of direct fetal brain infection, is associated with an increased risk of neuropsychiatric disorders in affected offspring. The cell types mediating the fetal brain response to maternal inflammation are largely unknown, hindering the development of novel treatment strategies. Here, we show that microglia, the resident phagocytes of the brain, highly express receptors for relevant pathogens and cytokines throughout embryonic development. Using a rodent maternal immune activation (MIA) model in which polyinosinic:polycytidylic acid is injected into pregnant mice, we demonstrate long-lasting transcriptional changes in fetal microglia that persist into postnatal life. We find that MIA induces widespread gene expression changes in neuronal and non-neuronal cells; importantly, these responses are abolished by selective genetic deletion of microglia, indicating that microglia are required for the transcriptional response of other cortical cell types to MIA. These findings demonstrate that microglia play a crucial durable role in the fetal response to maternal inflammation, and should be explored as potential therapeutic cell targets.
Asunto(s)
Encéfalo , Inflamación , Microglía , Poli I-C , Animales , Microglía/metabolismo , Microglía/inmunología , Femenino , Embarazo , Ratones , Encéfalo/patología , Encéfalo/inmunología , Encéfalo/metabolismo , Inflamación/patología , Inflamación/genética , Poli I-C/farmacología , Feto , Ratones Endogámicos C57BL , Regulación del Desarrollo de la Expresión Génica , Neuronas/metabolismoRESUMEN
The mammalian cerebral cortex comprises a complex neuronal network that maintains a delicate balance between excitatory neurons and inhibitory interneurons. Previous studies, including our own research, have shown that specific interneuron subtypes are closely associated with particular pyramidal neuron types, forming stereotyped local inhibitory microcircuits. However, the developmental processes that establish these precise networks are not well understood. Here we show that pyramidal neuron types are instrumental in driving the terminal differentiation and maintaining the survival of specific associated interneuron subtypes. In a wild-type cortex, the relative abundance of different interneuron subtypes aligns precisely with the pyramidal neuron types to which they synaptically target. In Fezf2 mutant cortex, characterized by the absence of layer 5 pyramidal tract neurons and an expansion of layer 6 intratelencephalic neurons, we observed a corresponding decrease in associated layer 5b interneurons and an increase in layer 6 subtypes. Interestingly, these shifts in composition are achieved through mechanisms specific to different interneuron types. While SST interneurons adjust their abundance to the change in pyramidal neuron prevalence through the regulation of programmed cell death, parvalbumin interneurons alter their identity. These findings illustrate two key strategies by which the dynamic interplay between pyramidal neurons and interneurons allows local microcircuits to be sculpted precisely. These insights underscore the precise roles of extrinsic signals from pyramidal cells in the establishment of interneuron diversity and their subsequent integration into local cortical microcircuits.
RESUMEN
Receptors for advanced glycation end-products (RAGE) are multiligand surface receptors detected abundantly in pulmonary tissue. Our previous work revealed increased RAGE expression in cells and lungs exposed to tobacco smoke and RAGE-mediated cytokine expression via proinflammatory mechanisms involving NF-κB. RAGE expression is elevated in various pathological states, including chronic obstructive pulmonary disease; however, precise contributions of RAGE to the progression of emphysema and pulmonary inflammation in the adult lung are unknown. In the current study, we generated a RAGE transgenic (RAGE TG) mouse and conditionally induced adult alveolar epithelium to overexpress RAGE. RAGE was induced after the period of alveologenesis, from weaning (20 d of age) until animals were killed at 50, 80, and 110 days (representing 30, 60, and 90 d of RAGE overexpression). Hematoxylin and eosin staining and mean chord length revealed incremental dilation of alveolar spaces as RAGE overexpression persisted. TUNEL staining and electron microscopy confirmed increased apoptosis and blebbing of alveolar epithelium in lungs from RAGE TG mice when compared with control mice. Immunohistochemistry for matrix metalloproteinase 9 revealed an overall increase in matrix metalloproteinase 9, which correlated with decreased elastin expression in RAGE TG mice. Furthermore, RAGE TG mice manifested significant inflammation measured by elevated bronchoalveolar lavage protein, leukocyte infiltration, and secreted cytokines. These data support the concept that innovative transgenic mice that overexpress RAGE may model pulmonary inflammation and alveolar destabilization independent of tobacco smoke and validate RAGE signaling as a target pathway in the prevention or attenuation of smoke-related inflammatory lung diseases.
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Pulmón/patología , Neumonía/patología , Alveolos Pulmonares/ultraestructura , Enfisema Pulmonar/patología , Receptores Inmunológicos/metabolismo , Animales , Apoptosis , Doxiciclina/farmacología , Elastina/genética , Elastina/metabolismo , Regulación de la Expresión Génica , Inmunohistoquímica , Etiquetado Corte-Fin in Situ , Pulmón/metabolismo , Metaloproteinasa 9 de la Matriz/genética , Metaloproteinasa 9 de la Matriz/metabolismo , Ratones , Ratones Transgénicos , Microscopía Electrónica , Neumonía/metabolismo , Proteolisis , Alveolos Pulmonares/efectos de los fármacos , Receptor para Productos Finales de Glicación Avanzada , Receptores Inmunológicos/genética , Coloración y Etiquetado , Regulación hacia Arriba , DesteteRESUMEN
BACKGROUND: Receptors for advanced glycation end-products (RAGE) are cell surface receptors prominently expressed by lung epithelium. Previous research demonstrated that over-expression of RAGE by murine alveolar epithelial cells during embryogenesis caused severe lung hypoplasia and neonatal lethality. However, the effects of RAGE over-expression on adjacent matrix and endothelial cells remained unknown. METHODS: RAGE transgenic (TG) mice were generated that conditionally over-expressed RAGE in alveolar type II cells when fed doxycycline (dox) from conception to E18.5. To evaluate effects on the basement membrane, immunostaining and immunoblotting were performed for collagen IV and MMP-9, a matrix metalloprotease capable of degrading basement membranes. To assess changes in vasculature, immunostaining, immunoblotting and qRT-PCR were performed for Pecam-1, a platelet endothelial cell adhesion marker also known as CD31. Lastly, to characterize potential regulatory mechanisms of endothelial cell differentiation, immunoblotting and qRT-PCR for FoxM1, a key endothelium-specific transcription factor of the Forkhead Box (Fox) family, were completed. RESULTS: Qualitative immunostaining for collagen IV was less in RAGE TG mice compared to controls and immunoblotting revealed decreased collagen IV in the RAGE TG mouse lung. Additionally, elevated MMP-9 detected via immunostaining and immunoblotting implicated MMP-9 as a possible down stream effector in matrix destabilization mediated by RAGE signaling. Lastly, Pecam-1 assessment revealed a decrease in the prevalence of microvascular endothelial cells coincident with FoxM1 abrogation in RAGE TG mice compared to controls. CONCLUSIONS: RAGE over-expression by alveolar epithelium weakened the basement membrane and associated matrix via increased MMP-9 activity. Furthermore, over-expression of RAGE inhibited FoxM1, suggesting that anomalous transcriptional control contributes to decreased endothelial cell prevalence in the TG mouse lung.
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Membrana Basal/metabolismo , Endotelio/patología , Alveolos Pulmonares/embriología , Alveolos Pulmonares/metabolismo , Receptores Inmunológicos/metabolismo , Animales , Diferenciación Celular/fisiología , Colágeno/metabolismo , Endotelio/metabolismo , Epitelio/embriología , Epitelio/metabolismo , Factores de Transcripción Forkhead/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Modelos Animales , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/metabolismo , Receptor para Productos Finales de Glicación Avanzada , Receptores Inmunológicos/genéticaRESUMEN
Astrocytes are the most abundant glial cell type in the central nervous system and are essential to the development, plasticity, and maintenance of neural circuits. Astrocytes are heterogeneous, with their diversity rooted in developmental programs modulated by the local brain environment. Astrocytes play integral roles in regulating and coordinating neural activity extending far beyond their metabolic support of neurons and other brain cell phenotypes. Both gray and white matter astrocytes occupy critical functional niches capable of modulating brain physiology on time scales slower than synaptic activity but faster than those adaptive responses requiring a structural change or adaptive myelination. Given their many associations and functional roles, it is not surprising that astrocytic dysfunction has been causally implicated in a broad set of neurodegenerative and neuropsychiatric disorders. In this review, we focus on recent discoveries concerning the contributions of astrocytes to the function of neural networks, with a dual focus on the contribution of astrocytes to synaptic development and maturation, and on their role in supporting myelin integrity, and hence conduction and its regulation. We then address the emerging roles of astrocytic dysfunction in disease pathogenesis and on potential strategies for targeting these cells for therapeutic purposes.
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Astrocitos , Neuroglía , Humanos , Astrocitos/fisiología , Neuronas/metabolismo , Vaina de Mielina , Redes Neurales de la ComputaciónRESUMEN
Receptors for advanced glycation end-products (RAGEs) are multiligand cell surface receptors highly expressed in the lung that contribute to alveolar epithelial cell differentiation during embryogenesis and the modulation of pulmonary inflammation during disease. When RAGEs are overexpressed throughout embryogenesis, severe lung hypoplasia ensues, culminating in perinatal lethality. However, the possible mechanisms that lead to the disappearance of pulmonary tissue remain unclear. A time course of lung organogenesis, commencing on Embryonic Day (E) 12.5, demonstrated that increased RAGE expression primarily alters lung morphogenesis beginning on E16.5. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) immunohistochemistry and immunoblotting for active caspase-3 confirmed a shift toward apoptosis in lungs from RAGE-overexpressing mice, compared with wild-type control mice. This observation supports previous work where electron microscopy identified the cellular blebbing of alveolar epithelium in embryonic RAGE-overexpressing mice. Assaying for NF-κB also revealed elevated nuclear translocation in lungs from transgenic mice compared with control mice. An RT-PCR assessment of genes regulated by NF-κB demonstrated the elevated expression of Fas ligand, suggesting increased activity of the Fas-mediated signal transduction pathway in which ligand-receptor interactions trigger cell death. These data provide evidence that the expression of RAGEs must be tightly regulated during homeostatic organogenesis. Further elucidations of the RAGE signaling potentially involved in cell-cycle abnormalities may provide insights into the progression of RAGE-mediated lung diseases.
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Células Epiteliales Alveolares/citología , Células Epiteliales Alveolares/metabolismo , Apoptosis , Proliferación Celular , Pulmón/embriología , Receptores Inmunológicos/metabolismo , Animales , Núcleo Celular/metabolismo , Proteína Ligando Fas/biosíntesis , Pulmón/anomalías , Pulmón/citología , Pulmón/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , FN-kappa B/metabolismo , Organogénesis , Receptor para Productos Finales de Glicación Avanzada , Receptores Inmunológicos/genética , Transducción de SeñalRESUMEN
Receptors for advanced glycation end-products (RAGE) are cell-surface receptors expressed by pulmonary tissue that influence alveolar type (AT) II-ATI transition required for normal alveolar formation. However, the precise contribution of RAGE in interactions between pulmonary epithelium and splanchnic mesenchyme during lung organogenesis remains uncertain. To test the hypothesis that RAGE misexpression adversely affects lung morphogenesis, conditional transgenic mice were generated that overexpress RAGE. Mice that overexpress RAGE throughout embryogenesis experienced 100% mortality and significant lung hypoplasia coincident with large, vacuous areas in the periphery when compared with normal airway and alveolar architecture observed in control mouse lungs. Flow cytometry and immunohistochemistry employing cell-specific markers for distal (forkhead box protein A2) and respiratory (thyroid transcription factor-1) epithelium, ATII cells (pro-surfactant protein-C), and ATI cells (T1-α) demonstrated anomalies in key epithelial cell populations resulting from RAGE up-regulation. These results reveal that precise regulation of RAGE expression is required during lung formation. Furthermore, abundant RAGE results in profound alterations in epithelial cell differentiation that culminate in severe respiratory distress and perinatal lethality.
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Diferenciación Celular , Alveolos Pulmonares/metabolismo , Receptores Inmunológicos/biosíntesis , Síndrome de Dificultad Respiratoria del Recién Nacido/metabolismo , Mucosa Respiratoria/metabolismo , Regulación hacia Arriba , Animales , Animales Recién Nacidos , Antígenos de Diferenciación/genética , Antígenos de Diferenciación/metabolismo , Humanos , Recién Nacido , Ratones , Ratones Noqueados , Organogénesis/genética , Alveolos Pulmonares/patología , Receptor para Productos Finales de Glicación Avanzada , Receptores Inmunológicos/genética , Síndrome de Dificultad Respiratoria del Recién Nacido/genética , Síndrome de Dificultad Respiratoria del Recién Nacido/patología , Mucosa Respiratoria/patologíaRESUMEN
Chronic obstructive pulmonary disease (COPD) is a progressive condition characterized by chronic airway inflammation and airspace remodeling, leading to airflow limitation that is not completely reversible. Smoking is the leading risk factor for compromised lung function stemming from COPD pathogenesis. First- and second-hand cigarette smoke contain thousands of constituents, including several carcinogens and cytotoxic chemicals that orchestrate chronic lung inflammation and destructive alveolar remodeling. Receptors for advanced glycation end-products (RAGE) are multi-ligand cell surface receptors primarily expressed by diverse lung cells. RAGE expression increases following cigarette smoke exposure and expression is elevated in the lungs of patients with COPD. RAGE is responsible in part for inducing pro-inflammatory signaling pathways that culminate in expression and secretion of several cytokines, chemokines, enzymes, and other mediators. In the current review, new transgenic mouse models that conditionally over-express RAGE in pulmonary epithelium are discussed. When RAGE is over-expressed throughout embryogenesis, apoptosis in the peripheral lung causes severe lung hypoplasia. Interestingly, apoptosis in RAGE transgenic mice occurs via conserved apoptotic pathways also known to function in advanced stages of COPD. RAGE over-expression in the adult lung models features of COPD including pronounced inflammation and loss of parenchymal tissue. Understanding the biological contributions of RAGE during cigarette smoke-induced inflammation may provide critically important insight into the pathology of COPD.