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The placenta is a selective maternal-fetal barrier that provides nourishment and protection from infections. However, certain pathogens can attach to and even cross the placenta, causing pregnancy complications with potential lifelong impacts on the child's health. Here, we profiled at the single-cell level the placental responses to three pathogens associated with intrauterine complications-Plasmodium falciparum, Listeria monocytogenes, and Toxoplasma gondii. We found that upon exposure to the pathogens, all placental lineages trigger inflammatory responses that may compromise placental function. Additionally, we characterized the responses of fetal macrophages known as Hofbauer cells (HBCs) to each pathogen and propose that they are the probable niche for T. gondii. Finally, we revealed how P. falciparum adapts to the placental microenvironment by modulating protein export into the host erythrocyte and nutrient uptake pathways. Altogether, we have defined the cellular networks and signaling pathways mediating acute placental inflammatory responses that could contribute to pregnancy complications.
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Placenta , Análise de Célula Única , Humanos , Feminino , Gravidez , Placenta/microbiologia , Placenta/imunologia , Análise de Célula Única/métodos , Plasmodium falciparum , Listeria monocytogenes/patogenicidade , Listeria monocytogenes/fisiologia , Toxoplasma/patogenicidade , Macrófagos/microbiologia , Macrófagos/imunologia , Macrófagos/metabolismo , Toxoplasmose/imunologia , Toxoplasmose/metabolismo , InflamaçãoRESUMO
Human limbs emerge during the fourth post-conception week as mesenchymal buds, which develop into fully formed limbs over the subsequent months1. This process is orchestrated by numerous temporally and spatially restricted gene expression programmes, making congenital alterations in phenotype common2. Decades of work with model organisms have defined the fundamental mechanisms underlying vertebrate limb development, but an in-depth characterization of this process in humans has yet to be performed. Here we detail human embryonic limb development across space and time using single-cell and spatial transcriptomics. We demonstrate extensive diversification of cells from a few multipotent progenitors to myriad differentiated cell states, including several novel cell populations. We uncover two waves of human muscle development, each characterized by different cell states regulated by separate gene expression programmes, and identify musculin (MSC) as a key transcriptional repressor maintaining muscle stem cell identity. Through assembly of multiple anatomically continuous spatial transcriptomic samples using VisiumStitcher, we map cells across a sagittal section of a whole fetal hindlimb. We reveal a clear anatomical segregation between genes linked to brachydactyly and polysyndactyly, and uncover transcriptionally and spatially distinct populations of the mesenchyme in the autopod. Finally, we perform single-cell RNA sequencing on mouse embryonic limbs to facilitate cross-species developmental comparison, finding substantial homology between the two species.
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T cells develop from circulating precursors, which enter the thymus and migrate throughout specialised sub-compartments to support maturation and selection. This process starts already in early fetal development and is highly active until the involution of the thymus in adolescence. To map the micro-anatomical underpinnings of this process in pre- vs. post-natal states, we undertook a spatially resolved analysis and established a new quantitative morphological framework for the thymus, the Cortico-Medullary Axis. Using this axis in conjunction with the curation of a multimodal single-cell, spatial transcriptomics and high-resolution multiplex imaging atlas, we show that canonical thymocyte trajectories and thymic epithelial cells are highly organised and fully established by post-conception week 12, pinpoint TEC progenitor states, find that TEC subsets and peripheral tissue genes are associated with Hassall's Corpuscles and uncover divergence in the pace and drivers of medullary entry between CD4 vs. CD8 T cell lineages. These findings are complemented with a holistic toolkit for spatial analysis and annotation, providing a basis for a detailed understanding of T lymphocyte development.
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The extraembryonic yolk sac (YS) ensures delivery of nutritional support and oxygen to the developing embryo but remains ill-defined in humans. We therefore assembled a comprehensive multiomic reference of the human YS from 3 to 8 postconception weeks by integrating single-cell protein and gene expression data. Beyond its recognized role as a site of hematopoiesis, we highlight roles in metabolism, coagulation, vascular development, and hematopoietic regulation. We reconstructed the emergence and decline of YS hematopoietic stem and progenitor cells from hemogenic endothelium and revealed a YS-specific accelerated route to macrophage production that seeds developing organs. The multiorgan functions of the YS are superseded as intraembryonic organs develop, effecting a multifaceted relay of vital functions as pregnancy proceeds.
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Desenvolvimento Embrionário , Saco Vitelino , Feminino , Humanos , Gravidez , Coagulação Sanguínea/genética , Macrófagos , Saco Vitelino/citologia , Saco Vitelino/metabolismo , Desenvolvimento Embrionário/genética , Atlas como Assunto , Expressão Gênica , Perfilação da Expressão Gênica , Hematopoese/genética , Fígado/embriologiaRESUMO
The relationship between the human placenta-the extraembryonic organ made by the fetus, and the decidua-the mucosal layer of the uterus, is essential to nurture and protect the fetus during pregnancy. Extravillous trophoblast cells (EVTs) derived from placental villi infiltrate the decidua, transforming the maternal arteries into high-conductance vessels1. Defects in trophoblast invasion and arterial transformation established during early pregnancy underlie common pregnancy disorders such as pre-eclampsia2. Here we have generated a spatially resolved multiomics single-cell atlas of the entire human maternal-fetal interface including the myometrium, which enables us to resolve the full trajectory of trophoblast differentiation. We have used this cellular map to infer the possible transcription factors mediating EVT invasion and show that they are preserved in in vitro models of EVT differentiation from primary trophoblast organoids3,4 and trophoblast stem cells5. We define the transcriptomes of the final cell states of trophoblast invasion: placental bed giant cells (fused multinucleated EVTs) and endovascular EVTs (which form plugs inside the maternal arteries). We predict the cell-cell communication events contributing to trophoblast invasion and placental bed giant cell formation, and model the dual role of interstitial EVTs and endovascular EVTs in mediating arterial transformation during early pregnancy. Together, our data provide a comprehensive analysis of postimplantation trophoblast differentiation that can be used to inform the design of experimental models of the human placenta in early pregnancy.
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Multiômica , Primeiro Trimestre da Gravidez , Trofoblastos , Feminino , Humanos , Gravidez , Movimento Celular , Placenta/irrigação sanguínea , Placenta/citologia , Placenta/fisiologia , Primeiro Trimestre da Gravidez/fisiologia , Trofoblastos/citologia , Trofoblastos/metabolismo , Trofoblastos/fisiologia , Decídua/irrigação sanguínea , Decídua/citologia , Relações Materno-Fetais/fisiologia , Análise de Célula Única , Miométrio/citologia , Miométrio/fisiologia , Diferenciação Celular , Organoides/citologia , Organoides/fisiologia , Células-Tronco/citologia , Transcriptoma , Fatores de Transcrição/metabolismo , Comunicação CelularRESUMO
Spatial transcriptomic technologies promise to resolve cellular wiring diagrams of tissues in health and disease, but comprehensive mapping of cell types in situ remains a challenge. Here we present Ñell2location, a Bayesian model that can resolve fine-grained cell types in spatial transcriptomic data and create comprehensive cellular maps of diverse tissues. Cell2location accounts for technical sources of variation and borrows statistical strength across locations, thereby enabling the integration of single-cell and spatial transcriptomics with higher sensitivity and resolution than existing tools. We assessed cell2location in three different tissues and show improved mapping of fine-grained cell types. In the mouse brain, we discovered fine regional astrocyte subtypes across the thalamus and hypothalamus. In the human lymph node, we spatially mapped a rare pre-germinal center B cell population. In the human gut, we resolved fine immune cell populations in lymphoid follicles. Collectively, our results present Ñell2location as a versatile analysis tool for mapping tissue architectures in a comprehensive manner.
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Análise de Célula Única , Transcriptoma , Animais , Teorema de Bayes , Camundongos , Análise de Célula Única/métodos , Transcriptoma/genéticaRESUMO
The endometrium, the mucosal lining of the uterus, undergoes dynamic changes throughout the menstrual cycle in response to ovarian hormones. We have generated dense single-cell and spatial reference maps of the human uterus and three-dimensional endometrial organoid cultures. We dissect the signaling pathways that determine cell fate of the epithelial lineages in the lumenal and glandular microenvironments. Our benchmark of the endometrial organoids reveals the pathways and cell states regulating differentiation of the secretory and ciliated lineages both in vivo and in vitro. In vitro downregulation of WNT or NOTCH pathways increases the differentiation efficiency along the secretory and ciliated lineages, respectively. We utilize our cellular maps to deconvolute bulk data from endometrial cancers and endometriotic lesions, illuminating the cell types dominating in each of these disorders. These mechanistic insights provide a platform for future development of treatments for common conditions including endometriosis and endometrial carcinoma.
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Endométrio/fisiologia , Ciclo Menstrual , Diferenciação Celular , Linhagem da Célula , Microambiente Celular , Neoplasias do Endométrio/patologia , Endométrio/embriologia , Endométrio/patologia , Feminino , Hormônios Esteroides Gonadais/metabolismo , Humanos , Técnicas In Vitro , Organoides , Receptores Notch/metabolismo , Transdução de Sinais , Análise Espaço-Temporal , Técnicas de Cultura de Tecidos , Transcriptoma , Útero/patologia , Proteínas Wnt/metabolismoRESUMO
The thymus provides a nurturing environment for the differentiation and selection of T cells, a process orchestrated by their interaction with multiple thymic cell types. We used single-cell RNA sequencing to create a cell census of the human thymus across the life span and to reconstruct T cell differentiation trajectories and T cell receptor (TCR) recombination kinetics. Using this approach, we identified and located in situ CD8αα+ T cell populations, thymic fibroblast subtypes, and activated dendritic cell states. In addition, we reveal a bias in TCR recombination and selection, which is attributed to genomic position and the kinetics of lineage commitment. Taken together, our data provide a comprehensive atlas of the human thymus across the life span with new insights into human T cell development.
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Atlas como Assunto , Linfócitos T CD8-Positivos/imunologia , Diferenciação Celular , Timo/crescimento & desenvolvimento , Timo/imunologia , Linfócitos T CD8-Positivos/citologia , Células Dendríticas/citologia , Células Dendríticas/imunologia , Fibroblastos/citologia , Fibroblastos/imunologia , Humanos , RNA-Seq/métodos , Receptores de Antígenos de Linfócitos T/metabolismo , Análise de Célula Única/métodos , Timo/citologiaRESUMO
The Deciphering the Mechanisms of Developmental Disorders (DMDD) program uses a systematic and standardised approach to characterise the phenotype of embryos stemming from mouse lines, which produce embryonically lethal offspring. Our study aims to provide detailed phenotype descriptions of homozygous Col4a2em1(IMPC)Wtsi mutants produced in DMDD and harvested at embryonic day 14.5. This shall provide new information on the role Col4a2 plays in organogenesis and demonstrate the capacity of the DMDD database for identifying models for researching inherited disorders. The DMDD Col4a2em1(IMPC)Wtsi mutants survived organogenesis and thus revealed the full spectrum of organs and tissues, the development of which depends on Col4a2 encoded proteins. They showed defects in the brain, cranial nerves, visual system, lungs, endocrine glands, skeleton, subepithelial tissues and mild to severe cardiovascular malformations. Together, this makes the DMDD Col4a2em1(IMPC)Wtsi line a useful model for identifying the spectrum of defects and for researching the mechanisms underlying autosomal dominant porencephaly 2 (OMIM # 614483), a rare human disease. Thus we demonstrate the general capacity of the DMDD approach and webpage as a valuable source for identifying mouse models for rare diseases.
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Large-scale phenotyping efforts have demonstrated that approximately 25-30% of mouse gene knockouts cause intrauterine lethality. Analysis of these mutants has largely focused on the embryo and not the placenta, despite the crucial role of this extraembryonic organ for developmental progression. Here we screened 103 embryonic lethal and sub-viable mouse knockout lines from the Deciphering the Mechanisms of Developmental Disorders program for placental phenotypes. We found that 68% of knockout lines that are lethal at or after mid-gestation exhibited placental dysmorphologies. Early lethality (embryonic days 9.5-14.5) is almost always associated with severe placental malformations. Placental defects correlate strongly with abnormal brain, heart and vascular development. Analysis of mutant trophoblast stem cells and conditional knockouts suggests that a considerable number of factors that cause embryonic lethality when ablated have primary gene function in trophoblast cells. Our data highlight the hugely under-appreciated importance of placental defects in contributing to abnormal embryo development and suggest key molecular nodes that govern placenta formation.
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Perda do Embrião/genética , Perda do Embrião/patologia , Mutação , Placenta/patologia , Placentação/genética , Animais , Feminino , Camundongos , Camundongos Knockout , Gravidez , Células-Tronco/metabolismo , Células-Tronco/patologia , Trofoblastos/metabolismo , Trofoblastos/patologiaRESUMO
Metastasis is the leading cause of death for cancer patients. This multi-stage process requires tumour cells to survive in the circulation, extravasate at distant sites, then proliferate; it involves contributions from both the tumour cell and tumour microenvironment ('host', which includes stromal cells and the immune system). Studies suggest the early steps of the metastatic process are relatively efficient, with the post-extravasation regulation of tumour growth ('colonization') being critical in determining metastatic outcome. Here we show the results of screening 810 mutant mouse lines using an in vivo assay to identify microenvironmental regulators of metastatic colonization. We identify 23 genes that, when disrupted in mouse, modify the ability of tumour cells to establish metastatic foci, with 19 of these genes not previously demonstrated to play a role in host control of metastasis. The largest reduction in pulmonary metastasis was observed in sphingosine-1-phosphate (S1P) transporter spinster homologue 2 (Spns2)-deficient mice. We demonstrate a novel outcome of S1P-mediated regulation of lymphocyte trafficking, whereby deletion of Spns2, either globally or in a lymphatic endothelial-specific manner, creates a circulating lymphopenia and a higher percentage of effector T cells and natural killer (NK) cells present in the lung. This allows for potent tumour cell killing, and an overall decreased metastatic burden.
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Proteínas de Transporte de Ânions/genética , Proteínas de Transporte de Ânions/metabolismo , Genoma/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/secundário , Metástase Neoplásica/genética , Metástase Neoplásica/patologia , Animais , Proteínas de Transporte de Ânions/deficiência , Linhagem Celular Tumoral , Movimento Celular , Modelos Animais de Doenças , Feminino , Genômica , Células Matadoras Naturais/citologia , Células Matadoras Naturais/imunologia , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Linfopenia/genética , Linfopenia/patologia , Lisofosfolipídeos/metabolismo , Masculino , Camundongos , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Linfócitos T/citologia , Linfócitos T/imunologia , Microambiente TumoralRESUMO
Background: Identifying genes that are essential for mouse embryonic development and survival through term is a powerful and unbiased way to discover possible genetic determinants of human developmental disorders. Characterising the changes in mouse embryos that result from ablation of lethal genes is a necessary first step towards uncovering their role in normal embryonic development and establishing any correlates amongst human congenital abnormalities. Methods: Here we present results gathered to date in the Deciphering the Mechanisms of Developmental Disorders (DMDD) programme, cataloguing the morphological defects identified from comprehensive imaging of 220 homozygous mutant and 114 wild type embryos from 42 lethal and subviable lines, analysed at E14.5. Results: Virtually all mutant embryos show multiple abnormal phenotypes and amongst the 42 lines these affect most organ systems. Within each mutant line, the phenotypes of individual embryos form distinct but overlapping sets. Subcutaneous edema, malformations of the heart or great vessels, abnormalities in forebrain morphology and the musculature of the eyes are all prevalent phenotypes, as is loss or abnormal size of the hypoglossal nerve.Conclusions: Overall, the most striking finding is that no matter how profound the malformation, each phenotype shows highly variable penetrance within a mutant line. These findings have challenging implications for efforts to identify human disease correlates.
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Knowledge of the expression profile of a gene is a critical piece of information required to build an understanding of the normal and essential functions of that gene and any role it may play in the development or progression of disease. High-throughput, large-scale efforts are on-going internationally to characterise reporter-tagged knockout mouse lines. As part of that effort, we report an open access adult mouse expression resource, in which the expression profile of 424 genes has been assessed in up to 47 different organs, tissues and sub-structures using a lacZ reporter gene. Many specific and informative expression patterns were noted. Expression was most commonly observed in the testis and brain and was most restricted in white adipose tissue and mammary gland. Over half of the assessed genes presented with an absent or localised expression pattern (categorised as 0-10 positive structures). A link between complexity of expression profile and viability of homozygous null animals was observed; inactivation of genes expressed in ≥ 21 structures was more likely to result in reduced viability by postnatal day 14 compared with more restricted expression profiles. For validation purposes, this mouse expression resource was compared with Bgee, a federated composite of RNA-based expression data sets. Strong agreement was observed, indicating a high degree of specificity in our data. Furthermore, there were 1207 observations of expression of a particular gene in an anatomical structure where Bgee had no data, indicating a large amount of novelty in our data set. Examples of expression data corroborating and extending genotype-phenotype associations and supporting disease gene candidacy are presented to demonstrate the potential of this powerful resource.
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Perfilação da Expressão Gênica/métodos , Genes Reporter , Sequenciamento de Nucleotídeos em Larga Escala , Óperon Lac , Fatores Etários , Animais , Biologia Computacional , Bases de Dados Genéticas , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Estudo de Associação Genômica Ampla , Homozigoto , Masculino , Camundongos Knockout , Mutação , Especificidade de Órgãos , FenótipoRESUMO
Limb-girdle muscular dystrophy type 1D (LGMD1D) is caused by dominantly inherited missense mutations in DNAJB6, an Hsp40 co-chaperone. LGMD1D muscle has rimmed vacuoles and inclusion bodies containing DNAJB6, Z-disc proteins and TDP-43. DNAJB6 is expressed as two isoforms; DNAJB6a and DNAJB6b. Both isoforms contain LGMD1D mutant residues and are expressed in human muscle. To identify which mutant isoform confers disease pathogenesis and generate a mouse model of LGMD1D, we evaluated DNAJB6 expression and localization in skeletal muscle as well as generating DNAJB6 isoform specific expressing transgenic mice. DNAJB6a localized to myonuclei while DNAJB6b was sarcoplasmic. LGMD1D mutations in DNAJB6a or DNAJB6b did not alter this localization in mouse muscle. Transgenic mice expressing the LGMD1D mutant, F93L, in DNAJB6b under a muscle-specific promoter became weak, had early lethality and developed muscle pathology consistent with myopathy after 2 months; whereas mice expressing the same F93L mutation in DNAJB6a or overexpressing DNAJB6a or DNAJB6b wild-type transgenes remained unaffected after 1 year. DNAJB6b localized to the Z-disc and DNAJB6b-F93L expressing mouse muscle had myofibrillar disorganization and desmin inclusions. Consistent with DNAJB6 dysfunction, keratin 8/18, a DNAJB6 client also accumulated in DNAJB6b-F93L expressing mouse muscle. The RNA-binding proteins hnRNPA1 and hnRNPA2/B1 accumulated and co-localized with DNAJB6 at sarcoplasmic stress granules suggesting that these proteins maybe novel DNAJB6b clients. Similarly, hnRNPA1 and hnRNPA2/B1 formed sarcoplasmic aggregates in patients with LGMD1D. Our data support that LGMD1D mutations in DNAJB6 disrupt its sarcoplasmic function suggesting a role for DNAJB6b in Z-disc organization and stress granule kinetics.
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Proteínas de Choque Térmico HSP40/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas Grupo A-B/metabolismo , Chaperonas Moleculares/metabolismo , Distrofia Muscular do Cíngulo dos Membros/metabolismo , Miofibrilas/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Animais , Modelos Animais de Doenças , Proteínas de Choque Térmico HSP40/genética , Humanos , Queratinas/metabolismo , Camundongos , Camundongos Transgênicos , Chaperonas Moleculares/genética , Distrofia Muscular do Cíngulo dos Membros/patologia , Mutação , Miofibrilas/patologia , Proteínas do Tecido Nervoso/genética , Regiões Promotoras Genéticas , Isoformas de Proteínas/metabolismoRESUMO
The pacemaking activity of specialized tissues in the heart and gut results in lifelong rhythmic contractions. Here we describe a new syndrome characterized by Chronic Atrial and Intestinal Dysrhythmia, termed CAID syndrome, in 16 French Canadians and 1 Swede. We show that a single shared homozygous founder mutation in SGOL1, a component of the cohesin complex, causes CAID syndrome. Cultured dermal fibroblasts from affected individuals showed accelerated cell cycle progression, a higher rate of senescence and enhanced activation of TGF-ß signaling. Karyotypes showed the typical railroad appearance of a centromeric cohesion defect. Tissues derived from affected individuals displayed pathological changes in both the enteric nervous system and smooth muscle. Morpholino-induced knockdown of sgol1 in zebrafish recapitulated the abnormalities seen in humans with CAID syndrome. Our findings identify CAID syndrome as a novel generalized dysrhythmia, suggesting a new role for SGOL1 and the cohesin complex in mediating the integrity of human cardiac and gut rhythm.
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Anormalidades Múltiplas/genética , Arritmias Cardíacas/genética , Proteínas de Ciclo Celular/genética , Proteínas Cromossômicas não Histona/genética , Enteropatias/genética , Contração Muscular/fisiologia , Transdução de Sinais/genética , Animais , Arritmias Cardíacas/patologia , Ciclo Celular/genética , Sistema Nervoso Entérico/patologia , Fibroblastos , Efeito Fundador , Trato Gastrointestinal/fisiopatologia , Técnicas de Silenciamento de Genes , Humanos , Enteropatias/fisiopatologia , Cariotipagem , Contração Muscular/genética , Músculo Liso Vascular/patologia , Mutação/genética , Quebeque , Síndrome , Fator de Crescimento Transformador beta/metabolismo , Peixe-Zebra , CoesinasRESUMO
The Mouse Genetics Project (MGP) at the Wellcome Trust Sanger Institute aims to generate and phenotype over 800 genetically modified mouse lines over the next 5 years to gain a better understanding of mammalian gene function and provide an invaluable resource to the scientific community for follow-up studies. Phenotyping includes the generation of a standardized biobank of paraffin-embedded tissues for each mouse line, but histopathology is not routinely performed. In collaboration with the Pathology Core of the Centre for Modeling Human Disease (CMHD) we report the utility of histopathology in a high-throughput primary phenotyping screen. Histopathology was assessed in an unbiased selection of 50 mouse lines with (n=30) or without (n=20) clinical phenotypes detected by the standard MGP primary phenotyping screen. Our findings revealed that histopathology added correlating morphological data in 19 of 30 lines (63.3%) in which the primary screen detected a phenotype. In addition, seven of the 50 lines (14%) presented significant histopathology findings that were not associated with or predicted by the standard primary screen. Three of these seven lines had no clinical phenotype detected by the standard primary screen. Incidental and strain-associated background lesions were present in all mutant lines with good concordance to wild-type controls. These findings demonstrate the complementary and unique contribution of histopathology to high-throughput primary phenotyping of mutant mice.
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Ensaios de Triagem em Larga Escala/métodos , Patologia , Fenótipo , Alelos , Animais , Feminino , Masculino , Camundongos , Camundongos Knockout , Especificidade de ÓrgãosRESUMO
Alterations in mitochondrial dynamics (fission, fusion, and movement) are implicated in many neurodegenerative diseases, from rare genetic disorders such as Charcot-Marie-Tooth disease, to common conditions including Alzheimer's disease. However, the relationship between altered mitochondrial dynamics and neurodegeneration is incompletely understood. Here we show that disease associated MFN2 proteins suppressed both mitochondrial fusion and transport, and produced classic features of segmental axonal degeneration without cell body death, including neurofilament filled swellings, loss of calcium homeostasis, and accumulation of reactive oxygen species. By contrast, depletion of Opa1 suppressed mitochondrial fusion while sparing transport, and did not induce axonal degeneration. Axon degeneration induced by mutant MFN2 proteins correlated with the disruption of the proper mitochondrial positioning within axons, rather than loss of overall mitochondrial movement, or global mitochondrial dysfunction. We also found that augmenting expression of MFN1 rescued the axonal degeneration caused by MFN2 mutants, suggesting a possible therapeutic strategy for Charcot-Marie-Tooth disease. These experiments provide evidence that the ability of mitochondria to sense energy requirements and localize properly within axons is key to maintaining axonal integrity, and may be a common pathway by which disruptions in axonal transport contribute to neurodegeneration.
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Axônios/ultraestrutura , GTP Fosfo-Hidrolases/genética , Mitocôndrias/patologia , Proteínas Mitocondriais/genética , Degeneração Neural/genética , Neurônios/citologia , Mutação Puntual/genética , Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Animais , Arginina/genética , Cálcio/metabolismo , Desoxiglucose/metabolismo , Embrião de Mamíferos , Fluoresceínas , GTP Fosfo-Hidrolases/metabolismo , Gânglios Espinais/citologia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Glutamina/genética , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Humanos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/ultraestrutura , Degeneração Neural/patologia , Degeneração Neural/prevenção & controle , Proteínas do Tecido Nervoso/metabolismo , Neurônios/ultraestrutura , RNA Interferente Pequeno/farmacologia , Ratos , Espécies Reativas de Oxigênio/metabolismo , Bloqueadores dos Canais de Sódio/uso terapêutico , Tetrodotoxina/uso terapêutico , TransfecçãoRESUMO
Successful axonal repair following injury is critical for nerve regeneration and functional recovery. Nerve repair relies on three functionally distinct events involving membrane trafficking. First, axonally transported vesicles accumulate, while others are generated at the cut end to restore a selective barrier to the severed axon. Then, retrograde transport of vesicles along microtubules informs the cell body that damage has occurred in the distal axon. Finally, membrane addition to a newly formed growth cone, or to the axonal membrane is required to promote axonal re-growth and elongation. Yet, how these membrane trafficking events are regulated and what are the identities of the molecules and organelles involved remains largely unknown. Several potential factors have been recently identified. Members of the SNARE machinery appear to regulate fusion of vesicles in a calcium-dependent manner to promote axolemmal resealing. Retrograde transport of endosomes powered by the dynein-dynactin molecular motor complex represents a potential injury-signaling platform. Several classes of secretory and endocytic vesicles may coordinate axonal membrane extension and re-growth. Here we discuss recent advances in understanding the mechanisms of the membrane trafficking involved in nerve repair.
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Many night-flying insects evolved ultrasound sensitive ears in response to acoustic predation by echolocating bats . Noctuid moths are most sensitive to frequencies at 20-40 kHz , the lower range of bat ultrasound . This may disadvantage the moth because noctuid-hunting bats in particular echolocate at higher frequencies shortly before prey capture and thus improve their echolocation and reduce their acoustic conspicuousness . Yet, moth hearing is not simple; the ear's nonlinear dynamic response shifts its mechanical sensitivity up to high frequencies. Dependent on incident sound intensity, the moth's ear mechanically tunes up and anticipates the high frequencies used by hunting bats. Surprisingly, this tuning is hysteretic, keeping the ear tuned up for the bat's possible return. A mathematical model is constructed for predicting a linear relationship between the ear's mechanical stiffness and sound intensity. This nonlinear mechanical response is a parametric amplitude dependence that may constitute a feature common to other sensory systems. Adding another twist to the coevolutionary arms race between moths and bats, these results reveal unexpected sophistication in one of the simplest ears known and a novel perspective for interpreting bat echolocation calls.