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1.
J Anat ; 2024 May 23.
Artículo en Inglés | MEDLINE | ID: mdl-38783643

RESUMEN

Much has been learned over the last half century regarding the molecular and genetic changes that take place during cardiac development. As yet, however, these advances have not been translated into knowledge regarding the marked changes that take place in the anatomical arrangements of the different cardiac components. As such, therefore, many aspects of cardiac development are still described on the basis of speculation rather than evidence. In this review, we show how controversial aspects of development can readily be arbitrated by the interested spectator by taking advantage of the material now gathered together in the Human Developmental Biology Resource; HDBR. We use the material to demonstrate the changes taking place during the formation of the ventricular loop, the expansion of the atrioventricular canal, the incorporation of the systemic venous sinus, the formation of the pulmonary vein, the process of atrial septation, the remodelling of the pharyngeal arches, the major changes occurring during formation of the outflow tract, the closure of the embryonic interventricular communication, and the formation of the ventricular walls. We suggest that access to the resource makes it possible for the interested observer to arbitrate, for themselves, the ongoing controversies that continue to plague the understanding of cardiac development.

2.
J Anat ; 244(3): 497-513, 2024 03.
Artículo en Inglés | MEDLINE | ID: mdl-37957890

RESUMEN

The separation of the outflow tract of the developing heart into the systemic and pulmonary arterial channels remains controversial and poorly understood. The definitive outflow tracts have three components. The developing outflow tract, in contrast, has usually been described in two parts. When the tract has exclusively myocardial walls, such bipartite description is justified, with an obvious dogleg bend separating proximal and distal components. With the addition of non-myocardial walls distally, it becomes possible to recognise three parts. The middle part, which initially still has myocardial walls, contains within its lumen a pair of intercalated valvar swellings. The swellings interdigitate with the distal ends of major outflow cushions, formed by the remodelling of cardiac jelly, to form the primordiums of the arterial roots. The proximal parts of the major cushions, occupying the proximal part of the outflow tract, which also has myocardial walls, themselves fuse and muscularise. The myocardial shelf thus formed remodels to become the free-standing subpulmonary infundibulum. Details of all these processes are currently lacking. In this account, we describe the anatomical changes seen during the overall remodelling. Our interpretations are based on the interrogation of serially sectioned histological and high-resolution episcopic microscopy datasets prepared from developing human and mouse embryos, with some of the datasets processed and reconstructed to reveal the specific nature of the tissues contributing to the separation of the outflow channels. Our findings confirm that the tripartite postnatal arrangement can be correlated with the changes occurring during development.


Asunto(s)
Estructuras Embrionarias , Matriz Extracelular , Cardiopatías Congénitas , Corazón , Ratones , Animales , Humanos , Ventrículos Cardíacos , Arteria Pulmonar
3.
PLoS Genet ; 19(11): e1010777, 2023 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-38011284

RESUMEN

Abnormalities of the arterial valves, including bicuspid aortic valve (BAV) are amongst the most common congenital defects and are a significant cause of morbidity as well as predisposition to disease in later life. Despite this, and compounded by their small size and relative inaccessibility, there is still much to understand about how the arterial valves form and remodel during embryogenesis, both at the morphological and genetic level. Here we set out to address this in human embryos, using Spatial Transcriptomics (ST). We show that ST can be used to investigate the transcriptome of the developing arterial valves, circumventing the problems of accurately dissecting out these tiny structures from the developing embryo. We show that the transcriptome of CS16 and CS19 arterial valves overlap considerably, despite being several days apart in terms of human gestation, and that expression data confirm that the great majority of the most differentially expressed genes are valve-specific. Moreover, we show that the transcriptome of the human arterial valves overlaps with that of mouse atrioventricular valves from a range of gestations, validating our dataset but also highlighting novel genes, including four that are not found in the mouse genome and have not previously been linked to valve development. Importantly, our data suggests that valve transcriptomes are under-represented when using commonly used databases to filter for genes important in cardiac development; this means that causative variants in valve-related genes may be excluded during filtering for genomic data analyses for, for example, BAV. Finally, we highlight "novel" pathways that likely play important roles in arterial valve development, showing that mouse knockouts of RBP1 have arterial valve defects. Thus, this study has confirmed the utility of ST for studies of the developing heart valves and broadens our knowledge of the genes and signalling pathways important in human valve development.


Asunto(s)
Enfermedad de la Válvula Aórtica Bicúspide , Enfermedades de las Válvulas Cardíacas , Humanos , Ratones , Animales , Enfermedades de las Válvulas Cardíacas/genética , Válvula Aórtica/anomalías , Enfermedad de la Válvula Aórtica Bicúspide/metabolismo , Perfilación de la Expresión Génica , Transcriptoma/genética
4.
Nature ; 614(7948): 509-520, 2023 02.
Artículo en Inglés | MEDLINE | ID: mdl-36543322

RESUMEN

The segmented body plan of vertebrates is established during somitogenesis, a well-studied process in model organisms; however, the details of this process in humans remain largely unknown owing to ethical and technical limitations. Despite recent advances with pluripotent stem cell-based approaches1-5, models that robustly recapitulate human somitogenesis in both space and time remain scarce. Here we introduce a pluripotent stem cell-derived mesoderm-based 3D model of human segmentation and somitogenesis-which we termed 'axioloid'-that captures accurately the oscillatory dynamics of the segmentation clock and the morphological and molecular characteristics of sequential somite formation in vitro. Axioloids show proper rostrocaudal patterning of forming segments and robust anterior-posterior FGF-WNT signalling gradients and retinoic acid signalling components. We identify an unexpected critical role of retinoic acid signalling in the stabilization of forming segments, indicating distinct, but also synergistic effects of retinoic acid and extracellular matrix on the formation and epithelialization of somites. Comparative analysis demonstrates marked similarities of axioloids to the human embryo, further validated by the presence of a Hox code in axioloids. Finally, we demonstrate the utility of axioloids for studying the pathogenesis of human congenital spine diseases using induced pluripotent stem cells with mutations in HES7 and MESP2. Our results indicate that axioloids represent a promising platform for the study of axial development and disease in humans.


Asunto(s)
Tipificación del Cuerpo , Técnicas de Cultivo Tridimensional de Células , Somitos , Humanos , Tipificación del Cuerpo/efectos de los fármacos , Matriz Extracelular/metabolismo , Factores de Crecimiento de Fibroblastos/metabolismo , Técnicas In Vitro , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/metabolismo , Modelos Biológicos , Mutación , Somitos/citología , Somitos/efectos de los fármacos , Somitos/embriología , Somitos/metabolismo , Enfermedades de la Columna Vertebral/patología , Tretinoina/metabolismo , Tretinoina/farmacología , Vía de Señalización Wnt/efectos de los fármacos
5.
J Anat ; 241(1): 33-41, 2022 07.
Artículo en Inglés | MEDLINE | ID: mdl-35224745

RESUMEN

Several strategies have been recently introduced to improve the practicality of multiple immunolabeling and RNA in situ hybridization protocols. Tyramide signal amplification (TSA) is a powerful method used to improve the detection sensitivity of immunohistochemistry. RNAScope is a novel commercially available in situ hybridization assay for the detection of RNA expression. In this work, we describe the use of TSA and RNAScope in situ hybridization as extremely sensitive and specific methods for the evaluation of protein and RNA expression in formaldehyde-fixed paraffin-embedded human fetal brain sections. These two techniques, when properly optimized, were highly compatible with routine formaldehyde-fixed paraffin-embedded tissue that preserves the best morphological characteristics of delicate fetal brain samples, enabling an unparalleled ability to simultaneously visualize the expression of multiple protein and mRNA of genes that are sparsely expressed in the human fetal telencephalon.


Asunto(s)
Formaldehído , ARN , Encéfalo/metabolismo , Humanos , Hibridación in Situ , Adhesión en Parafina/métodos , ARN/genética
6.
Semin Cell Dev Biol ; 76: 3-14, 2018 04.
Artículo en Inglés | MEDLINE | ID: mdl-28834762

RESUMEN

The cerebral cortex is divided stereotypically into a number of functionally distinct areas. According to the protomap hypothesis formulated by Rakic neural progenitors in the ventricular zone form a mosaic of proliferative units that provide a primordial species-specific cortical map. Positional information of newborn neurons is maintained during their migration to the overlying cortical plate. Much evidence has been found to support this hypothesis from studies of primary cortical areas in mouse models in particular. Differential expansion of cortical areas and the introduction of new functional modules during evolution might be the result of changes in the progenitor cells. The human cerebral cortex shows a wide divergence from the mouse containing a much higher proportion of association cortex and a more complicated regionalised repertoire of neuron sub-types. To what extent does the protomap hypothesis hold true for the primate brain? This review summarises a growing number of studies exploring arealised gene expression in the early developing human telencephalon. The evidence so far is that the human and mouse brain do share fundamental mechanisms of areal specification, however there are subtle differences which could lead us to a better understanding of cortical evolution and the origins of neurodevelopmental diseases.


Asunto(s)
Corteza Cerebral/crecimiento & desarrollo , Neurogénesis/genética , Telencéfalo/crecimiento & desarrollo , Diferenciación Celular , Humanos
7.
Brain Struct Funct ; 222(5): 2309-2328, 2017 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-27905023

RESUMEN

The extent of similarities and differences between cortical GABAergic interneuron generation in rodent and primate telencephalon remains contentious. We examined expression of three interneuron precursor transcription factors, alongside other markers, using immunohistochemistry on 8-12 post-conceptional weeks (PCW) human telencephalon sections. NKX2.1, OLIG2, and COUP-TFII expression occupied distinct (although overlapping) neurogenic domains which extended into the cortex and revealed three CGE compartments: lateral, medial, and ventral. NKX2.1 expression was very largely confined to the MGE, medial CGE, and ventral septum confirming that, at this developmental stage, interneuron generation from NKX2.1+ precursors closely resembles the process observed in rodents. OLIG2 immunoreactivity was observed in GABAergic cells of the proliferative zones of the MGE and septum, but not necessarily co-expressed with NKX2.1, and OLIG2 expression was also extensively seen in the LGE, CGE, and cortex. At 8 PCW, OLIG2+ cells were only present in the medial and anterior cortical wall suggesting a migratory pathway for interneuron precursors via the septum into the medial cortex. By 12 PCW, OLIG2+ cells were present throughout the cortex and many were actively dividing but without co-expressing cortical progenitor markers. Dividing COUP-TFII+ progenitor cells were localized to ventral CGE as previously described but were also numerous in adjacent ventral cortex; in both the cases, COUP-TFII was co-expressed with PAX6 in proliferative zones and TBR1 or calretinin in post-mitotic cortical neurons. Thus COUP-TFII+ progenitors gave rise to pyramidal cells, but also interneurons which not only migrated posteriorly into the cortex from ventral CGE but also anteriorly via the LGE.


Asunto(s)
Factor de Transcripción COUP II/metabolismo , Interneuronas/metabolismo , Factor de Transcripción 2 de los Oligodendrocitos/metabolismo , Telencéfalo/metabolismo , Factor Nuclear Tiroideo 1/metabolismo , Factores de Transcripción/metabolismo , Calbindina 2/metabolismo , Regulación del Desarrollo de la Expresión Génica , Humanos , Inmunohistoquímica , Neurogénesis/fisiología , Telencéfalo/embriología
8.
Sci Rep ; 6: 27714, 2016 Jun 21.
Artículo en Inglés | MEDLINE | ID: mdl-27323879

RESUMEN

Hypertrophic cardiomyopathy (HCM) is caused by mutations in sarcomeric proteins, the commonest being MYBPC3 encoding myosin-binding protein C. It is characterised by left ventricular hypertrophy but there is an important pre-hypertrophic phenotype with features including crypts, abnormal mitral leaflets and trabeculae. We investigated these during mouse cardiac development using high-resolution episcopic microscopy. In embryonic hearts from wildtype, homozygous (HO) and heterozygous (HET) Mybpc3-targeted knock-out (KO) mice we show that crypts (one or two) are a normal part of wildtype development but they almost all resolve by birth. By contrast, HO and HET embryos had increased crypt presence, abnormal mitral valve formation and alterations in the compaction process. In scarce normal human embryos, crypts were sometimes present. This study shows that features of the human pre-hypertrophic HCM phenotype occur in the mouse. In an animal model we demonstrate that there is an embryological HCM phenotype. Crypts are a normal part of cardiac development but, along with the mitral valve and trabeculae, their developmental trajectory is altered by the presence of HCM truncating Mybpc3 gene mutation.


Asunto(s)
Cardiomiopatía Hipertrófica/genética , Proteínas Portadoras/genética , Corazón/crecimiento & desarrollo , Hipertrofia Ventricular Izquierda/genética , Animales , Cardiomiopatía Hipertrófica/embriología , Cardiomiopatía Hipertrófica/patología , Modelos Animales de Enfermedad , Genotipo , Corazón/embriología , Corazón/fisiopatología , Humanos , Hipertrofia Ventricular Izquierda/embriología , Hipertrofia Ventricular Izquierda/patología , Ratones , Válvula Mitral/patología , Mutación
9.
J Anat ; 217(4): 289-99, 2010 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-20979583

RESUMEN

We are developing a three-dimensional (3D) atlas of the human embryonic brain using anatomical landmarks and gene expression data to define major subdivisions through 12 stages of development [Carnegie Stages (CS) 12-23; approximately 26-56 days post conception (dpc)]. Virtual 3D anatomical models are generated from intact specimens using optical projection tomography (OPT). Using MAPAINT software, selected gene expression data, gathered using standard methods of in situ hybridization and immunohistochemistry, are mapped to a representative 3D model for each chosen Carnegie stage. In these models, anatomical domains, defined on the basis of morphological landmarks and comparative knowledge of expression patterns in vertebrates, are linked to a developmental neuroanatomic ontology. Human gene expression patterns for genes with characteristic expression in different vertebrates (e.g. PAX6, GAD65 and OLIG2) are being used to confirm and/or refine the human anatomical domain boundaries. We have also developed interpolation software that digitally generates a full domain from partial data. Currently, the 3D models and a preliminary set of anatomical domains and ontology are available on the atlas pages along with gene expression data from approximately 100 genes in the HUDSEN Human Spatial Gene Expression Database (http://www.hudsen.org). The aim is that full 3D data will be generated from expression data used to define a more detailed set of anatomical domains linked to a more advanced anatomy ontology and all of these will be available online, contributing to the long-term goal of the atlas, which is to help maximize the effective use and dissemination of data wherever it is generated.


Asunto(s)
Encéfalo/embriología , Encéfalo/metabolismo , Bases de Datos Genéticas , Regulación del Desarrollo de la Expresión Génica/fisiología , Expresión Génica/fisiología , Imagenología Tridimensional/métodos , Factores de Transcripción/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Mapeo Encefálico/métodos , Gráficos por Computador , Proteínas del Ojo/genética , Proteínas del Ojo/metabolismo , Perfilación de la Expresión Génica/métodos , Glutamato Descarboxilasa/genética , Glutamato Descarboxilasa/metabolismo , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Humanos , Modelos Neurológicos , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Factor de Transcripción 2 de los Oligodendrocitos , Factor de Transcripción PAX6 , Factores de Transcripción Paired Box/genética , Factores de Transcripción Paired Box/metabolismo , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Tomografía Óptica/métodos
10.
Cell ; 140(1): 74-87, 2010 Jan 08.
Artículo en Inglés | MEDLINE | ID: mdl-20074521

RESUMEN

We report that eight heterozygous missense mutations in TUBB3, encoding the neuron-specific beta-tubulin isotype III, result in a spectrum of human nervous system disorders that we now call the TUBB3 syndromes. Each mutation causes the ocular motility disorder CFEOM3, whereas some also result in intellectual and behavioral impairments, facial paralysis, and/or later-onset axonal sensorimotor polyneuropathy. Neuroimaging reveals a spectrum of abnormalities including hypoplasia of oculomotor nerves and dysgenesis of the corpus callosum, anterior commissure, and corticospinal tracts. A knock-in disease mouse model reveals axon guidance defects without evidence of cortical cell migration abnormalities. We show that the disease-associated mutations can impair tubulin heterodimer formation in vitro, although folded mutant heterodimers can still polymerize into microtubules. Modeling each mutation in yeast tubulin demonstrates that all alter dynamic instability whereas a subset disrupts the interaction of microtubules with kinesin motors. These findings demonstrate that normal TUBB3 is required for axon guidance and maintenance in mammals.


Asunto(s)
Tubulina (Proteína)/metabolismo , Secuencia de Aminoácidos , Animales , Axones/metabolismo , Encéfalo/embriología , Encéfalo/metabolismo , Supervivencia Celular , Niño , Discapacidades del Desarrollo , Femenino , Humanos , Cinesinas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Microtúbulos/metabolismo , Modelos Moleculares , Datos de Secuencia Molecular , Mutación Missense , Transporte de Proteínas , Tubulina (Proteína)/química , Tubulina (Proteína)/genética
11.
Brain Res Bull ; 66(4-6): 449-53, 2005 Sep 15.
Artículo en Inglés | MEDLINE | ID: mdl-16144630

RESUMEN

As human brain development proceeds, there are complex changes in size and shape, most notably in the developing forebrain. Molecular technologies enable us to characterise the gene expression patterns that underlie these changes. To interpret these patterns the location of expression must be identified and, often, gene expression patterns compared for several genes or across several developmental stages. To facilitate interpretation we have generated a set of three-dimensional models using a recently developed technique, optical projection tomography. The models act as a framework onto which gene expression patterns are mapped and anatomical domains identified using custom-designed software, MAPaint. Here, we demonstrate their use to compare forebrain development at two embryonic stages (Carnegie stages 18 and 21; 44 and 52 days post conception, respectively) and as a means of recording, storing and visualising gene expression data for three example genes EMX1, EMX2 and OTX2. Anatomical domains were also mapped to the models and the comparison of gene expression and anatomical data is demonstrated at Carnegie stage 21. The three-dimensional models and sophisticated software facilitate the analysis and visualisation of morphological changes and gene expression patterns during early brain development and can be applied to the development of other complex structures.


Asunto(s)
Mapeo Encefálico , Encéfalo/metabolismo , Mapeo Cromosómico , Regulación del Desarrollo de la Expresión Génica/fisiología , Encéfalo/embriología , Embrión de Mamíferos , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Humanos , Imagenología Tridimensional/métodos , Complejo Mediador , Modelos Anatómicos , Factores de Transcripción Otx/genética , Factores de Transcripción Otx/metabolismo , Transactivadores/genética , Transactivadores/metabolismo , Factores de Transcripción
12.
BMC Bioinformatics ; 6: 47, 2005 Mar 09.
Artículo en Inglés | MEDLINE | ID: mdl-15757508

RESUMEN

BACKGROUND: Many three-dimensional (3D) images are routinely collected in biomedical research and a number of digital atlases with associated anatomical and other information have been published. A number of tools are available for viewing this data ranging from commercial visualization packages to freely available, typically system architecture dependent, solutions. Here we discuss an atlas viewer implemented to run on any workstation using the architecture neutral Java programming language. RESULTS: We report the development of a freely available Java based viewer for 3D image data, descibe the structure and functionality of the viewer and how automated tools can be developed to manage the Java Native Interface code. The viewer allows arbitrary re-sectioning of the data and interactive browsing through the volume. With appropriately formatted data, for example as provided for the Electronic Atlas of the Developing Human Brain, a 3D surface view and anatomical browsing is available. The interface is developed in Java with Java3D providing the 3D rendering. For efficiency the image data is manipulated using the Woolz image-processing library provided as a dynamically linked module for each machine architecture. CONCLUSION: We conclude that Java provides an appropriate environment for efficient development of these tools and techniques exist to allow computationally efficient image-processing libraries to be integrated relatively easily.


Asunto(s)
Biología Computacional/métodos , Bases de Datos de Proteínas , Anatomía , Anatomía Transversal , Animales , Encéfalo/anatomía & histología , Atlas Cervical , Gráficos por Computador , Simulación por Computador , Instrucción por Computador , Computadores , Presentación de Datos , Humanos , Interpretación de Imagen Asistida por Computador , Procesamiento de Imagen Asistido por Computador , Imagenología Tridimensional , Almacenamiento y Recuperación de la Información , Ratones , Modelos Anatómicos , Lenguajes de Programación , Programas Informáticos , Interfaz Usuario-Computador
13.
BMC Neurosci ; 5: 27, 2004 Aug 06.
Artículo en Inglés | MEDLINE | ID: mdl-15298700

RESUMEN

BACKGROUND: As development proceeds the human embryo attains an ever more complex three dimensional (3D) structure. Analyzing the gene expression patterns that underlie these changes and interpreting their significance depends on identifying the anatomical structures to which they map and following these patterns in developing 3D structures over time. The difficulty of this task greatly increases as more gene expression patterns are added, particularly in organs with complex 3D structures such as the brain. Optical Projection Tomography (OPT) is a new technology which has been developed for rapidly generating digital 3D models of intact specimens. We have assessed the resolution of unstained neuronal structures within a Carnegie Stage (CS)17 OPT model and tested its use as a framework onto which anatomical structures can be defined and gene expression data mapped. RESULTS: Resolution of the OPT models was assessed by comparison of digital sections with physical sections stained, either with haematoxylin and eosin (H&E) or by immunocytochemistry for GAP43 or PAX6, to identify specific anatomical features. Despite the 3D models being of unstained tissue, peripheral nervous system structures from the trigeminal ganglion (approximately 300 microm by approximately 150 microm) to the rootlets of cranial nerve XII (approximately 20 microm in diameter) were clearly identifiable, as were structures in the developing neural tube such as the zona limitans intrathalamica (core is approximately 30 microm thick). Fourteen anatomical domains have been identified and visualised within the CS17 model. Two 3D gene expression domains, known to be defined by Pax6 expression in the mouse, were clearly visible when PAX6 data from 2D sections were mapped to the CS17 model. The feasibility of applying the OPT technology to all stages from CS12 to CS23, which encompasses the major period of organogenesis for the human developing central nervous system, was successfully demonstrated. CONCLUSION: In the CS17 model considerable detail is visible within the developing nervous system at a minimum resolution of approximately 20 microm and 3D anatomical and gene expression domains can be defined and visualised successfully. The OPT models and accompanying technologies for manipulating them provide a powerful approach to visualising and analysing gene expression and morphology during early human brain development.


Asunto(s)
Encéfalo/embriología , Imagenología Tridimensional/métodos , Modelos Neurológicos , Tomografía/métodos , Gráficos por Computador , Edad Gestacional , Humanos , Programas Informáticos
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