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Found 67 matches for Mommersteeg
Tbx18 and the fate of epicardial progenitors.
Uncovering the origins of myocardial cells is important for understanding and treating heart diseases. Cai et al. suggest that Tbx18-expressing epicardium provides a substantial contribution to myocytes in the ventricular septum and the atrial and ventricular walls. Here we show that the T-box transcription factor gene 18 (Tbx18) itself is expressed in the myocardium, showing that their genetic lineage tracing system does not allow conclusions of an epicardial origin of cardiomyocytes in vivo to be drawn.
Molecular pathway for the localized formation of the sinoatrial node.
The sinoatrial node, which resides at the junction of the right atrium and the superior caval vein, contains specialized myocardial cells that initiate the heart beat. Despite this fundamental role in heart function, the embryonic origin and mechanisms of localized formation of the sinoatrial node have not been defined. Here we show that subsequent to the formation of the Nkx2-5-positive heart tube, cells bordering the inflow tract of the heart tube give rise to the Nkx2-5-negative myocardial cells of the sinoatrial node and the sinus horns. Using genetic models, we show that as the myocardium of the heart tube matures, Nkx2-5 suppresses pacemaker channel gene Hcn4 and T-box transcription factor gene Tbx3, thereby enforcing a progressive confinement of their expression to the forming Nkx2-5-negative sinoatrial node and sinus horns. Thus, Nkx2-5 is essential for establishing a gene expression border between the atrium and sinoatrial node. Tbx3 was found to suppress chamber differentiation, providing an additional mechanism by which the Tbx3-positive sinoatrial node is shielded from differentiating into atrial myocardium. Pitx2c-deficient fetuses form sinoatrial nodes with indistinguishable molecular signatures at both the right and left sinuatrial junction, indicating that Pitx2c functions within the left/right pathway to suppress a default program for sinuatrial node formation on the left. Our molecular pathway provides a mechanism for how pacemaker activity becomes progressively relegated to the most recently added components of the venous pole of the heart and, ultimately, to the junction of the right atrium and superior caval vein.
Formation of the sinus node head and differentiation of sinus node myocardium are independently regulated by Tbx18 and Tbx3.
The sinus node (or sinoatrial node [SAN]), the pacemaker of the heart, is a functionally and structurally heterogeneous tissue, which consists of a large "head" within the right caval vein myocardium and a "tail" along the terminal crest. Here, we investigated its cellular origin and mechanism of formation. Using genetic lineage analysis and explant assays, we identified T-box transcription factor Tbx18-expressing mesenchymal progenitors in the inflow tract region that differentiate into pacemaker myocardium to form the SAN. We found that the head and tail represent separate regulatory domains expressing distinctive gene programs. Tbx18 is required to establish the large head structure, as seen by the existence of a very small but still functional tail piece in Tbx18-deficient fetuses. In contrast, Tbx3-deficient embryos formed a morphologically normal SAN, which, however, aberrantly expressed Cx40 and other atrial genes, demonstrating that Tbx3 controls differentiation of SAN head and tail cardiomyocytes but also demonstrating that Tbx3 is not required for the formation of the SAN structure. Our data establish a functional order for Tbx18 and Tbx3 in SAN formation, in which Tbx18 controls the formation of the SAN head from mesenchymal precursors, on which Tbx3 subsequently imposes the pacemaker gene program.
Transcription factor Tbx3 is required for the specification of the atrioventricular conduction system.
The cardiac conduction system consists of distinctive heart muscle cells that initiate and propagate the electric impulse required for coordinated contraction. The conduction system expresses the transcriptional repressor Tbx3, which is required for vertebrate development and controls the formation of the sinus node. In humans, mutations in Tbx3 cause ulnar-mammary syndrome. Here, we investigated the role of Tbx3 in the molecular specification of the atrioventricular conduction system. Expression analysis revealed early delineation of the atrioventricular bundle and proximal bundle branches by Tbx3 expression in human, mouse, and chicken. Tbx3-deficient mice, which die between embryonic day 12.5 and 15.5, ectopically expressed genes for connexin (Cx)43, atrial natriuretic factor (Nppa), Tbx18, and Tbx20 in the atrioventricular bundle and proximal bundle branches. Cx40 was precociously upregulated in the atrioventricular bundle of Tbx3 mutants. Moreover, the atrioventricular bundle and branches failed to exit the cell cycle in Tbx3 mutant embryos. Finally, Tbx3-deficient embryos developed outflow tract malformations and ventricular septal defects. These data reveal that Tbx3 is required for the molecular specification of the atrioventricular bundle and bundle branches and for the development of the ventricular septum and outflow tract. Our data suggest a mechanism in which Tbx3 represses differentiation into ventricular working myocardium, thereby imposing the conduction system phenotype on cells within its expression domain.
The sinus venosus progenitors separate and diversify from the first and second heart fields early in development.
AIMS: During development, the heart tube grows by differentiation of Isl1(+)/Nkx2-5(+) progenitors to the arterial and venous pole and dorsal mesocardium. However, after the establishment of the heart tube, Tbx18(+) progenitors were proposed to form the Tbx18(+)/Nkx2-5(-) sinus venosus and proepicardium. To elucidate the relationship between these contributions, we investigated the origin of the Tbx18(+) sinus venosus progenitor population in the cardiogenic mesoderm and its spatial and temporal relation to the second heart field during murine heart development. METHODS AND RESULTS: Explant culture revealed that the Tbx18(+) cell population has the potential to form Nkx2-5(-) sinus venosus myocardium. Three-dimensional reconstruction of expression patterns showed that during heart tube elongation, the Tbx18(+) progenitors remained spatially and temporally separate from the Isl1(+) second heart field, only overlapping with the Isl1(+) domain at the right lateral side of the inflow tract, where the sinus node developed. Consistently, genetic lineage analysis revealed that the Tbx18(+) descendants formed the sinus venosus myocardium, but did not contribute to the pulmonary vein myocardium that developed in the Isl1(+) second heart field. By means of DiI labelling and expression analysis, the origin of the sinus venosus progenitor population was traced to the lateral rim of splanchnic mesoderm that down-regulated Nkx2-5 expression approximately 2 days before its differentiation into sinus venosus myocardium. CONCLUSION: Our data indicate that the cardiogenic mesoderm contains an additional progenitor subpopulation that contributes to the sinus venosus myocardium. After patterning of the cardiogenic mesoderm, this progenitor population remains spatially separated and genetically distinctive from the second heart field subpopulation.
Slit-roundabout signaling regulates the development of the cardiac systemic venous return and pericardium.
RATIONALE: The Slit-Roundabout (Robo) signaling pathway has pleiotropic functions during Drosophila heart development. However, its role in mammalian heart development is largely unknown. OBJECTIVE: To analyze the role of Slit-Robo signaling in the formation of the pericardium and the systemic venous return in the murine heart. METHODS AND RESULTS: Expression of genes encoding Robo1 and Robo2 receptors and their ligands Slit2 and Slit3 was found in or around the systemic venous return and pericardium during development. Analysis of embryos lacking Robo1 revealed partial absence of the pericardium, whereas Robo1/2 double mutants additionally showed severely reduced sinus horn myocardium, hypoplastic caval veins, and a persistent left inferior caval vein. Mice lacking Slit3 recapitulated the defects in the myocardialization, alignment, and morphology of the caval veins. Ligand binding assays confirmed Slit3 as the preferred ligand for the Robo1 receptor, whereas Slit2 showed preference for Robo2. Sinus node development was mostly unaffected in all mutants. In addition, we show absence of cross-regulation with previously identified regulators Tbx18 and Wt1. We provide evidence that pericardial defects are created by abnormal localization of the caval veins combined with ectopic pericardial cavity formation. Local increase in neural crest cell death and impaired neural crest adhesive and migratory properties underlie the ectopic pericardium formation. CONCLUSIONS: A novel Slit-Robo signaling pathway is involved in the development of the pericardium, the sinus horn myocardium, and the alignment of the caval veins. Reduced Slit3 binding in the absence of Robo1, causing impaired cardiac neural crest survival, adhesion, and migration, underlies the pericardial defects.
Disrupted Slit-Robo signalling results in membranous ventricular septum defects and bicuspid aortic valves.
AIMS: The mesenchymal cushions lining the early embryonic heart undergo complex remodelling to form the membranous ventricular septum as well as the atrioventricular and semilunar valves in later life. Disruption of this process underlies the most common congenital heart defects. Here, we identified a novel role for Slit-Robo signalling in the development of the murine membranous ventricular septum and cardiac valves. METHODS AND RESULTS: Expression of Robo1 and Robo2 receptors and their ligands, Slit2 and Slit3, was present in or adjacent to all cardiac cushions/valves. Loss of Robo1 or both Robo1 and Robo2 resulted in membranous ventricular septum defects at birth, a defect also found in Slit3, but not in Slit2 mutants. Additionally, Robo1;Robo2 double mutants showed thickened immature semilunar and atrioventricular valves as well as highly penetrant bicuspid aortic valves. Slit2 mutants recapitulated the semilunar phenotype, whereas Slit3 mutants displayed thickened atrioventricular valves. Bicuspid aortic cushions were already observed at E12.5 in the Robo1;Robo2 double mutants. Expression of Notch- and downstream Hey and Hes genes was down-regulated in Robo1 mutants, suggesting that reduced Notch signalling in mice lacking Robo might underlie the defects. Luciferase assays confirmed regulation of Notch signalling by Robo. CONCLUSION: Cardiac defects in mutants for Robo or Slit range from membranous ventricular septum defects to bicuspid aortic valves. These ligands and receptors have unique functions during development of specific cardiac cushion derivatives, and the Slit-Robo signalling pathway likely enforces its role by regulating Notch signalling, making these mutants a valuable new model to study cardiac valve formation.
Heart Regeneration in the Mexican Cavefish.
Although Astyanax mexicanus surface fish regenerate their hearts after injury, their Pachón cave-dwelling counterparts cannot and, instead, form a permanent fibrotic scar, similar to the human heart. Myocardial proliferation peaks at similar levels in both surface fish and Pachón 1 week after injury. However, in Pachón, this peak coincides with a strong scarring and immune response, and ultimately, cavefish cardiomyocytes fail to replace the scar. We identified lrrc10 to be upregulated in surface fish compared with Pachón after injury. Similar to cavefish, knockout of lrrc10 in zebrafish impairs heart regeneration without affecting wound cardiomyocyte proliferation. Furthermore, using quantitative trait locus (QTL) analysis, we have linked the degree of heart regeneration to three loci in the genome, identifying candidate genes fundamental to the difference between scarring and regeneration. Our study provides evidence that successful heart regeneration entails a delicate interplay between cardiomyocyte proliferation and scarring.
T-box transcription factor 3 governs a transcriptional program for the function of the mouse atrioventricular conduction system.
Genome-wide association studies have identified noncoding variants near TBX3 that are associated with PR interval and QRS duration, suggesting that subtle changes in TBX3 expression affect atrioventricular conduction system function. To explore whether and to what extent the atrioventricular conduction system is affected by Tbx3 dose reduction, we first characterized electrophysiological properties and morphology of heterozygous Tbx3 mutant (Tbx3+/-) mouse hearts. We found PR interval shortening and prolonged QRS duration, as well as atrioventricular bundle hypoplasia after birth in heterozygous mice. The atrioventricular node size was unaffected. Transcriptomic analysis of atrioventricular nodes isolated by laser capture microdissection revealed hundreds of deregulated genes in Tbx3 +/- mutants. Notably, Tbx3 +/- atrioventricular nodes showed increased expression of working myocardial gene programs (mitochondrial and metabolic processes, muscle contractility) and reduced expression of pacemaker gene programs (neuronal, Wnt signaling, calcium/ion channel activity). By integrating chromatin accessibility profiles (ATAC sequencing) of atrioventricular tissue and other epigenetic data, we identified Tbx3-dependent atrioventricular regulatory DNA elements (REs) on a genome-wide scale. We used transgenic reporter assays to determine the functionality of candidate REs near Ryr2, an up-regulated chamber-enriched gene, and in Cacna1g, a down-regulated conduction system-specific gene. Using genome editing to delete candidate REs, we showed that a strong intronic bipartite RE selectively governs Cacna1g expression in the conduction system in vivo. Our data provide insights into the multifactorial Tbx3-dependent transcriptional network that regulates the structure and function of the cardiac conduction system, which may underlie the differences in PR duration and QRS interval between individuals carrying variants in the TBX3 locus.
Runx1 promotes scar deposition and inhibits myocardial proliferation and survival during zebrafish heart regeneration.
Runx1 is a transcription factor that plays a key role in determining the proliferative and differential state of multiple cell types, during both development and adulthood. Here, we report how Runx1 is specifically upregulated at the injury site during zebrafish heart regeneration, and that absence of runx1 results in increased myocardial survival and proliferation, and overall heart regeneration, accompanied by decreased fibrosis. Using single cell sequencing, we found that the wild-type injury site consists of Runx1-positive endocardial cells and thrombocytes that induce expression of smooth muscle and collagen genes. Both these populations cannot be identified in runx1 mutant wounds that contain less collagen and fibrin. The reduction in fibrin in the mutant is further explained by reduced myofibroblast formation and upregulation of components of the fibrin degradation pathway, including plasminogen receptor annexin 2A as well as downregulation of plasminogen activator inhibitor serpine1 in myocardium and endocardium, resulting in increased levels of plasminogen. Our findings suggest that Runx1 controls the regenerative response of multiple cardiac cell types and that targeting Runx1 is a novel therapeutic strategy for inducing endogenous heart repair.
Extraordinary model systems for regeneration
ABSTRACT Regeneration is the remarkable phenomenon through which an organism can regrow lost or damaged parts with fully functional replacements, including complex anatomical structures, such as limbs. In 2019, Development launched its ‘Model systems for regeneration’ collection, a series of articles introducing some of the most popular model organisms for studying regeneration in vivo. To expand this topic further, this Perspective conveys the voices of five expert biologists from the field of regenerative biology, each of whom showcases some less well-known, but equally extraordinary, species for studying regeneration.
Unlocking the Secrets of the Regenerating Fish Heart: Comparing Regenerative Models to Shed Light on Successful Regeneration.
The adult human heart cannot repair itself after injury and, instead, forms a permanent fibrotic scar that impairs cardiac function and can lead to incurable heart failure. The zebrafish, amongst other organisms, has been extensively studied for its innate capacity to repair its heart after injury. Understanding the signals that govern successful regeneration in models such as the zebrafish will lead to the development of effective therapies that can stimulate endogenous repair in humans. To date, many studies have investigated cardiac regeneration using a reverse genetics candidate gene approach. However, this approach is limited in its ability to unbiasedly identify novel genes and signalling pathways that are essential to successful regeneration. In contrast, drawing comparisons between different models of regeneration enables unbiased screens to be performed, identifying signals that have not previously been linked to regeneration. Here, we will review in detail what has been learnt from the comparative approach, highlighting the techniques used and how these studies have influenced the field. We will also discuss what further comparisons would enhance our knowledge of successful regeneration and scarring. Finally, we focus on the Astyanax mexicanus, an intraspecies comparative fish model that holds great promise for revealing the secrets of the regenerating heart.
Discordant Genome Assemblies Drastically Alter the Interpretation of Single-Cell RNA Sequencing Data Which Can Be Mitigated by a Novel Integration Method
Advances in sequencing and assembly technology have led to the creation of genome assemblies for a wide variety of non-model organisms. The rapid production and proliferation of updated, novel assembly versions can create vexing problems for researchers when multiple-genome assembly versions are available at once, requiring researchers to work with more than one reference genome. Multiple-genome assemblies are especially problematic for researchers studying the genetic makeup of individual cells, as single-cell RNA sequencing (scRNAseq) requires sequenced reads to be mapped and aligned to a single reference genome. Using the Astyanax mexicanus, this study highlights how the interpretation of a single-cell dataset from the same sample changes when aligned to its two different available genome assemblies. We found that the number of cells and expressed genes detected were drastically different when aligning to the different assemblies. When the genome assemblies were used in isolation with their respective annotations, cell-type identification was confounded, as some classic cell-type markers were assembly-specific, whilst other genes showed differential patterns of expression between the two assemblies. To overcome the problems posed by multiple-genome assemblies, we propose that researchers align to each available assembly and then integrate the resultant datasets to produce a final dataset in which all genome alignments can be used simultaneously. We found that this approach increased the accuracy of cell-type identification and maximised the amount of data that could be extracted from our single-cell sample by capturing all possible cells and transcripts. As scRNAseq becomes more widely available, it is imperative that the single-cell community is aware of how genome assembly alignment can alter single-cell data and their interpretation, especially when reviewing studies on non-model organisms.
Tissue-Specific Roles for the Slit-Robo Pathway During Heart, Caval Vein, and Diaphragm Development.
Background Binding of Slit ligands to their Robo receptors regulates signaling pathways that are important for heart development. Genetic variants in ROBO1and ROBO4 have been linked to congenital heart defects in humans. These defects are recapitulated in mouse models with ubiquitous deletions of the Slit ligands or Robo receptors and include additional heart defects not currently linked to SLIT or ROBO mutations in humans. Given the broad expression patterns of these genes, the question remains open which tissue-specific ligand-receptor interactions are important for the correct development of different cardiac structures. Methods and Results We used tissue-specific knockout mouse models of Robo1/Robo2, Robo4, Slit2 andSlit3 and scored cardiac developmental defects in perinatal mice. Knockout of Robo2 in either the whole heart, endocardium and its derivatives, or the neural crest in ubiquitous Robo1 knockout background resulted in ventricular septal defects. Neural crest-specific removal of Robo2 in Robo1 knockouts showed fully penetrant bicuspid aortic valves (BAV). Endocardial knock-out of either Slit2or Robo4 caused low penetrant BAV. In contrast, endocardial knockout of Slit3 using a newly generated line resulted in fully penetrant BAV, while removal from smooth muscle cells also resulted in BAV. Caval vein and diaphragm defects observed in ubiquitous Slit3 mutants were recapitulated in the tissue-specific knockouts. Conclusions Our data will help understand defects observed in patients with variants in ROBO1 and ROBO4. The results strongly indicate interaction between endocardial Slit3and neural crest Robo2 in the development of BAV, highlighting the need for further studies of this connection.
Loss of function in ROBO1 is associated with tetralogy of Fallot and septal defects.
BACKGROUND: Congenital heart disease (CHD) is a common birth defect affecting approximately 1% of newborns. Great progress has been made in elucidating the genetic aetiology of CHD with advances in genomic technology, which we leveraged in recovering a new pathway affecting heart development in humans previously known to affect heart development in an animal model. METHODS: Four hundred and sixteen individuals from Thailand and the USA diagnosed with CHD and/or congenital diaphragmatic hernia were evaluated with chromosomal microarray and whole exome sequencing. The DECIPHER Consortium and medical literature were searched for additional patients. Murine hearts from ENU-induced mouse mutants and transgenic mice were evaluated using both episcopic confocal histopathology and troponin I stained sections. RESULTS: Loss of function ROBO1 variants were identified in three families; each proband had a ventricular septal defect, and one proband had tetralogy of Fallot. Additionally, a microdeletion in an individual with CHD was found in the medical literature. Mouse models showed perturbation of the Slit-Robo signalling pathway, causing septation and outflow tract defects and craniofacial anomalies. Two probands had variable facial features consistent with the mouse model. CONCLUSION: Our findings identify Slit-Robo as a significant pathway in human heart development and CHD.
Slit-Robo signalling in heart development.
The Slit ligands and their Robo receptors are well-known for their roles during axon guidance in the central nervous system but are still relatively unknown in the cardiac field. However, data from different animal models suggest a broad involvement of the pathway in many aspects of heart development, from cardiac cell migration and alignment, lumen formation, chamber formation, to the formation of the ventricular septum, semilunar and atrioventricular valves, caval veins, and pericardium. Absence of one or more of the genes in the pathway results in defects ranging from bicuspid aortic valves to ventricular septal defects and abnormal venous connections to the heart. Congenital heart defects are the most common congenital malformations found in life new-born babies and progress in methods for large scale human genetic testing has significantly enhanced the identification of new causative genes involved in human congenital heart disease. Recently, loss of function variants in ROBO1 have also been linked to ventricular septal defects and tetralogy of Fallot in patients. Here, we will give an overview of the role of the Slit-Robo signalling pathway in Drosophila, zebrafish, and mouse heart development. The extent of these data warrant further attention on the SLIT-ROBO signalling pathway as a candidate for an array of human congenital heart defects.
The developmental origin of heart size and shape differences in Astyanax mexicanus populations.
Regulation of heart size and shape is one of the least understood processes in developmental biology. We have for the first time analysed the hearts of Astyanax mexicanus and identified several differences in heart morphology between the surface (epigean morph) and cave-dwelling (troglomorph) morphs. Examination of the adult revealed that the troglomorph possesses a smaller heart with a rounder ventricle in comparison to the epigean morph. The size differences identified appear to arise early in development, as early as 24 h post-fertilisation (hpf), while shape differences begin to appear at 2 days post-fertilisation. The heart of the first-generation cross between the cave-dwelling and river-dwelling morph shows uncoupling of different phenotypes observed in the parental populations and indicates that the cardiac differences have become embedded in the genome during evolution. The differences in heart morphology are accompanied by functional changes between the two morphs, with the cave-dwelling morph exhibiting a slower heart rate than the river-dwelling morph. The identification of morphological and functional differences in the A. mexicanus heart could allow us to gain more insight into how such parameters are regulated during cardiac development, with potential relevance to cardiac pathologies in humans.