Richard Harvey

PDGFRα signaling in cardiac fibroblasts modulates quiescence, metabolism and self-renewal, and promotes anatomical and functional repair

Publisher: Cold Spring Harbor Laboratory

Date: 28-11-2017

DOI: 10.1101/225979

Abstract: The interstitial and perivascular spaces of the mammalian heart contain a highly interactive tissue community essential for cardiac homeostasis, repair and regeneration. Mesenchymal cells (fibroblasts) are one of the most abundant cell types, playing key roles as sentinels, tissue architects, paracrine signaling hubs and lineage precursors, and are linked to heart disease through their roles in inflammation and fibrosis. Platelet-derived growth factors (PDGFs) are secreted by several cell types involved in cardiac injury and repair, and are recognized mitogens for cardiac fibroblasts and mesenchymal stem cells. However, their roles are complex and investigations of their impact on heart repair have produced contrasting outcomes, leaving therapeutic potential uncertain. Here, we use new approaches and tools, including single cell RNA sequencing, to explore cardiac fibroblast heterogeneity and how PDGF receptor α (PDGFRα) signaling impacts fibroblasts during heart repair. Short-term systemic delivery of PDGF-AB to mice from the time of myocardial infarction (MI) led to enhanced anatomical and functional recovery. Underpinning these benefits was a priming effect, in which PDGF-AB accelerated exit of fibroblasts from quiescence and induced a higher translational biosynthetic capacity in both fibroblasts and macrophages without triggering fibrosis. Our study highlights the significant biosynthetic heterogeneity and plasticity in cardiac fibroblast populations, and suggests a rationale for a novel therapeutic approach to cardiac injury involving controlled stimulation of fibroblast activation.

Microinjection of synthetic Xhox-1A homeobox mRNA disrupts somite formation in developing Xenopus embryos

Publisher: Elsevier BV

Date: 06-1988

DOI: 10.1016/0092-8674(88)90087-6

Abstract: The structural similarity between Drosophila and vertebrate homeobox genes begs the question of whether the vertebrate gene products affect cell fate and pattern formation. To study the function of the Xenopus homeobox protein, Xhox-1A, we microinjected fertilized Xenopus eggs with an excess of synthetic Xhox-RNA and assayed for effects on development. The predominant phenotype is a disturbance in somite formation. When embryos are injected with Xhox-1A mRNA, but not with control mRNAs, morphogenesis of somites occurs chaotically and in idual segments are lost. Histological staining, in situ hybridization, and immunohistochemistry indicate that the disorganized somitic tissue has differentiated into muscle cells. Overall, these results suggest that correct regulation of the Xhox-1A gene may be important for the normal development of the segmented somite pattern in early embryos. Moreover, the inferred role of Xhox-1A in somite formation indicates that there may be molecular parallels between mechanisms of segmentation in flies and vertebrates.

Point mutations in murine Nkx2-5 phenocopy human congenital heart disease and induce pathogenic Wnt signaling

Publisher: American Society for Clinical Investigation

Date: 23-03-2017

DOI: 10.1172/JCI.INSIGHT.88271

Cardiac outflow tract development relies on the complex function of Sox4 and Sox11 in multiple cell types

Publisher: Springer Science and Business Media LLC

Date: 06-12-2013

DOI: 10.1007/S00018-013-1523-X

Abstract: Congenital heart defects represent the most common human birth defects and are often life-threatening. Frequently, they are caused by abnormalities of the outflow tract whose formation results from coordinated development of cells from mesodermal and neural crest origin and depends on the activity of many different transcription factors. However, place, time, and mode of action have only been analyzed for a few of them. Here we assess the contribution of the closely related high-mobility-group transcription factors Sox4 and Sox11 to outflow tract development and determine their function. Using cell-type-specific deletion in the mouse, we show that Sox11 is required for proper development in both mesodermal cells and neural crest cells. Deletion in either mesoderm or neural crest, or both, leads to outflow tract defects ranging from double outlet right ventricle to common arterial trunk. Sox4 supports Sox11 in its function, but has additional roles with relevance for outflow tract formation in other cell types. The two Sox proteins are dispensable during early phases of cardiac neural crest development including neural tube emigration, proliferation, and migration through the pharyngeal arches. They become essential after arrival of the neural crest cells in the outflow tract for their proper differentiation and interaction with each other as well as with the environment through regulation of cytoskeletal, cell adhesion, and extracellular matrix molecules. Our results demonstrate that Sox4 and Sox11 have multiple functions in several cell types during outflow tract formation and may thus help to understand the basis of congenital heart defects in humans.

Cloning and expression of a cDNA coding for the anticoagulant hirudin from the bloodsucking leech, Hirudo medicinalis.

Publisher: Proceedings of the National Academy of Sciences

Date: 02-1986

DOI: 10.1073/PNAS.83.4.1084

Abstract: Cloned cDNAs have been isolated that encode a variant of hirudin, a potent thrombin inhibitor that is secreted by the salivary glands of the medicinal leech, Hirudo medicinalis. This variant probably corresponds to a form that has been purified from leech heads but differs in amino acid sequence from the hirudin purified from whole leeches. There are at least three hirudin transcripts detectable in leech RNAs that are different in size, site of synthesis, inducibility by starvation, and relationship to hirudin activity. The new hirudin variant predicted by the cDNA and the heterodisperse transcription products suggest a hirudin protein family. The hirudin cDNA was expressed in Escherichia coli under the control of the bacteriophage lambda PL promoter. The recombinant product is biologically active, inhibiting the cleavage by thrombin of fibrinogen and a synthetic tripeptide substrate.

Cardiac fibroblast heterogeneity and dynamics through the lens of single-cell dual ‘omics

Publisher: Oxford University Press (OUP)

Date: 20-04-2022

DOI: 10.1093/CVR/CVAC037

Controlling Endosomal Escape Using pH-Responsive Nanoparticles with Tunable Disassembly

Publisher: American Chemical Society (ACS)

Date: 05-06-2018

DOI: 10.1021/ACSANM.8B00338

Quantitative Trait Loci Modifying Cardiac Atrial Septal Morphology and Risk of Patent Foramen Ovale in the Mouse

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 17-03-2006

DOI: 10.1161/01.RES.0000209965.59312.AA

Abstract: Atrial septal defect (ASD) is a common congenital heart disease (CHD) occurring in 5 to 7 per 10 000 live births. Mutations in 5 human genes ( NKX2.5 , TBX5 , GATA4 , MYHC , ACTC ) are known to cause dominant ASD, but these account for a minority of cases. Human and mouse data suggest that ASD exists in an anatomical continuum with milder septal variants patent foramen ovale (PFO) and atrial septal aneurysm, strongly associated with ischemic stroke and migraine. We have previously shown in inbred mice that the incidence of PFO strongly correlates with length of the interatrial septum primum , defining a quantitative trait underlying PFO risk. To better understand genetic causation of atrial septal abnormalities, we mapped quantitative trait loci (QTL) influencing septal morphology using mouse strains (QSi5 and 129T2/SvEms) maximally informative for PFO incidence and 3 quantitative septal anatomical traits including septum primum length. [QSi5×129T2/SvEms]F2 intercross animals (n=1437) were phenotyped and a whole genome scan performed at an average 17-cM interval. Statistical methodology scoring PFO as a binary phenotype was developed as a confirmatory mapping technique. We mapped 7 significant and 6 suggestive QTL modifying quantitative phenotypes, with 4 supported by binary analysis. Quantitative traits, although strongly associated with PFO ( P .001), correlated poorly with each other and in all but 1 case QTL for different traits were nonoverlapping. Thus, multiple anatomical processes under separate genetic control contribute to risk of PFO. Our findings demonstrate the feasibility of modeling the genetic basis of common CHD using animal genetic and genomic technologies.

Biased DNA Segregation and Cardiac Stem Cell Therapies

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 14-09-2012

DOI: 10.1161/CIRCRESAHA.112.277764

High Resolution Melting Analysis Is a More Sensitive and Effective Alternative to Gel-Based Platforms in Analysis of SSR – An Example in Citrus

Publisher: Public Library of Science (PLoS)

Date: 30-08-2012

DOI: 10.1371/JOURNAL.PONE.0044202

Pressure Overload by Transverse Aortic Constriction Induces Maladaptive Hypertrophy in a Titin-Truncated Mouse Model

Publisher: Hindawi Limited

Date: 2015

DOI: 10.1155/2015/163564

Abstract: Mutations in the giant sarcomeric protein titin (TTN) are a major cause for inherited forms of dilated cardiomyopathy (DCM). We have previously developed a mouse model that imitates a TTN truncation mutation we found in a large pedigree with DCM. While heterozygous Ttn knock-in mice do not display signs of heart failure under sedentary conditions, they recapitulate the human phenotype when exposed to the pharmacological stressor angiotensin II or isoproterenol. In this study we investigated the effects of pressure overload by transverse aortic constriction (TAC) in heterozygous (Het) Ttn knock-in mice. Two weeks after TAC, Het mice developed marked impairment of left ventricular ejection fraction ( p 0.05 ) , while wild-type (WT) TAC mice did not. Het mice also trended toward increased ventricular end diastolic pressure and volume compared to WT littermates. We found an increase in histologically diffuse cardiac fibrosis in Het compared to WT in TAC mice. This study shows that a pattern of DCM can be induced by TAC-mediated pressure overload in a TTN-truncated mouse model. This model enlarges our arsenal of cardiac disease models, adding a valuable tool to understand cardiac pathophysiological remodeling processes and to develop therapeutic approaches to combat heart failure.

Molecular pathways in myocardial development: a stem cell perspective

Publisher: Oxford University Press (OUP)

Date: 05-2003

DOI: 10.1016/S0008-6363(03)00286-4

Abstract: The heart has long been considered to adapt to increased work or pathology through the cellular growth process of hypertrophy. However, recent evidence for the existence of endogenous stem cells and regenerative capacity in the adult heart has given new impetus to the quest for cell therapies for heart failure, which remains the number one killer in Western cultures. The molecular cues driving cardiac development are now being explored in detail and will come into sharp focus as regimes for stem cell differentiation and efforts to augment endogenous regeneration are trialed. This review briefly outlines the current state of knowledge on the molecular basis of the four modalities of myogenesis that have been identified in the developing vertebrate heart. Stem cell-mediated myogenic repair in the heart represents a fifth modality, and an exciting frontier with basic and practical implications that remain to be explored.

Formation of the Venous Pole of the Heart From an Nkx2–5 –Negative Precursor Population Requires Tbx18

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 23-06-2006

DOI: 10.1161/01.RES.0000227571.84189.65

Abstract: The venous pole of the mammalian heart is a structurally and electrically complex region, yet the lineage and molecular mechanisms underlying its formation have remained largely unexplored. In contrast to classical studies that attribute the origin of the myocardial sinus horns to the embryonic venous pole, we find that the sinus horns form only after heart looping by differentiation of mesenchymal cells of the septum transversum region into myocardium. The myocardial sinus horns and their mesenchymal precursor cells never express Nkx2–5 , a transcription factor critical for heart development. In addition, lineage studies show that the sinus horns do not derive from cells previously positive for Nkx2–5 . In contrast, the sinus horns express the T-box transcription factor gene Tbx18 . Mice deficient for Tbx18 fail to form sinus horns from the pericardial mesenchyme and have defective caval veins, whereas the pulmonary vein and atrial structures are unaffected. Our studies define a novel heart precursor population that contributes exclusively to the myocardium surrounding the sinus horns or systemic venous tributaries of the developing heart, which are a source of congenital malformation and cardiac arrhythmias.

Hypoplastic Left Heart Syndrome

Publisher: Elsevier BV

Date: 03-2009

DOI: 10.1016/J.JACC.2008.12.024

NKX2-5 regulates human cardiomyogenesis via a HEY2 dependent transcriptional network

Publisher: Springer Science and Business Media LLC

Date: 10-04-2018

DOI: 10.1038/S41467-018-03714-X

Abstract: Congenital heart defects can be caused by mutations in genes that guide cardiac lineage formation. Here, we show deletion of NKX2-5 , a critical component of the cardiac gene regulatory network, in human embryonic stem cells (hESCs), results in impaired cardiomyogenesis, failure to activate VCAM1 and to downregulate the progenitor marker PDGFRα. Furthermore, NKX2-5 null cardiomyocytes have abnormal physiology, with asynchronous contractions and altered action potentials. Molecular profiling and genetic rescue experiments demonstrate that the bHLH protein HEY2 is a key mediator of NKX2-5 function during human cardiomyogenesis. These findings identify HEY2 as a novel component of the NKX2-5 cardiac transcriptional network, providing tangible evidence that hESC models can decipher the complex pathways that regulate early stage human heart development. These data provide a human context for the evaluation of pathogenic mutations in congenital heart disease.

A RhoA-FRET Biosensor Mouse for Intravital Imaging in Normal Tissue Homeostasis and Disease Contexts

Publisher: Elsevier BV

Date: 10-2017

DOI: 10.1016/J.CELREP.2017.09.022

Abstract: The small GTPase RhoA is involved in a variety of fundamental processes in normal tissue. Spatiotemporal control of RhoA is thought to govern mechanosensing, growth, and motility of cells, while its deregulation is associated with disease development. Here, we describe the generation of a RhoA-fluorescence resonance energy transfer (FRET) biosensor mouse and its utility for monitoring real-time activity of RhoA in a variety of native tissues in vivo. We assess changes in RhoA activity during mechanosensing of osteocytes within the bone and during neutrophil migration. We also demonstrate spatiotemporal order of RhoA activity within crypt cells of the small intestine and during different stages of mammary gestation. Subsequently, we reveal co-option of RhoA activity in both invasive breast and pancreatic cancers, and we assess drug targeting in these disease settings, illustrating the potential for utilizing this mouse to study RhoA activity in vivo in real time.

Cardiomyogenic Precursor Cells in the Mammalian Embryo: Induction, Heterogeneity, and Morphogenesis

Publisher: Elsevier

Date: 2004

DOI: 10.1016/B978-012436643-5/50037-7

NK-2 Class Homeodomain Proteins

Publisher: Elsevier

Date: 2010

DOI: 10.1016/B978-0-12-381332-9.00026-8

Conditional (loxP-flanked) allele for the gene encoding the retinoic acid-synthesizing enzyme retinaldehyde dehydrogenase 2 (RALDH2)

Publisher: Wiley

Date: 2006

DOI: 10.1002/GENE.20195

Abstract: Retinoic acid, the active vitamin A derivative, has pleiotropic functions during vertebrate development and postnatal life. Retinaldehyde dehydrogenase 2 (RALDH2) acts as the main retinoic acid-synthesizing enzyme during development. Mouse Raldh2 germline null mutants are early embryonic lethal and exhibit complex abnormalities that include defective heart looping morphogenesis. To investigate later functions of this enzyme, we have engineered a "floxed" (loxP-flanked) allele allowing Cre-mediated somatic gene inactivations. Mice heterozygous or homozygous for the floxed Raldh2 allele are viable and fertile. We tested whether the novel Raldh2 allele behaves as a null mutation after Cre-mediated in vivo excision by crossing the conditional mutants with CMV-Cre transgenic mice. An embryonic lethal phenotype indistinguishable from that of germline mutants was obtained. The conditional allele described herein is a genetic tool for studying tissue-specific, RALDH2-dependent functions of retinoic acid during development and in adult life.

Author response: Quantitative trait and transcriptome analysis of genetic complexity underpinning cardiac interatrial septation in mice using an advanced intercross line

Publisher: eLife Sciences Publications, Ltd

Date: 05-05-2023

DOI: 10.7554/ELIFE.83606.SA2

Single cell analysis of the developing mouse kidney provides deeper insight into marker gene expression and ligand-receptor crosstalk

Publisher: The Company of Biologists

Date: 12-06-2019

DOI: 10.1242/DEV.178673

Abstract: Recent advances in the generation of kidney organoids and the culture of primary nephron progenitors from mouse and human have been based on knowledge of the molecular basis of kidney development in mice. Although gene expression during kidney development has been intensely investigated, single cell profiling provides new opportunities to further subsect component cell types and the signalling networks at play. Here, we describe the generation and analysis of 6732 single cell transcriptomes from the fetal mouse kidney [embryonic day (E)18.5] and 7853 sorted nephron progenitor cells (E14.5). These datasets provide improved resolution of cell types and specific markers, including sub ision of the renal stroma and heterogeneity within the nephron progenitor population. Ligand-receptor interaction and pathway analysis reveals novel crosstalk between cellular compartments and associates new pathways with differentiation of nephron and ureteric epithelium cell types. We identify transcriptional congruence between the distal nephron and ureteric epithelium, showing that most markers previously used to identify ureteric epithelium are not specific. Together, this work improves our understanding of metanephric kidney development and provides a template to guide the regeneration of renal tissue.

Phenotypic effects of maternal immune activation and early postnatal milieu in mice mutant for the schizophrenia risk gene neuregulin-1

Publisher: Elsevier BV

Date: 09-2014

DOI: 10.1016/J.NEUROSCIENCE.2014.06.028

Abstract: Risk of schizophrenia is likely to involve gene × environment (G × E) interactions. Neuregulin 1 (NRG1) is a schizophrenia risk gene, hence any interaction with environmental adversity, such as maternal infection, may provide further insights into the basis of the disease. This study examined the in idual and combined effects of prenatal immune activation with polyriboinosinic-polyribocytidilic acid (Poly I:C) and disruption of the schizophrenia risk gene NRG1 on the expression of behavioral phenotypes related to schizophrenia. NRG1 heterozygous (NRG1 HET) mutant breeding pairs were time-mated. Pregnant dams received a single injection (5mg/kg i.p.) of Poly I:C or vehicle on gestation day 9 (GD9). Offspring were then cross-fostered to vehicle-treated or Poly I:C-treated dams. Expression of schizophrenia-related behavioral endophenotypes was assessed at adolescence and in adulthood. Combining NRG1 disruption and prenatal environmental insult (Poly I:C) caused developmental stage-specific deficits in social behavior, spatial working memory and prepulse inhibition (PPI). However, combining Poly I:C and cross-fostering produced a number of behavioral deficits in the open field, social behavior and PPI. This became more complex by combining NRG1 deletion with both Poly I:C exposure and cross-fostering, which had a robust effect on PPI. These findings suggest that concepts of G × E interaction in risk of schizophrenia should be elaborated to multiple interactions that involve in idual genes interacting with erse biological and psychosocial environmental factors over early life, to differentially influence particular domains of psychopathology, sometimes over specific stages of development.

Architectural defects in the spleens of Nkx2-3-deficient mice are intrinsic and associated with defects in both B cell maturation and T cell-dependent immune responses.

Publisher: The American Association of Immunologists

Date: 15-04-2003

DOI: 10.4049/JIMMUNOL.170.8.4002

Abstract: Mice lacking the homeodomain transcription factor Nkx2-3 are either asplenic or develop a spleen of significantly reduced size with poorly organized white pulp. In this report, we analyze the effect of this mutation on B lymphocyte development and differentiation. Follicular dendritic cells in spleen, but not lymph node, of Nkx2-3−/− mice fail to express a developmental Ag (follicular dendritic cell-M2) and show an abnormal association with B cells, despite essentially normal expression of several chemokine genes. Bone marrow reconstitution studies show the splenic disorganization and absence of marginal zone B cells to be of stromal rather than hemopoietic origin. Furthermore, Nkx2-3−/− mice show an excess of conventional B cells in mesenteric lymph node and peritoneal cavity, whereas transitional B cells are rare in spleen but overrepresented in bone marrow. Finally, immunization of Nkx2-3−/− mice with a T cell-dependent Ag elicits clusters of germinal center B cells, although these fail to develop to the same extent as in controls and there is no evidence of affinity maturation in serum Ab. Similarly, Ab-forming cells fail to aggregate into foci early in the response. Collectively, these data indicate a substantial role for Nkx2-3 in the correct association of lymphocytes and splenic stromal elements that is independent of chemokine expression.

Combined Mutation Screening of NKX2-5, GATA4, and TBX5 in Congenital Heart Disease: Multiple Heterozygosity and Novel Mutations

Publisher: Computers, Materials and Continua (Tech Science Press)

Date: 20-10-2011

DOI: 10.1111/J.1747-0803.2011.00573.X

Control of cardiac jelly dynamics by NOTCH1 and NRG1 defines the building plan for trabeculation

Publisher: Springer Science and Business Media LLC

Date: 05-2018

DOI: 10.1038/S41586-018-0110-6

Abstract: In vertebrate hearts, the ventricular trabecular myocardium develops as a sponge-like network of cardiomyocytes that is critical for contraction and conduction, ventricular septation, papillary muscle formation and wall thickening through the process of compaction

Homeodomain factor Nkx2-5 controls left/right asymmetric expression of bHLH gene eHand during murine heart development.

Publisher: Cold Spring Harbor Laboratory

Date: 06-1997

DOI: 10.1101/GAD.11.11.1357

Abstract: One of the first morphological manifestations of left/right (L/R) asymmetry in mammalian embryos is a pronounced rightward looping of the linear heart tube. The direction of looping is thought to be controlled by signals from an embryonic L/R axial system. We report here that morphological L/R asymmetry in the murine heart first became apparent at the linear tube stage as a leftward displacement of its caudal aspect. Beginning at the same stage, the basic helix-loop-helix (bHLH) factor gene eHand was expressed in a strikingly left-dominant pattern in myocardium, reflecting an intrinsic molecular asymmetry. In hearts of embryos lacking the homeobox gene Nkx2-5, which do not loop, left-sided eHand expression was abolished. However, expression was unaffected in Sc1-/- hearts that loop poorly because of hematopoietic insufficiency, and was right-sided in hearts of inv/inv embryos that display situs inversus. The data predict that eHand expression is enhanced in descendants of the left heart progenitor pool as one response to inductive signaling from the L/R axial system, and that eHand controls intrinsic morphogenetic pathways essential for looping. One aspect of the intrinsic response to L/R information falls under Nkx2-5 homeobox control.

Update on the use of stem cells for cardiac disease

Publisher: Wiley

Date: 13-05-2005

DOI: 10.1111/J.1445-5994.2005.00840.X

Abstract: Major advances have recently been made in our understanding of stem cell biology, and in the application of stem cells to treat cardiac disease. Resident cardiac stem cells have now been described and the long-accepted paradigm of the adult mammalian heart as an organ without regenerative capacity has been questioned. Various stem-cell-based approaches for ameliorating cardiac disease have been shown to be beneficial in animal models and are now being trialled in humans, with several phase I clinical studies already completed. Although these clinical studies lacked adequate placebo controls, they have consistently shown promising results. If confirmed by larger phase II/III trials, it is possible that within a few years a powerful new therapeutic option may be available for the burgeoning number of patients suffering from myocardial ischaemia and/or other cardiac disorders.

Phenotype of spontaneous orofacial dyskinesia in neuregulin-1 ‘knockout’ mice

Publisher: Elsevier BV

Date: 03-2009

DOI: 10.1016/J.PNPBP.2008.12.010

Abstract: Studies in antipsychotic-naïve patients with schizophrenia indicate a baseline level of spontaneous involuntary movements, particularly orofacial dyskinesia. Neuregulin-1 is associated with risk for schizophrenia and its functional role can be studied in 'knockout' mice. We have shown previously that neuregulin-1 'knockouts' evidence disruption in social behaviour. Neuregulin-1 'knockouts' were assessed for four topographies of orofacial movement, both spontaneously and under challenge with the D(1)-like dopamine receptor agonist SKF 83959. Neuregulin-1 'knockouts' evidenced an increase in spontaneous incisor chattering, particularly among males. SKF 83959 induced incisor chattering, vertical jaw movements and tongue protrusions the level of horizontal jaw movements was increased and that of tongue protrusions decreased in neuregulin-1 'knockouts'. These findings indicate that the schizophrenia risk gene neuregulin-1 is involved in the regulation of not only social behaviour but also orofacial dyskinesia. Orofacial dyskinesia in neuregulin-1 mutants may indicate some modest genetic relationship between risk for schizophrenia and vulnerability to spontaneous movement disorder.

Loss of Rearranged L-Myc Fusion (RLF) results in defects in heart development in the mouse

Publisher: Elsevier BV

Date: 03-2017

DOI: 10.1016/J.DIFF.2016.11.004

Abstract: Recently we reported that Rearranged L-Myc Fusion, RLF, acts as an epigenetic modifier maintaining low levels of DNA methylation at CpG island shores and enhancers across the genome. Here we focus on the phenotype of Rlf null mutant mice generated via an ENU mutagenesis screen, to identify genes required for epigenetic regulation. RLF is expressed in a range of fetal mouse tissues, including the fetal heart. Comprehensive timed-mating studies are consistent with our previously reported findings that Rlf homozygous mutant mice rarely survive to adulthood, with the majority dying shortly after birth. Histological analysis of two independent Rlf ENU mutant lines at E11.5-E14.5 showed heart defects resembling those present in humans with Left Ventricular Non-Compaction (LVNC). In situ hybridisation analysis localized expression of Rlf to the endocardium and epicardium of embryonic and postnatal hearts, and transiently to cardiomyocytes during heart looping and early chamber formation stages. RNA-seq analysis of Rlf mutant hearts highlighted defective NOTCH pathway signalling, recently describe as one cause of LVNC. This study provides the first evidence that RLF is required for normal heart development in the mouse. The heart morphological defects present at high penetrance in Rlf mutants are consistent with features of LVNC in humans, and molecular analysis identified attenuated JAGGED 1 expression and NOTCH signalling as likely contributors to these defects. Our study highlights the importance of RLF-dependent epigenetic modifications to DNA for maintaining correct gene regulatory network and intercellular signalling interactions during heart chamber and septal development. Further investigations are needed to define the biochemical role of RLF in the developing heart, and whether RLF mutations are a cause of heart defects in humans.

Author response: Nkx2.5 marks angioblasts that contribute to hemogenic endothelium of the endocardium and dorsal aorta

Publisher: eLife Sciences Publications, Ltd

Date: 30-01-2017

DOI: 10.7554/ELIFE.20994.035

Large-Scale Production of Cardiomyocytes from Human Pluripotent Stem Cells Using a Highly Reproducible Small Molecule-Based Differentiation Protocol

Publisher: MyJove Corporation

Date: 25-07-2016

DOI: 10.3791/54276

Quantifying intrinsic and extrinsic control of single-cell fates in cancer and stem/progenitor cell pedigrees with competing risks analysis

Publisher: Springer Science and Business Media LLC

Date: 06-2016

DOI: 10.1038/SREP27100

Abstract: The molecular control of cell fate and behaviour is a central theme in biology. Inherent heterogeneity within cell populations requires that control of cell fate is studied at the single-cell level. Time-lapse imaging and single-cell tracking are powerful technologies for acquiring cell lifetime data, allowing quantification of how cell-intrinsic and extrinsic factors control single-cell fates over time. However, cell lifetime data contain complex features. Competing cell fates, censoring, and the possible inter-dependence of competing fates, currently present challenges to modelling cell lifetime data. Thus far such features are largely ignored, resulting in loss of data and introducing a source of bias. Here we show that competing risks and concordance statistics, previously applied to clinical data and the study of genetic influences on life events in twins, respectively, can be used to quantify intrinsic and extrinsic control of single-cell fates. Using these statistics we demonstrate that 1) breast cancer cell fate after chemotherapy is dependent on p53 genotype 2) granulocyte macrophage progenitors and their differentiated progeny have concordant fates and 3) cytokines promote self-renewal of cardiac mesenchymal stem cells by symmetric isions. Therefore, competing risks and concordance statistics provide a robust and unbiased approach for evaluating hypotheses at the single-cell level.

The Ontogeny of Cardiac Regeneration

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 27-05-2011

DOI: 10.1161/RES.0B013E318222BA1A

Lack of Genetic Interaction between Tbx20 and Tbx3 in Early Mouse Heart Development

Publisher: Public Library of Science (PLoS)

Date: 25-07-2013

DOI: 10.1371/JOURNAL.PONE.0070149

Seeking a regulatory roadmap for heart morphogenesis

Publisher: Elsevier BV

Date: 02-1999

DOI: 10.1006/SCDB.1998.0277

Abstract: Despite descriptive studies spanning centuries, we are still lacking an integrated concept of how molecular patterning information is developed in the heart, and how this information guides morphogenesis. With an increasing number of regionally-expressed cardiac genes and transgenes being identified, along with new animal models of cardiac dysmorphogenesis and an exciting array of genetic tools for further dissection, the need for an integrated morphogenetic concept is acute. Models invoking a linear array of cardiac segments are difficult to reconcile with the observation that those segments are evident only on the outer curvature of the heart tube. Molecular and anatomical evidence supports the view that chamber specification is achieved by interpretation of dorso-ventral (inner curvature/outer curvature), as well as anterior-posterior patterning information in the primary heart tube. In this essay, I examine some of the issues influencing and perhaps confusing our view of cardiac morphogenesis and briefly discuss regulatory genes in the context of an evolving morphogenetic model.

Live cell imaging and single cell tracking of mesenchymal stromal cellsin vitro

Publisher: Wiley

Date: 26-11-2016

DOI: 10.1002/9781118907474.CH24

The Cardiac Expression of Striated Muscle LIM Protein 1 (SLIM1) is Restricted to the Outflow Tract of the Developing Heart

Publisher: Elsevier BV

Date: 04-1999

DOI: 10.1006/JMCC.1998.0922

Abstract: LIM proteins perform critical roles in development and tissue differentiation. The skeletal muscle LIM protein 1 (SLIM1) comprises four and a half LIM domains. Northern blot analysis demonstrated high level expression of SLIM1 mRNA in adult human skeletal muscle with intermediate expression in adult heart and lower expression in other tissues. Western blot analysis using specific affinity-purified anti-SLIM1 antipeptide antibodies demonstrated a 32 kDa polypeptide in the aorta and atria of rabbit heart, but not in vena cava, interventricular septum or ventricular muscle. SLIM1 was also demonstrated in rabbit skeletal muscle. In situ hybridization of whole mouse embryos confirmed the cardiac expression of SLIM1 was restricted to the cardiac outflow tract from embryonic day 8.5-11. No expression was seen in atrial or ventricular muscle. SLIM1 mRNA was also demonstrated in the hindbrain, neural tube and somites. The localized expression of SLIM1 to the outflow tract of the embryonic heart implies an important role for the protein in the development of this region and possibly in congenital heart anomalies involving the separation and formation of the aortic and pulmonary trunks.

Quantitative trait and transcriptome analysis of genetic complexity underpinning cardiac interatrial septation in mice using an advanced intercross line

Publisher: Cold Spring Harbor Laboratory

Date: 11-2022

DOI: 10.1101/2022.10.31.514499

Abstract: Unlike single-gene mutations leading to Mendelian conditions, common human diseases are likely emergent phenomena arising from multilayer, multiscale and highly interconnected interactions. Atrial and ventricular septal defects are the most common forms of cardiac congenital anomalies in humans. Atrial septal defects (ASD) show an open communication between left and right atria postnatally, potentially resulting in serious hemodynamic consequences if untreated. A milder form of atrial septal defect, patent foramen ovale (PFO), exists in about one quarter of the human population, strongly associated with ischaemic stroke and migraine. The anatomic liabilities and genetic and molecular basis of atrial septal defects remain unclear. Here, we advance our previous analysis of atrial septal variation through quantitative trait locus (QTL) mapping of an advanced intercross line (AIL) established between the inbred QSi5 and 129T2/SvEms mouse strains, that show extremes of septal phenotypes. Analysis resolved 37 unique septal QTL with high overlap between QTL for distinct septal traits. Whole genome sequencing of parental strains identified high confidence candidate deleterious variants, including in known human congenital heart disease genes, whereas transcriptome analysis of developing septa revealed networks involving ribosome, nucleosome, mitochondrial and extracellular matrix biosynthesis underlying septal variation. Analysis of variant architecture across different gene features, including enhancers and promoters, provided evidence for involvement of non-coding as well as protein coding variants. Our study provides the first high resolution picture of genetic complexity and network liability underlying common congenital heart disease, with relevance to human ASD and PFO.

Peripheral nervous system defects in erbB2 mutants following genetic rescue of heart development

Publisher: Cold Spring Harbor Laboratory

Date: 10-1999

DOI: 10.1101/GAD.13.19.2538

Abstract: The ErbB2 tyrosine kinase functions as coreceptor for the neuregulin receptors ErbB3 and ErbB4 and can participate in signaling of EGF receptor (ErbB1), interleukin receptor gp130, and G-protein coupled receptors. ErbB2(-/-) mice die at midgestation because of heart malformation. Here, we report a genetic rescue of their heart development by myocardial expression of erbB2 cDNA that allows survival of the mutants to birth. In rescued erbB2 mutants, Schwann cells are lacking. Motoneurons form and can project to muscle, but nerves are poorly fasciculated and disorganized. Neuromuscular junctions form, as reflected in clustering of AChR and postsynaptic expression of the genes encoding the alpha-AChR, AChE, epsilon-AChR, and the RI subunit of the cAMP protein kinase. However, a severe loss of motoneurons on cervical and lumbar, but not on thoracic levels occurs. Our results define the roles of Schwann cells during motoneuron and synapse development, and reveal different survival requirements for distinct motoneuron populations.

Histone genes are clustered with a 15-kilobase repeat in the chicken genome

Publisher: Springer Science and Business Media LLC

Date: 05-1979

DOI: 10.1038/279132A0

Abstract: Histone mRNA isolated from 5-day-old chick embryos has been used as a template for complementary DNA (cDNA) synthesis. The resultant cDNA, after removal of sequences complementary to rRNA, was used to detect histone genes in adult chicken genomal DNA. Hybridisation data indicate that the histone genes are repeated about 10-fold in the chicken genome. Restriction endonuclease analysis reveals some sequence heterogeneity in these genes. However, the results show that chicken histone genes are clustered with a basic repeat unit of 15 kilobases.

A novel conditional mouse model for Nkx2-5 reveals transcriptional regulation of cardiac ion channels

Publisher: Elsevier BV

Date: 2016

DOI: 10.1016/J.DIFF.2015.12.003

Abstract: Nkx2-5 is one of the master regulators of cardiac development, homeostasis and disease. This transcription factor has been previously associated with a suite of cardiac congenital malformations and impairment of electrical activity. When disease causative mutations in transcription factors are considered, NKX2-5 gene dysfunction is the most common abnormality found in patients. Here we describe a novel mouse model and subsequent implications of Nkx2-5 loss for aspects of myocardial electrical activity. In this work we have engineered a new Nkx2-5 conditional knockout mouse in which flox sites flank the entire Nkx2-5 locus, and validated this line for the study of heart development, differentiation and disease using a full deletion strategy. While our homozygous knockout mice show typical embryonic malformations previously described for the lack of the Nkx2-5 gene, hearts of heterozygous adult mice show moderate morphological and functional abnormalities that are sufficient to sustain blood supply demands under homeostatic conditions. This study further reveals intriguing aspects of Nkx2-5 function in the control of cardiac electrical activity. Using a combination of mouse genetics, biochemistry, molecular and cell biology, we demonstrate that Nkx2-5 regulates the gene encoding Kcnh2 channel and others, shedding light on potential mechanisms generating electrical abnormalities observed in patients bearing NKX2-5 dysfunction and opening opportunities to the study of novel therapeutic targets for anti-arrhythmogenic therapies.

High-resolution genetic analysis of a deletion on mouse Chromosome 17 extending over the Fused, tufted, and homeobox Nkx2-5 loci

Publisher: Springer Science and Business Media LLC

Date: 12-1994

DOI: 10.1007/BF00292022

Abstract: Expansion microscopy (ExM) has been successfully used to improve the spatial resolution when imaging tissues by optical microscopy. In ExM, proteins of a fixed s le are crosslinked to a swellable acrylamide gel, which expands when incubated in water. Therefore, ExM allows enlarged subcellular structures to be resolved that would otherwise be hidden to standard confocal microscopy. Herein, we aim to validate ExM for the study of peroxisomes, mitochondria, nuclei and the plasma membrane. Upon comparison of the expansion factors of these cellular compartments in HEK293 cells within the same gel, we found significant differences, of a factor of above 2, in expansion factors. For peroxisomes, the expansion factor differed even between peroxisomal membrane and matrix marker this underlines the need for a thorough validation of expansion factors of this powerful technique. We further give an overview of possible quantification methods for the determination of expansion factors of intracellular organelles, and we highlight some potentials and challenges.

Responsiveness of Naive CD4 T Cells to Polarizing Cytokine Determines the Ratio of Th1 and Th2 Cell Differentiation

Publisher: The American Association of Immunologists

Date: 02-2006

DOI: 10.4049/JIMMUNOL.176.3.1553

Abstract: The intrinsic features of naive CD4 T cells that affect their ability to respond to polarizing signals for Th cell differentiation are not well understood. In this study, we show that naive CD4 T cells from mice transgenic for the Hlx gene expressed lower levels of IL-4Rα. The down-regulation of IL-4Rα diminished IL-4 signaling and the Th2 response and enhanced the Th1 response under suboptimal polarizing conditions. In nontransgenic CD4 T cells, blocking IL-4Rα with Abs had the same effect in an Ab dose-dependent manner. Conversely, Hlx haploinsufficiency caused higher expression of IL-4Rα to favor Th2 cell differentiation. Thus, the IL-4Rα level on naive CD4 T cells is genetically controlled by Hlx and determines the ratio of Th1 and Th2 cell differentiation.

A common Shox2-Nkx2-5 antagonistic mechanism primes the pacemaking cell fate in the pulmonary vein myocardium and sinoatrial node

Publisher: The Company of Biologists

Date: 2015

DOI: 10.1242/DEV.120220

Abstract: In humans, atrial fibrillation is often triggered by ectopic pacemaking activity in the myocardium sleeves of the pulmonary vein (PV) and systemic venous return. However, the genetic programs that abnormally reinforce pacemaker properties at these sites and how this relates to normal sinoatrial node (SAN) development remain uncharacterized. It was noted previously that Nkx2-5, which is expressed in the PV myocardium and reinforces a chamber-liker myocardial identity in the PV, is lacking in the SAN. Here we present evidence that Shox2 antagonizes the transcription output of Nkx2-5 in the PV myocardium and in a functional Nkx2-5+ domain within the SAN to determine the cell fate. Shox2 deletion in the Nkx2-5+ domain of the SAN caused sick sinus syndrome, associated with the loss of pacemaker program. Explanted Shox2+ cells from the embryonic PV myocardium exhibited pacemaker characteristics including node-like electrophysiological properties and the capability to pace surrounding Shox2− cells. Shox2 deletion led to Hcn4 obliteration in the developing PV myocardium. Nkx2-5 hypomorphism rescued the requirement for Shox2 for the expression of genes essential for SAN development in Shox2 mutants. Similarly, the pacemaker-like phenotype induced in the PV myocardium in Nkx2-5 hypomorphs reverted back to a working myocardial phenotype when Shox2 was simultaneously deleted. A similar mechanism is also adopted in differentiated embryoid-bodies. Moreover, we found that Shox2 interacts with Nkx2-5 directly, and discovered a substantial genome wide co-occupancy of Shox2, Nkx2-5, and Tbx5, further supporting a pivotal role for Shox2 in the core myogenic program orchestrating venous pole and pacemaker development.

Generation of conditionalCited2 null alleles

Publisher: Wiley

Date: 2006

DOI: 10.1002/DVG.20251

Abstract: Cited2 is a transcriptional co-factor that is widely expressed in both embryonic and extraembryonic cells during early development. It is essential for embryonic development with Cited2 null embryos showing abnormal development of organs including heart, neural tube, adrenal glands, and placenta (both in trophoblast derivatives and invading fetal vasculature), as well as having defects in the establishment of the left-right body axis. We report the generation of two conditional null alleles allowing Cre-recombinase-mediated somatic cell gene inactivation. Mice heterozygous or homozygous for these alleles are viable and fertile. Crossing conditional mutants with CMV-Cre transgenic mice produces an embryonic-lethal phenotype in the offspring indistinguishable from germline null mutants. We also demonstrate that conditional deletion results in lacZ expression under the control of the Cited2 promoter. These alleles are therefore useful genetic tools for dissecting the functions of Cited2 in the formation of different organs and patterning of the developing embryo. genesis

Advances in the Genetics of Congenital Heart Disease: A Clinician's Guide

Publisher: Elsevier BV

Date: 02-2017

DOI: 10.1016/J.JACC.2016.11.060

Abstract: Our understanding of the genetics of congenital heart disease (CHD) is rapidly expanding however, many questions, particularly those relating to sporadic forms of disease, remain unanswered. Massively parallel sequencing technology has made significant contributions to the field, both from a diagnostic perspective for patients and, importantly, also from the perspective of disease mechanism. The importance of de novo variation in sporadic disease is a recent highlight, and the genetic link between heart and brain development has been established. Furthermore, evidence of an underlying burden of genetic variation contributing to sporadic and familial forms of CHD has been identified. Although we are still unable to identify the cause of CHD for most patients, recent findings have provided us with a much clearer understanding of the types of variants and their in idual contributions and collectively mark an important milestone in our understanding of both familial and sporadic forms of disease.

Tinman/Nkx2-5 acts via miR-1 and upstream of Cdc42 to regulate heart function across species

Publisher: Rockefeller University Press

Date: 20-06-2011

DOI: 10.1083/JCB.201006114

Abstract: Unraveling the gene regulatory networks that govern development and function of the mammalian heart is critical for the rational design of therapeutic interventions in human heart disease. Using the Drosophila heart as a platform for identifying novel gene interactions leading to heart disease, we found that the Rho-GTPase Cdc42 cooperates with the cardiac transcription factor Tinman/Nkx2-5. Compound Cdc42, tinman heterozygous mutant flies exhibited impaired cardiac output and altered myofibrillar architecture, and adult heart–specific interference with Cdc42 function is sufficient to cause these same defects. We also identified K+ channels, encoded by dSUR and slowpoke, as potential effectors of the Cdc42–Tinman interaction. To determine whether a Cdc42–Nkx2-5 interaction is conserved in the mammalian heart, we examined compound heterozygous mutant mice and found conduction system and cardiac output defects. In exploring the mechanism of Nkx2-5 interaction with Cdc42, we demonstrated that mouse Cdc42 was a target of, and negatively regulated by miR-1, which itself was negatively regulated by Nkx2-5 in the mouse heart and by Tinman in the fly heart. We conclude that Cdc42 plays a conserved role in regulating heart function and is an indirect target of Tinman/Nkx2-5 via miR-1.

Somatic mutations in NKX2-5, GATA4, and HAND1 are not a common cause of tetralogy of Fallot or hypoplastic left heart

Publisher: Wiley

Date: 16-08-2011

DOI: 10.1002/AJMG.A.34187

Abstract: The majority of congenital heart disease (CHD) occurs as a sporadic finding, with a minority of cases associated with a known genetic abnormality. Combinations of genetic and environmental factors are implicated, with the recent and intriguing hypothesis that an apparently high rate of somatic mutations might explain some sporadic CHD. We used s les of right ventricular myocardium from patients undergoing surgical repair of tetralogy of Fallot (TOF) and hypoplastic left heart (HLH) to examine the incidence of somatic mutation in cardiac tissue. TOF is a common form of cyanotic CHD, occurring in 3.3 per 10,000 live births. HLH is a rare defect in which the left side of the heart is severely under-developed. Both are severe malformations whose genetic etiology is largely unknown. We carried out direct sequence analysis of the NKX2–5 and GATA4 genes from fresh frozen cardiac tissues and matched blood s les of nine TOF patients. Analysis of NKX2–5, GATA4, and HAND1 was performed from cardiac tissue of 24 HLH patients and three matched blood s les. No somatic or germline mutations were identified in the TOF or HLH patients. Although limited by s le size, our study suggests that somatic mutations in NKX2–5 and GATA4 are not a common cause of isolated TOF or HLH.

Wnt inhibition promotes vascular specification of embryonic cardiac progenitors

Publisher: The Company of Biologists

Date: 2017

DOI: 10.1242/DEV.159905

Abstract: Several studies have demonstrated a multiphasic role for Wnt signaling during embryonic cardiogenesis (Naito et al., 2006 Qyang et al., 2007) and developed protocols that enrich for cardiac derivatives during in vitro differentiation of human pluripotent stem cells (hPSC) (Elliott et al., 2011 Iyer et al., 2016 Lian et al., 2012 Paige et al., 2010 Willems et al., 2011 Witty et al., 2014), however, few studies have investigated the role of Wnt signaling in specification of cardiac progenitor cells (CPC) toward downstream fates. Using transgenic mice and hPSCs, we tracked endothelial cells (EC) that originated from CPCs expressing Nkx2.5. Analysis of EC-fated CPCs at discrete phenotypic milestones during hPSC differentiation identified reduced Wnt activity as a hallmark of EC specification, and enforced activation or inhibition of Wnt reduced or increased, respectively, the degree of vascular commitment within the CPC population during both hPSC differentiation and mouse embryogenesis. Wnt5a, which has been shown to play an inhibitory role on Wnt during cardiac development, was dynamically expressed during vascular commitment of hPSC-derived CPCs and ectopic Wnt5a promoted vascular specification of hPSC-derived and mouse embryonic CPCs.

Platelet‐Derived Growth Factor Receptor Type α Activation Drives Pulmonary Vascular Remodeling Via Progenitor Cell Proliferation and Induces Pulmonary Hypertension

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 05-04-2022

DOI: 10.1161/JAHA.121.023021

Abstract: Platelet‐derived growth factor is a major regulator of the vascular remodeling associated with pulmonary arterial hypertension. We previously showed that protein widely 1 (PW1 + ) vascular progenitor cells participate in early vessel neomuscularization during experimental pulmonary hypertension (PH) and we addressed the role of the platelet‐derived growth factor receptor type α (PDGFRα) pathway in progenitor cell‐dependent vascular remodeling and in PH development. Remodeled pulmonary arteries from patients with idiopathic pulmonary arterial hypertension showed an increased number of perivascular and vascular PW1 + cells expressing PDGFRα. PW1 nLacZ reporter mice were used to follow the fate of pulmonary PW1 + progenitor cells in a model of chronic hypoxia–induced PH development. Under chronic hypoxia, PDGFRα inhibition prevented the increase in PW1 + progenitor cell proliferation and differentiation into vascular smooth muscle cells and reduced pulmonary vessel neomuscularization, but did not prevent an increased right ventricular systolic pressure or the development of right ventricular hypertrophy. Conversely, constitutive PDGFRα activation led to neomuscularization via PW1 + progenitor cell differentiation into new smooth muscle cells and to PH development in male mice without fibrosis. In vitro, PW1 + progenitor cell proliferation, but not differentiation, was dependent on PDGFRα activity. These results demonstrate a major role of PDGFRα signaling in progenitor cell–dependent lung vessel neomuscularization and vascular remodeling contributing to PH development, including in idiopathic pulmonary arterial hypertension patients. Our findings suggest that PDGFRα blockers may offer a therapeutic add‐on strategy to combine with current pulmonary arterial hypertension treatments to reduce vascular remodeling. Furthermore, our study highlights constitutive PDGFRα activation as a novel experimental PH model.

Zac1 Is an Essential Transcription Factor for Cardiac Morphogenesis

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 02-04-2010

DOI: 10.1161/CIRCRESAHA.109.214130

Abstract: Rationale : The transcriptional networks guiding heart development remain poorly understood, despite the identification of several essential cardiac transcription factors. Objective : To isolate novel cardiac transcription factors, we performed gene chip analysis and found that Zac1 , a zinc finger-type transcription factor, was strongly expressed in the developing heart. This study was designed to investigate the molecular and functional role of Zac1 as a cardiac transcription factor. Methods and Results : Zac1 was strongly expressed in the heart from cardiac crescent stages and in the looping heart showed a chamber-restricted pattern. Zac1 stimulated luciferase reporter constructs driven by ANF, BNP , or α MHC promoters. Strong functional synergy was seen between Zac1 and Nkx2-5 on the ANF promoter, which carries adjacent Zac1 and Nkx2-5 DNA-binding sites. Zac1 directly associated with the ANF promoter in vitro and in vivo, and Zac1 and Nkx2-5 physically associated through zinc fingers 5 and 6 in Zac1, and the homeodomain in Nkx2-5. Zac1 is a maternally imprinted gene and is the first such gene found to be involved in heart development. Homozygous and paternally derived heterozygous mice carrying an interruption in the Zac1 locus showed decreased levels of chamber and myofilament genes, increased apoptotic cells, partially penetrant lethality and morphological defects including atrial and ventricular septal defects, and thin ventricular walls. Conclusions : Zac1 plays an essential role in the cardiac gene regulatory network. Our data provide a potential mechanistic link between Zac1 in cardiogenesis and congenital heart disease manifestations associated with genetic or epigenetic defects in an imprinted gene network.

Preface

Publisher: Elsevier

Date: 2010

DOI: 10.1016/B978-0-12-381332-9.00048-7

Loss of Cited2 causes congenital heart disease by perturbing left–right patterning of the body axis

Publisher: Oxford University Press (OUP)

Date: 24-12-2010

DOI: 10.1093/HMG/DDQ554

Abstract: Cited2 is a transcriptional coactivator that is required for normal development of the embryo and placenta. Cited2-null mice die during gestation with fully penetrant heart defects and partially penetrant laterality defects. The laterality defects occur due to the loss of Nodal expression in the left lateral plate mesoderm (LPM). The cause of the heart defects that arise independently of laterality defects is unknown they might occur due to an intrinsic requirement for Cited2 in the developing heart, or to disturbances in left-right patterning of the early embryo. Herein it is established that deletion of Cited2 from the heart progenitors does not alter development, and that heart defects in Cited2-null embryos arise due to an extra-cardiac requirement for Cited2 in establishing the left-right body axis. In addition, we provide evidence supporting a role for Cited2 in tissues of the embryo vital for left-right patterning (the node and LPM). Molecular and genetic analysis reveals that Cited2 is required for the initiation, but not propagation of, the left-sided determinant Nodal in the LPM. Moreover, a new role for Cited2 is identified as a potentiator of bone morphogenetic protein (BMP) signalling, counteracting the initiation of Nodal expression in the LPM. These data define Cited2 as a key regulator of left-right patterning in the mammalian embryo, and reveal that the role of Cited2 in cardiac development lies in its extra-cardiac functions. The clinical relevance of these findings lies in the fact that heterozygous mutation of human CITED2 is associated with congenital heart disease and laterality defects.

Irreversible Triggers for Hypertrophic Cardiomyopathy Are Established in the Early Postnatal Period

Publisher: Elsevier BV

Date: 02-2015

DOI: 10.1016/J.JACC.2014.10.069

Abstract: Hypertrophic cardiomyopathy (HCM) is caused by mutations in sarcomere protein genes, and left ventricular hypertrophy (LVH) develops as an adaptive response to sarcomere dysfunction. It remains unclear whether persistent expression of the mutant gene is required for LVH or whether early gene expression acts as an immutable inductive trigger. The aim of this study was to use a regulatable murine model of HCM to study the reversibility of pathological LVH. The authors generated a double-transgenic mouse model, tTAxαMHCR403Q, in which expression of the HCM-causing Arg403Gln mutation in the α-myosin heavy chain (MHC) gene is inhibited by doxycycline administration. Cardiac structure and function were evaluated in groups of mice that received doxycycline for varying periods from 0 to 40 weeks of age. Untreated tTAxαMHCR403Q mice showed increased left ventricular (LV) mass, contractile dysfunction, myofibrillar disarray, and fibrosis. In contrast, mice treated with doxycycline from conception to 6 weeks had markedly less LVH and fibrosis at 40 weeks. Transgene inhibition from 6 weeks reduced fibrosis but did not prevent LVH or functional changes. There were no differences in LV parameters at 40 weeks between mice with transgene inhibition from 20 weeks and mice with continuous transgene expression. These findings highlight the critical role of the early postnatal period in HCM pathogenesis and suggest that mutant sarcomeres manifest irreversible cardiomyocyte defects that induce LVH. In HCM, mutation-silencing therapies are likely to be ineffective for hypertrophy regression and would have to be administered very early in life to prevent hypertrophy development.

α-Cardiac myosin heavy chain (MYH6) mutations affecting myofibril formation are associated with congenital heart defects

Publisher: Oxford University Press (OUP)

Date: 23-07-2010

DOI: 10.1093/HMG/DDQ315

Abstract: Congenital heart defects (CHD) are collectively the most common form of congenital malformation. Studies of human cases and animal models have revealed that mutations in several genes are responsible for both familial and sporadic forms of CHD. We have previously shown that a mutation in MYH6 can cause an autosomal dominant form of atrial septal defect (ASD), whereas others have identified mutations of the same gene in patients with hypertrophic and dilated cardiomyopathy. In the present study, we report a mutation analysis of MYH6 in patients with a wide spectrum of sporadic CHD. The mutation analysis of MYH6 was performed in DNA s les from 470 cases of isolated CHD using denaturing high-performance liquid chromatography and sequence analysis to detect point mutations and small deletions or insertions, and multiplex lifiable probe hybridization to detect partial or complete copy number variations. One non-sense mutation, one splicing site mutation and seven non-synonymous coding mutations were identified. Transfection of plasmids encoding mutant and non-mutant green fluorescent protein-MYH6 fusion proteins in mouse myoblasts revealed that the mutations A230P and A1366D significantly disrupt myofibril formation, whereas the H252Q mutation significantly enhances myofibril assembly in comparison with the non-mutant protein. Our data indicate that functional variants of MYH6 are associated with cardiac malformations in addition to ASD and provide a novel potential mechanism. Such phenotypic heterogeneity has been observed in other genes mutated in CHD.

Nkx2–5 transactivates the Ets-related protein 71 gene and specifies an endothelial/endocardial fate in the developing embryo

Publisher: Proceedings of the National Academy of Sciences

Date: 20-01-2009

DOI: 10.1073/PNAS.0807583106

Abstract: Recent studies support the existence of a common progenitor for the cardiac and endothelial cell lineages, but the underlying transcriptional networks responsible for specification of these cell fates remain unclear. Here we demonstrated that Ets-related protein 71 (Etsrp71), a newly discovered ETS family transcription factor, was a novel downstream target of the homeodomain protein, Nkx2–5. Using genetic mouse models and molecular biological techniques, we demonstrated that Nkx2–5 binds to an evolutionarily conserved Nkx2–5 response element in the Etsrp71 promoter and induces the Etsrp71 gene expression in vitro and in vivo. Etsrp71 was transiently expressed in the endocardium/endothelium of the developing embryo (E7.75-E9.5) and was extinguished during the latter stages of development. Using a gene disruption strategy, we found that Etsrp71 mutant embryos lacked endocardial/endothelial lineages and were nonviable. Moreover, using transgenic technologies and transcriptional and chromatin immunoprecipitation (ChIP) assays, we further established that Tie2 is a direct downstream target of Etsrp71. Collectively, our results uncover a novel functional role for Nkx2–5 and define a transcriptional network that specifies an endocardial/endothelial fate in the developing heart and embryo.

Investigation Between the S377G3 GATA-4 Polymorphism and Migraine

Publisher: Bentham Science Publishers Ltd.

Date: 25-09-2008

DOI: 10.2174/1874205X00802010035

Analysis of steric effects in DamID profiling of transcription factor target genes

Publisher: Elsevier BV

Date: 03-2017

DOI: 10.1016/J.YGENO.2017.01.006

Abstract: DNA adenine methyltransferase identification (DamID) is an enzymatic technology for detecting DNA regions targeted by chromatin-associated proteins. Proteins are fused to bacterial DNA adenine methyltransferase (Dam) and expressed in cultured cells or whole organisms. Here, we used DamID to detect DNA regions bound by the cardiac-restricted transcription factors (TFs) NKX2-5 and SRF, and ubiquitously-expressed co-factors ELK1 and ELK4. We compared targets bound by these TFs as N- and C-terminal fusions with Dam, for both wild type (WT) NKX2-5 and mutant proteins mimicking those found in congenital heart disease. Overall, DamID is highly robust: while the orientation of WT Dam fusions can affect the size of the target sets, their signatures remained largely reproducible. Furthermore, a severe NKX2-5 mutant lacking the homeodomain showed strong steric effects negatively impacting target discovery. The extent of steric effect is likely to be dependent on the protein in question and the orientation of Dam fusion.

Bioengineering and Stem Cell Technology in the Treatment of Congenital Heart Disease.

Publisher: MDPI AG

Date: 22-04-2015

DOI: 10.3390/JCM4040768

QTL mapping of complex binary traits in an advanced intercross line

Publisher: Wiley

Date: 28-06-2012

DOI: 10.1111/J.1365-2052.2012.02383.X

Abstract: An advanced intercross line (AIL) is an easier and more cost-effective approach compared to recombinant inbred lines for fine mapping of quantitative trait loci (QTL) identified by F(2) designs. In an AIL, a complex binary trait can be mapped through analysis of either continuously distributed proxy traits for the liability of the binary trait or the liability itself, the latter presenting the greater statistical challenge. In another work, we successfully applied both approaches in an AIL to fine map previously identified QTL underlying anatomical parameters of the cardiac inter-atrial septum including patent foramen ovale. Here, we describe the statistical methods that we used to analyse complex binary traits in our AIL design. This is achieved using a likelihood-based method, with the expectation-maximisation algorithm allowing use of standard logistic regression methods for model fitting.

Novel murine homeo box gene on chromosome 1 expressed in specific hematopoietic lineages and during embryogenesis.

Publisher: Cold Spring Harbor Laboratory

Date: 04-1991

DOI: 10.1101/GAD.5.4.509

Abstract: We describe here a new murine homeo box gene, denoted Hlx, which is expressed within specific hematopoietic lineages. The cDNA sequence indicates that Hlx differs markedly from known vertebrate homeo box genes, and linkage analysis of an interspecific murine backcross showed that it resides at a novel homeo box locus on the distal portion of mouse chromosome 1. The Hlx homeo domain is most similar to that of the Drosophila H2.0 gene, but outside this region the two polypeptides are related only within a few short segments, the most notable being a motif (denoted Hep) also partially conserved in the engrailed and invected homeo proteins and possibly related to an octapeptide in certain paired box proteins. The presence of an intron within the Hlx homeo box at the same position as in several ergent Drosophila genes (H2.0, labial, Distal-less, proboscipedia, Abdominal-B, NK-1) suggests an ancient evolutionary relationship between these genes. RNA analysis of 67 murine hematopoietic cell lines and normal hematopoietic cells revealed Hlx expression throughout the myeloid/macrophage lineage and at early stages of B lymphocyte development but not in T lymphocytes, erythroid cells, or mast cells. Hence, Hlx is a candidate regulator of hematopoietic lineage commitment and maturation. It probably also functions outside the hematopoietic system, however, because Hlx mRNA could be detected in erse adult tissues and in embryos from as early as day 8 of development.

Investigation of Association between PFO Complicated by Cryptogenic Stroke and a Common Variant of the Cardiac Transcription Factor GATA4

Publisher: Public Library of Science (PLoS)

Date: 06-06-2011

DOI: 10.1371/JOURNAL.PONE.0020711

Increasing the Tolerance of DCD Hearts to Warm Ischemia by Pharmacological Postconditioning

Publisher: Elsevier BV

Date: 08-2014

DOI: 10.1111/AJT.12782

Abstract: Donation after circulatory death (DCD) offers a potential additional source of cardiac allografts. We used a porcine asphyxia model to evaluate viability of DCD hearts subjected to warm ischemic times (WIT) of 20–40 min prior to flushing with Celsior (C) solution. We then assessed potential benefits of supplementing C with erythropoietin, glyceryl trinitrate and zoniporide (Cs), a combination that we have shown previously to activate ischemic postconditioning pathways. Hearts flushed with C/Cs were assessed for functional, biochemical and metabolic recovery on an ex vivo working heart apparatus. Hearts exposed to 20-min WIT showed full recovery of functional and metabolic profiles compared with control hearts (no WIT). Hearts subjected to 30- or 40-min WIT prior to C solution showed partial and no recovery, respectively. Hearts exposed to 30-min WIT and Cs solution displayed complete recovery, while hearts exposed to 40-min WIT and Cs solution demonstrated partial recovery. We conclude that DCD hearts flushed with C solution demonstrate complete recovery up to 20-min WIT after which there is rapid loss of viability. Cs extends the limit of WIT tolerability to 30 min. DCD hearts with ≤30-min WIT may be suitable for transplantation and warrant assessment in a transplant model.

Phenotypic characterization of spatial cognition and social behavior in mice with ‘knockout’ of the schizophrenia risk gene neuregulin 1

Publisher: Elsevier BV

Date: 06-2007

DOI: 10.1016/J.NEUROSCIENCE.2007.03.051

Abstract: Neuregulin-1 (NRG1) has been identified as a candidate susceptibility gene for schizophrenia. In the present study the functional role of the NRG1 gene, as it relates to cognitive and social processes known to be disrupted in schizophrenia, was assessed in mice with heterozygous deletion of transmembrane (TM)-domain NRG1 in comparison with wildtypes (WT). Social affiliative behavior was assessed using the sociability and preference for social novelty paradigm, in terms of time spent in: (i) a chamber containing an unfamiliar conspecific vs. an empty chamber (sociability), or (ii) a chamber containing an unfamiliar conspecific vs. a chamber containing a familiar conspecific (preference for social novelty). Social dominance and aggressive behavior were examined in the resident-intruder paradigm. Spatial learning and memory were assessed using the Barnes maze paradigm, while spatial working memory was measured using the continuous variant of the spontaneous alternation task. Barnes maze data revealed intact spatial learning in NRG1 mutants, with elevated baseline latency to enter the escape hole in male NRG1 mutants reflecting an increase in activity level. Similarly, although a greater number of overall arm entries were found, spontaneous alternation was unaffected in NRG1 mice. Social affiliation data revealed NRG1 mutants to evidence a specific loss of WT preference for spending time with an unfamiliar as opposed to a familiar conspecific. This suggests that NRG1 mutants show a selective impairment in response to social novelty. While spatial learning and working memory processes appear intact, heterozygous deletion of TM-domain NRG1 was associated with disruption to social novelty behavior. These data inform at a novel phenotypic level on the functional role of this gene in the context of its association with risk for schizophrenia.

Cardiac Regeneration Therapies - Targeting Neuregulin 1 Signalling

Publisher: Elsevier BV

Date: 2016

DOI: 10.1016/J.HLC.2015.08.014

Pitx2c and Nkx2-5 Are Required for the Formation and Identity of the Pulmonary Myocardium

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 26-10-2007

DOI: 10.1161/CIRCRESAHA.107.161182

Abstract: The pulmonary vein is sleeved by myocardium, which is a major source of atrial fibrillation and is involved in congenital sinus venosus defects. Little is known about the cellular origin and mechanism of formation of the pulmonary myocardium. We observed a biphasic process of pulmonary myocardium formation in mice. Firstly, a myocardial cell population forms de novo at the connection of the pulmonary vein and the atrium. Genetic labeling revealed that atrial cells do not contribute to this population, indicating it forms by differentiation of pulmonary mesenchymal cells. Secondly, these pulmonary myocardial cells initiate a phase of rapid proliferation and form the pulmonary myocardial sleeve. Pitx2c -deficient mice do not develop a pulmonary myocardial sleeve because they fail to form the initial pulmonary myocardial cells. Genetic-labeling analyses demonstrated that whereas the systemic venous return derives from Nkx2-5 –negative precursors, the pulmonary myocardium derives from Nkx2-5 –expressing precursors, indicating a distinct origin of the 2 venous systems. Nkx2-5 and its target gap-junction gene Cx40 are expressed in the atria and in the pulmonary myocardium but not in the systemic venous return, which expresses the essential pacemaker channel Hcn4 . When Nkx2-5 protein level was lowered in a hypomorphic model, the pulmonary myocardium switched to a Cx40 -negative, Hcn4 -positive phenotype resembling that of the systemic venous return. In conclusion, our data suggest a cellular mechanism for pulmonary myocardium formation and highlight the key roles played by Pitx2c and Nkx2-5 in its formation and identity.

Altered cytokine profile, pain sensitivity, and stress responsivity in mice with co-disruption of the developmental genes Neuregulin-1×DISC1

Publisher: Elsevier BV

Date: 03-2017

DOI: 10.1016/J.BBR.2016.11.049

Abstract: The complex genetic origins of many human disorders suggest that epistatic (gene×gene) interactions may contribute to a significant proportion of their heritability estimates and phenotypic heterogeneity. Simultaneous disruption of the developmental genes and schizophrenia risk factors Neuregulin-1 (NRG1) and Disrupted-in-schizophrenia 1 (DISC1) in mice has been shown to produce disease-relevant and domain-specific phenotypic profiles different from that observed following disruption of either gene alone. In the current study, anxiety and stress responsivity phenotypes in male and female mutant mice with simultaneous disruption of DISC1 and NRG1 were examined. NRG1×DISC1 mutant mice were generated and adult mice from each genotype were assessed for pain sensitivity (hot plate and tail flick tests), anxiety (light-dark box), and stress-induced hypothermia. Serum s les were assayed to measure circulating levels of pro-inflammatory cytokines. Mice with the NRG1 mutation, irrespective of DISC1 mutation, spent significantly more time in the light chamber, displayed increased core body temperature following acute stress, and decreased pain sensitivity. Basal serum levels of cytokines IL8, IL1β and IL10 were decreased in NRG1 mutants. Mutation of DISC1, in the absence of epistatic interaction with NRG1, was associated with increased serum levels of IL1β. Epistatic effects were evident for IL6, IL12 and TNFα. NRG1 mutation alters stress and pain responsivity, anxiety, and is associated with changes in basal cytokine levels. Epistasis resulting from synergistic NRG1 and DISC1 gene mutations altered pro-inflammatory cytokine levels relative to the effects of each of these genes in idually, highlighting the importance of epistatic mechanisms in immune-related pathology.

Time to mend a broken heart

Publisher: Elsevier BV

Date: 10-2007

DOI: 10.1016/J.SCR.2007.09.001

Heart Matters

Publisher: Elsevier BV

Date: 11-2001

DOI: 10.1016/S0092-8674(01)00556-6

Cardiac T-box factor Tbx20 directly interacts with Nkx2-5, GATA4, and GATA5 in regulation of gene expression in the developing heart

Publisher: Elsevier BV

Date: 10-2003

DOI: 10.1016/S0012-1606(03)00385-3

Abstract: Tbx20 is a member of the T-box transcription factor family expressed in the forming hearts of vertebrate and invertebrate embryos. We report here analysis of Tbx20 expression during murine cardiac development and assessment of DNA-binding and transcriptional properties of Tbx20 isoforms. Tbx20 was expressed in myocardium and endocardium, including high levels in endocardial cushions. cDNAs generated by alternative splicing encode at least four Tbx20 isoforms, and Tbx20a uniquely carried strong transactivation and transrepression domains in its C terminus. Isoforms with an intact T-box bound specifically to DNA sites resembling the consensus brachyury half site, although with less avidity compared with the related factor, Tbx5. Tbx20 physically interacted with cardiac transcription factors Nkx2-5, GATA4, and GATA5, collaborating to synergistically activate cardiac gene expression. Among cardiac GATA factors, there was preferential synergy with GATA5, implicated in endocardial differentiation. In Xenopus embryos, enforced expression of Tbx20a, but not Tbx20b, led to induction of mesodermal and endodermal lineage markers as well as cell migration, indicating that the long Tbx20a isoform uniquely bears functional domains that can alter gene expression and developmental behaviour in an in vivo context. We propose that Tbx20 plays an integrated role in the ancient myogenic program of the heart, and has been additionally coopted during evolution of vertebrates for endocardial cushion development.

Developmental origin and lineage plasticity of endogenous cardiac stem cells

Publisher: The Company of Biologists

Date: 15-04-2016

DOI: 10.1242/DEV.111591

Abstract: Over the past two decades, several populations of cardiac stem cells have been described in the adult mammalian heart. For the most part, however, their lineage origins and in vivo functions remain largely unexplored. This Review summarizes what is known about different populations of embryonic and adult cardiac stem cells, including KIT+, PDGFRα+, ISL1+ and SCA1+ cells, side population cells, cardiospheres and epicardial cells. We discuss their developmental origins and defining characteristics, and consider their possible contribution to heart organogenesis and regeneration. We also summarize the origin and plasticity of cardiac fibroblasts and circulating endothelial progenitor cells, and consider what role these cells have in contributing to cardiac repair.

Uncontrolled angiogenic precursor expansion causes coronary artery anomalies in mice lacking Pofut1

Publisher: Springer Science and Business Media LLC

Date: 18-09-2017

DOI: 10.1038/S41467-017-00654-W

Abstract: Coronary artery anomalies may cause life-threatening cardiac complications however, developmental mechanisms underpinning coronary artery formation remain ill-defined. Here we identify an angiogenic cell population for coronary artery formation in mice. Regulated by a DLL4/NOTCH1/VEGFA/VEGFR2 signaling axis, these angiogenic cells generate mature coronary arteries. The NOTCH modulator POFUT1 critically regulates this signaling axis. POFUT1 inactivation disrupts signaling events and results in excessive angiogenic cell proliferation and plexus formation, leading to anomalous coronary arteries, myocardial infarction and heart failure. Simultaneous VEGFR2 inactivation fully rescues these defects. These findings show that dysregulated angiogenic precursors link coronary anomalies to ischemic heart disease.

A rapid co-culture stamping device for studying intercellular communication

Publisher: Springer Science and Business Media LLC

Date: 18-10-2016

DOI: 10.1038/SREP35618

Abstract: Regulation of tissue development and repair depends on communication between neighbouring cells. Recent advances in cell micro-contact printing and microfluidics have facilitated the in-vitro study of homotypic and heterotypic cell-cell interaction. Nonetheless, these techniques are still complicated to perform and as a result, are seldom used by biologists. We report here development of a temporarily sealed microfluidic st ing device which utilizes a novel valve design for patterning two adherent cell lines with well-defined interlacing configurations to study cell-cell interactions. We demonstrate post-st ing cell viability of %, the st ing of multiple adherent cell types, and the ability to control the seeded cell density. We also show viability, proliferation and migration of cultured cells, enabling analysis of co-culture boundary conditions on cell fate. We also developed an in-vitro model of endothelial and cardiac stem cell interactions, which are thought to regulate coronary repair after myocardial injury. The st is fabricated using microfabrication techniques, is operated with a lab pipettor and uses very low reagent volumes of 20 μl with cell injection efficiency of %. This easy-to-use device provides a general strategy for micro-patterning of multiple cell types and will be important for studying cell-cell interactions in a multitude of applications.

Single allele mutations at the heart of congenital disease.

Publisher: American Society for Clinical Investigation

Date: 12-1999

DOI: 10.1172/JCI8825

Single-cell expression profiling reveals dynamic flux of cardiac stromal, vascular and immune cells in health and injury

Publisher: eLife Sciences Publications, Ltd

Date: 26-03-2019

DOI: 10.7554/ELIFE.43882

Abstract: Besides cardiomyocytes (CM), the heart contains numerous interstitial cell types which play key roles in heart repair, regeneration and disease, including fibroblast, vascular and immune cells. However, a comprehensive understanding of this interactive cell community is lacking. We performed single-cell RNA-sequencing of the total non-CM fraction and enriched (Pdgfra-GFP+) fibroblast lineage cells from murine hearts at days 3 and 7 post-sham or myocardial infarction (MI) surgery. Clustering of ,000 single cells identified populations representing nine cell lineages, including a previously undescribed fibroblast lineage trajectory present in both sham and MI hearts leading to a uniquely activated cell state defined in part by a strong anti-WNT transcriptome signature. We also uncovered novel myofibroblast subtypes expressing either pro-fibrotic or anti-fibrotic signatures. Our data highlight non-linear dynamics in myeloid and fibroblast lineages after cardiac injury, and provide an entry point for deeper analysis of cardiac homeostasis, inflammation, fibrosis, repair and regeneration.

Nkx2-5 Represses Gata1 Gene Expression and Modulates the Cellular Fate of Cardiac Progenitors During Embryogenesis

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 19-04-2011

DOI: 10.1161/CIRCULATIONAHA.110.008185

Abstract: Recent studies suggest that the hematopoietic and cardiac lineages have close ontogenic origins, and that an early mesodermal cell population has the potential to differentiate into both lineages. Studies also suggest that specification of these lineages is inversely regulated. However, the transcriptional networks that govern the cell fate specification of these progenitors are incompletely defined. Here, we show that Nkx2-5 regulates the hematopoietic/erythroid fate of the mesoderm precursors early during cardiac morphogenesis. Using transgenic technologies to isolate Nkx2-5 expressing cells, we observed an induction of the erythroid molecular program, including Gata1 , in the Nkx2-5 –null embryos. We further observed that overexpression of Nkx2-5 with an Nkx2-5–inducible embryonic stem cell system significantly repressed Gata1 gene expression and suppressed the hematopoietic/erythroid potential, but not the endothelial potential, of the embryonic stem cells. This suppression was cell-autonomous, and was partially rescued by overexpressing Gata1. In addition, we demonstrated that Nkx2-5 binds to the Gata1 gene enhancer and represses the transcriptional activity of the Gata1 gene. Our results demonstrate that the hematopoietic/erythroid cell fate is suppressed via Nkx2-5 during mesodermal fate determination, and that the Gata1 gene is one of the targets that are suppressed by Nkx2-5.

An endothelial contribution to coronary vessels

Publisher: Elsevier BV

Date: 11-2012

DOI: 10.1016/J.CELL.2012.11.004

Abstract: Despite the profound impact of coronary artery disease on human health, the origins of the coronary blood vessels are poorly understood. Wu et al. use imaging and genetic techniques to show that the endocardium contributes to the coronary vessels and that the coronary arteries and veins have multilineage origins.

Skeletal muscle hypertrophy is mediated by a Ca2+-dependent calcineurin signalling pathway

Publisher: Springer Science and Business Media LLC

Date: 08-1999

DOI: 10.1038/23054

Abstract: Skeletal muscle hypertrophy and regeneration are important adaptive responses to both physical activity and pathological stimuli. Failure to maintain these processes underlies the loss of skeletal muscle mass and strength that occurs with ageing and in myopathies. Here we show that stable expression of a gene encoding insulin-like growth factor 1 (IGF-1) in C2C12 skeletal muscle cells, or treatment of these cells with recombinant IGF-1 or with insulin and dexamethasone, results in hypertrophy of differentiated myotubes and a switch to glycolytic metabolism. Treatment with IGF-1 or insulin and dexamethasone mobilizes intracellular calcium, activates the Ca2+/calmodulin-dependent phosphatase calcineurin, and induces the nuclear translocation of the transcription factor NF-ATc1. Hypertrophy is suppressed by the calcineurin inhibitors cyclosporin A or FK506, but not by inhibitors of the MAP-kinase or phosphatidylinositol-3-OH kinase pathways. Injecting rat latissimus dorsi muscle with a plasmid encoding IGF-1 also activates calcineurin, mobilizes satellite cells and causes a switch to glycolytic metabolism. We propose that growth-factor-induced skeletal-muscle hypertrophy and changes in myofibre phenotype are mediated by calcium mobilization and are critically regulated by the calcineurin/NF-ATc1 signalling pathway.

Gene–environment interaction demonstrates the vulnerability of the embryonic heart

Publisher: Elsevier BV

Date: 07-2014

DOI: 10.1016/J.YDBIO.2014.03.005

PI(4,5)P ₂ -dependent regulation of endothelial tip cell specification contributes to angiogenesis

Publisher: American Association for the Advancement of Science (AAAS)

Date: 31-03-2023

DOI: 10.1126/SCIADV.ADD6911

Abstract: Dynamic positioning of endothelial tip and stalk cells, via the interplay between VEGFR2 and NOTCH signaling, is essential for angiogenesis. VEGFR2 activates PI3K, which phosphorylates PI(4,5)P 2 to PI(3,4,5)P 3 , activating AKT however, PI3K/AKT does not direct tip cell specification. We report that PI(4,5)P 2 hydrolysis by the phosphoinositide-5-phosphatase, INPP5K, contributes to angiogenesis. INPP5K ablation disrupted tip cell specification and impaired embryonic angiogenesis associated with enhanced DLL4/NOTCH signaling. INPP5K degraded a pool of PI(4,5)P 2 generated by PIP5K1C phosphorylation of PI(4)P in endothelial cells. INPP5K ablation increased PI(4,5)P 2 , thereby releasing β-catenin from the plasma membrane, and concurrently increased PI(3,4,5)P 3 -dependent AKT activation, conditions that licensed DLL4 / NOTCH transcription. Suppression of PI(4,5)P 2 in INPP5K -siRNA cells by PIP5K1C -siRNA, restored β-catenin membrane localization and normalized AKT signaling. Pharmacological NOTCH or AKT inhibition in vivo or genetic β-catenin attenuation rescued angiogenesis defects in INPP5K-null mice. Therefore, PI(4,5)P 2 is critical for β-catenin/DLL4/NOTCH signaling, which governs tip cell specification during angiogenesis.

Genetic networks governing heart development

Publisher: Cold Spring Harbor Laboratory

Date: 03-10-2014

DOI: 10.1101/CSHPERSPECT.A013839

Congenital heart disease: current knowledge about causes and inheritance.

Publisher: AMPCo

Date: 08-2012

DOI: 10.5694/MJA12.10811

Abstract: About 80% of congenital heart disease (CHD) is multifactorial and arises through various combinations of genetic and environmental contributors. About 20% of cases can be attributed to chromosomal anomalies, Mendelian syndromes, non-syndromal single gene disorders or teratogens. Down syndrome and velocardiofacial syndrome are the most commonly seen syndromes in patients with CHD. To date, more than 30 genes have been linked to non-syndromal forms of CHD. Their contribution to CHD remains unknown but is presumed to be relatively small. There is limited evidence for the contribution of specific environmental factors to CHD causation. However, folic acid supplementation in the pre- and peri-conception period, ensuring rubella vaccination has been completed before pregnancy, and maintaining good glycaemic control in mothers with diabetes may reduce the risk of CHD in infants. Recurrence risks vary between different types of non-syndromal CHD with multifactorial inheritance, and can be as high as 10% when two or more siblings are affected. Generally, the recurrence risk increases if a parent rather than a sibling is affected, particularly when the affected parent is the mother. In idualised recurrence risks can be generated for members of families affected by CHD after obtaining a detailed family history, including accurate cardiac diagnoses for all affected members. High-throughput genetic techniques can accelerate gene discovery and improve our ability to provide in idualised genetic counselling.

BMP/SMAD1 signaling sets a threshold for the left/right pathway in lateral plate mesoderm and limits availability of SMAD4

Publisher: Cold Spring Harbor Laboratory

Date: 11-2008

DOI: 10.1101/GAD.1682108

Abstract: Bistability in developmental pathways refers to the generation of binary outputs from graded or noisy inputs. Signaling thresholds are critical for bistability. Specification of the left/right (LR) axis in vertebrate embryos involves bistable expression of transforming growth factor β (TGFβ) member NODAL in the left lateral plate mesoderm (LPM) controlled by feed-forward and feedback loops. Here we provide evidence that bone morphogenetic protein (BMP)/SMAD1 signaling sets a repressive threshold in the LPM essential for the integrity of LR signaling. Conditional deletion of Smad1 in the LPM led to precocious and bilateral pathway activation. NODAL expression from both the left and right sides of the node contributed to bilateral activation, indicating sensitivity of mutant LPM to noisy input from the LR system. In vitro, BMP signaling inhibited NODAL pathway activation and formation of its downstream SMAD2/4–FOXH1 transcriptional complex. Activity was restored by overexpression of SMAD4 and in embryos, elevated SMAD4 in the right LPM robustly activated LR gene expression, an effect reversed by superactivated BMP signaling. We conclude that BMP/SMAD1 signaling sets a bilateral, repressive threshold for NODAL-dependent Nodal activation in LPM, limiting availability of SMAD4. This repressive threshold is essential for bistable output of the LR system.

Haemogenic endocardium contributes to transient definitive haematopoiesis

Publisher: Springer Science and Business Media LLC

Date: 05-03-2013

DOI: 10.1038/NCOMMS2569

Platelet-derived growth factor (PDGF) signaling directs cardiomyocyte movement toward the midline during heart tube assembly

Publisher: eLife Sciences Publications, Ltd

Date: 18-01-2017

DOI: 10.7554/ELIFE.21172

Abstract: Communication between neighboring tissues plays a central role in guiding organ morphogenesis. During heart tube assembly, interactions with the adjacent endoderm control the medial movement of cardiomyocytes, a process referred to as cardiac fusion. However, the molecular underpinnings of this endodermal-myocardial relationship remain unclear. Here, we show an essential role for platelet-derived growth factor receptor alpha (Pdgfra) in directing cardiac fusion. Mutation of pdgfra disrupts heart tube assembly in both zebrafish and mouse. Timelapse analysis of in idual cardiomyocyte trajectories reveals misdirected cells in zebrafish pdgfra mutants, suggesting that PDGF signaling steers cardiomyocytes toward the midline during cardiac fusion. Intriguingly, the ligand pdgfaa is expressed in the endoderm medial to the pdgfra-expressing myocardial precursors. Ectopic expression of pdgfaa interferes with cardiac fusion, consistent with an instructive role for PDGF signaling. Together, these data uncover a novel mechanism through which endodermal-myocardial communication can guide the cell movements that initiate cardiac morphogenesis.

Molecular analysis of PRKAG2, LAMP2, and NKX2‐5 genes in a cohort of 125 patients with accessory atrioventricular connection

Publisher: Wiley

Date: 16-06-2009

DOI: 10.1002/AJMG.A.32907

Nkx2.5 marks angioblasts that contribute to hemogenic endothelium of the endocardium and dorsal aorta

Publisher: eLife Sciences Publications, Ltd

Date: 08-03-2017

DOI: 10.7554/ELIFE.20994

Abstract: Novel regenerative therapies may stem from deeper understanding of the mechanisms governing cardiovascular lineage ersification. Using enhancer mapping and live imaging in avian embryos, and genetic lineage tracing in mice, we investigated the spatio-temporal dynamics of cardiovascular progenitor populations. We show that expression of the cardiac transcription factor Nkx2.5 marks a mesodermal population outside of the cardiac crescent in the extraembryonic and lateral plate mesoderm, with characteristics of hemogenic angioblasts. Extra-cardiac Nkx2.5 lineage progenitors migrate into the embryo and contribute to clusters of CD41+/CD45+ and RUNX1+ cells in the endocardium, the aorta-gonad-mesonephros region of the dorsal aorta and liver. We also demonstrated that ectopic expression of Nkx2.5 in chick embryos activates the hemoangiogenic gene expression program. Taken together, we identified a hemogenic angioblast cell lineage characterized by transient Nkx2.5 expression that contributes to hemogenic endothelium and endocardium, suggesting a novel role for Nkx2.5 in hemoangiogenic lineage specification and ersification.

Neuregulin 1 and Susceptibility to Schizophrenia

Publisher: Elsevier BV

Date: 10-2002

DOI: 10.1086/342734

Hop is an unusual homeobox gene that modulates cardiac development

Publisher: Elsevier BV

Date: 09-2002

DOI: 10.1016/S0092-8674(02)00932-7

Abstract: Hop is a small, ergent homeodomain protein that lacks certain conserved residues required for DNA binding. Hop gene expression initiates early in cardiogenesis and continues in cardiomyocytes throughout embryonic and postnatal development. Genetic and biochemical data indicate that Hop functions directly downstream of Nkx2-5. Inactivation of Hop in mice by homologous recombination results in a partially penetrant embryonic lethal phenotype with severe developmental cardiac defects involving the myocardium. Inhibition of Hop activity in zebrafish embryos likewise disrupts cardiac development and results in severely impaired cardiac function. Hop physically interacts with serum response factor (SRF) and inhibits activation of SRF-dependent transcription by inhibiting SRF binding to DNA. Hop encodes an unusual homeodomain protein that modulates SRF-dependent cardiac-specific gene expression and cardiac development.

Neuregulin 1 Sustains the Gene Regulatory Network in Both Trabecular and Nontrabecular Myocardium

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 17-09-2010

DOI: 10.1161/CIRCRESAHA.110.218693

Abstract: The cardiac gene regulatory network (GRN) is controlled by transcription factors and signaling inputs, but network logic in development and it unraveling in disease is poorly understood. In development, the membrane-tethered signaling ligand Neuregulin (Nrg)1, expressed in endocardium, is essential for ventricular morphogenesis. In adults, Nrg1 protects against heart failure and can induce cardiomyocytes to ide. To understand the role of Nrg1 in heart development through analysis of null and hypomorphic Nrg1 mutant mice. Chamber domains were correctly specified in Nrg1 mutants, although chamber-restricted genes Hand1 and Cited1 failed to be activated. The chamber GRN subsequently decayed with in idual genes exhibiting decay patterns unrelated to known patterning boundaries. Both trabecular and nontrabecular myocardium were affected. Network demise was spatiotemporally dynamic, the most sensitive region being the central part of the left ventricle, in which the GRN underwent complete collapse. Other regions were partially affected with graded sensitivity. In vitro, Nrg1 promoted phospho-Erk1/2–dependent transcription factor expression, cardiomyocyte maturation and cell cycle inhibition. We monitored cardiac pErk1/2 in embryos and found that expression was Nrg1-dependent and levels correlated with cardiac GRN sensitivity in mutants. The chamber GRN is fundamentally labile and dependent on signaling from extracardiac sources. Nrg1–ErbB1/4–Erk1/2 signaling critically sustains elements of the GRN in trabecular and nontrabecular myocardium, challenging our understanding of Nrg1 function. Transcriptional decay patterns induced by reduced Nrg1 suggest a novel mechanism for cardiac transcriptional regulation and dysfunction in disease, potentially linking biomechanical feedback to molecular pathways for growth and differentiation.

Long Noncoding RNAs in Cardiac Development and Pathophysiology

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 26-10-2012

DOI: 10.1161/CIRCRESAHA.112.268953

Abstract: Heart function requires sophisticated regulatory networks to orchestrate organ development, physiological responses, and environmental adaptation. Until recently, it was thought that these regulatory networks are composed solely of protein-mediated transcriptional control and signaling systems consequently, it was thought that cardiac disease involves perturbation of these systems. However, it is becoming evident that RNA, long considered to function primarily as the platform for protein production, may in fact play a major role in most, if not all, aspects of gene regulation, especially the epigenetic processes that underpin organogenesis. These include not only well-validated classes of regulatory RNAs, such as microRNAs, but also tens of thousands of long noncoding RNAs that are differentially expressed across the entire genome of humans and other animals. Here, we review this emerging landscape, summarizing what is known about their functions and their role in cardiac biology, and provide a toolkit to assist in exploring this previously hidden layer of gene regulation that may underpin heart adaptation and complex heart diseases.

Identification of clinically actionable variants from genome sequencing of families with congenital heart disease

Publisher: Elsevier BV

Date: 05-2019

DOI: 10.1038/S41436-018-0296-X

Molecular Pathway for the Localized Formation of the Sinoatrial Node

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 16-02-2007

DOI: 10.1161/01.RES.0000258019.74591.B3

Abstract: 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.

Prediction and validation of protein–protein interactors from genome-wide DNA-binding data using a knowledge-based machine-learning approach

Publisher: The Royal Society

Date: 09-2016

DOI: 10.1098/RSOB.160183

Abstract: The ability to accurately predict the DNA targets and interacting cofactors of transcriptional regulators from genome-wide data can significantly advance our understanding of gene regulatory networks. NKX2-5 is a homeodomain transcription factor that sits high in the cardiac gene regulatory network and is essential for normal heart development. We previously identified genomic targets for NKX2-5 in mouse HL-1 atrial cardiomyocytes using DNA-adenine methyltransferase identification (DamID). Here, we apply machine learning algorithms and propose a knowledge-based feature selection method for predicting NKX2-5 protein : protein interactions based on motif grammar in genome-wide DNA-binding data. We assessed model performance using leave-one-out cross-validation and a completely independent DamID experiment performed with replicates. In addition to identifying previously described NKX2-5-interacting proteins, including GATA, HAND and TBX family members, a number of novel interactors were identified, with direct protein : protein interactions between NKX2-5 and retinoid X receptor (RXR), paired-related homeobox (PRRX) and Ikaros zinc fingers (IKZF) validated using the yeast two-hybrid assay. We also found that the interaction of RXRα with NKX2-5 mutations found in congenital heart disease (Q187H, R189G and R190H) was altered. These findings highlight an intuitive approach to accessing protein–protein interaction information of transcription factors in DNA-binding experiments.

Expression of cardiac muscle markers in rat myocyte cell lines

Publisher: Springer Science and Business Media LLC

Date: 04-1996

DOI: 10.1007/BF00227884

Abstract: Baicalin, a natural bioactive flavonoid extracted from Human cementoblast cell line (HCEM) cells were cultured and treated with 0, 0.01, 0.1, or 1 μM of baicalin. The proliferative capacity of cultured HCEM cells was analyzed using bromodeoxyuridine immunoassay and cell counting. The baicalin effect on OPG and RANKL expression was determined using quantitative polymerase chain reaction (qPCR) and western blotting. Furthermore, OPG expression was measured in 1 μM baicalin-treated HCEM cells in the presence or absence of the Wnt signaling pathway inhibitor, Dickkopf (Dkk)-1, using qPCR and western blotting. The addition of 0.01, 0.1, and 1 μM of baicalin did not significantly change the proliferative capacity of cultured HCEM cells. Compared with the non-supplemented group, baicalin increased and suppressed OPG and RANKL gene and protein expression, respectively, in a concentration-dependent manner. OPG mRNA and protein expression levels were increased by 1 μM baicalin, which was suppressed by Dkk-1 addition. Baicalin enhanced OPG expression in HCEM cells through the Wnt/beta-catenin signaling pathway, which could contribute to periodontal tissue regeneration.

Perinatal angiogenesis from pre-existing coronary vessels via DLL4–NOTCH1 signalling

Publisher: Springer Science and Business Media LLC

Date: 09-2021

DOI: 10.1038/S41556-021-00747-1

Developmental paradigms in heart disease: insights from tinman

Publisher: Informa UK Limited

Date: 2002

DOI: 10.1080/ANN.34.3.148.156

Cardiac Repair with a Novel Population of Mesenchymal Stem Cells Resident in the Human Heart

Publisher: Oxford University Press (OUP)

Date: 29-07-2015

DOI: 10.1002/STEM.2101

Abstract: Cardiac resident stem cells (CRSCs) hold much promise to treat heart disease but this remains a controversial field. Here, we describe a novel population of CRSCs, which are positive for W8B2 antigen and were obtained from adult human atrial appendages. W8B2+ CRSCs exhibit a spindle-shaped morphology, are clonogenic and capable of self-renewal. W8B2+ CRSCs show high expression of mesenchymal but not hematopoietic nor endothelial markers. W8B2+ CRSCs expressed GATA4, HAND2, and TBX5, but not C-KIT, SCA-1, NKX2.5, PDGFRα, ISL1, or WT1. W8B2+ CRSCs can differentiate into cardiovascular lineages and secrete a range of cytokines implicated in angiogenesis, chemotaxis, inflammation, extracellular matrix remodeling, cell growth, and survival. In vitro, conditioned medium collected from W8B2+ CRSCs displayed prosurvival, proangiogenic, and promigratory effects on endothelial cells, superior to that of other adult stem cells tested, and additionally promoted survival and proliferation of neonatal rat cardiomyocytes. Intramyocardial transplantation of human W8B2+ CRSCs into immunocompromised rats 1 week after myocardial infarction markedly improved cardiac function (∼40% improvement in ejection fraction) and reduced fibrotic scar tissue 4 weeks after infarction. Hearts treated with W8B2+ CRSCs showed less adverse remodeling of the left ventricle, a greater number of proliferating cardiomyocytes (Ki67+cTnT+ cells) in the remote region, higher myocardial vascular density, and greater infiltration of CD163+ cells (a marker for M2 macrophages) into the border zone and scar regions. In summary, W8B2+ CRSCs are distinct from currently known CRSCs found in human hearts, and as such may be an ideal cell source to repair myocardial damage after infarction. Stem Cells 2015 :3100–3113

Normothermic Ex vivo perfusion provides superior organ preservation and enables viability assessment of hearts from DCD donors

Publisher: Elsevier BV

Date: 02-2015

DOI: 10.1111/AJT.12994

A Critical Time for Stem Cell Research in Australia

Publisher: Elsevier BV

Date: 02-2008

DOI: 10.1016/J.STEM.2008.01.019

Abstract: Australia's strong historical foundation in stem cell science was boosted in 2002 by the formation of a virtual government-supported institute now called the Australian Stem Cell Centre. However, the inevitable "brain drain" and government and community expectations of quick commercial gains and rapid cures necessitate careful reconsideration of the identity and future of stem cell science in Australia.

Hif-1a suppresses ROS-induced proliferation of cardiac fibroblasts following myocardial infarction

Publisher: Elsevier BV

Date: 02-2022

DOI: 10.1016/J.STEM.2021.10.009

Isolation of a genomal clone containing chicken histone genes

Publisher: Oxford University Press (OUP)

Date: 1979

DOI: 10.1093/NAR/7.7.1787

Abstract: We have used enriched chicken histone cDNA to select genomal clones from a chicken library. Because the cDNA probe also contained other sequences, a further screening of positive plagues with negative probes eliminated most non-histone gene clones. One 'positively-selected' genomal clone, lambda CH-01, hybridised with cloned sea-urchin histone genes and also detected histone genes in EcoRI-digested genomal sea-urchin DNA. Limited DNA sequencing of HaeIII fragments identified two sequences within the coding region of chicken histone H2A. A third fragment predicted an amino acid sequence with strong homology to an H1 histone sequence.

Conserved linkage of NK-2 homeobox gene pairs Nkx2-2/2-4 and Nkx2-1/2-9 in mammals

Publisher: Springer Science and Business Media LLC

Date: 06-2000

DOI: 10.1007/S003350010089

Quantitative trait and transcriptome analysis of genetic complexity underpinning cardiac interatrial septation in mice using an advanced intercross line

Publisher: eLife Sciences Publications, Ltd

Date: 05-06-2023

DOI: 10.7554/ELIFE.83606

Abstract: Unlike single-gene mutations leading to Mendelian conditions, common human diseases are likely to be emergent phenomena arising from multilayer, multiscale, and highly interconnected interactions. Atrial and ventricular septal defects are the most common forms of cardiac congenital anomalies in humans. Atrial septal defects (ASD) show an open communication between the left and right atria postnatally, potentially resulting in serious hemodynamic consequences if untreated. A milder form of atrial septal defect, patent foramen ovale (PFO), exists in about one-quarter of the human population, strongly associated with ischaemic stroke and migraine. The anatomic liabilities and genetic and molecular basis of atrial septal defects remain unclear. Here, we advance our previous analysis of atrial septal variation through quantitative trait locus (QTL) mapping of an advanced intercross line (AIL) established between the inbred QSi5 and 129T2/SvEms mouse strains, that show extremes of septal phenotypes. Analysis resolved 37 unique septal QTL with high overlap between QTL for distinct septal traits and PFO as a binary trait. Whole genome sequencing of parental strains and filtering identified predicted functional variants, including in known human congenital heart disease genes. Transcriptome analysis of developing septa revealed downregulation of networks involving ribosome, nucleosome, mitochondrial, and extracellular matrix biosynthesis in the 129T2/SvEms strain, potentially reflecting an essential role for growth and cellular maturation in septal development. Analysis of variant architecture across different gene features, including enhancers and promoters, provided evidence for the involvement of non-coding as well as protein-coding variants. Our study provides the first high-resolution picture of genetic complexity and network liability underlying common congenital heart disease, with relevance to human ASD and PFO.

The Murine Homeobox Genes Nkx2.3 and Nkx2.6 Are Located on Chromosomes 19 and 14, Respectively

Publisher: Elsevier BV

Date: 08-1994

DOI: 10.1006/GENO.1994.1444

Specialized Information Processing Deficits and Distinct Metabolomic Profiles Following TM-Domain Disruption of Nrg1

Publisher: Oxford University Press (OUP)

Date: 11-03-2017

DOI: 10.1093/SCHBUL/SBW189

Heart redevelopment

Publisher: Springer Science and Business Media LLC

Date: 09-2010

DOI: 10.1038/467039A

Homeodomain factor Nkx2-3 controls regional expression of leukocyte homing coreceptor MAdCAM-1 in specialized endothelial cells of the viscera.

Publisher: Elsevier BV

Date: 08-2000

DOI: 10.1006/DBIO.2000.9749

Assignment of the Human Helix-Loop-Helix Transcription Factor Gene Musculin/Activated B-Cell Factor-1 (MSC) to Chromosome 8q21 and Its Mouse Homologue (Msc) to the Proximal Region of Chromosome 1

Publisher: Elsevier BV

Date: 04-1999

DOI: 10.1006/GENO.1999.5764

Gene-environment interaction impacts on heart development and embryo survival.

Publisher: The Company of Biologists

Date: 15-02-2019

DOI: 10.1242/DEV.172957

Abstract: Congenital heart disease (CHD) is the most common type of birth defect. In recent years, research has focussed on identifying the genetic causes of CHD. However, only a minority of CHD cases can be attributed to single gene mutations. In addition, studies have identified different environmental stressors that promote CHD, but the additive effect of genetic susceptibility and environmental factors is poorly understood. In this context, we have investigated the effects of short-term gestational hypoxia on mouse embryos genetically predisposed to heart defects. Exposure of mouse embryos heterozygous for Tbx1 or Fgfr1/Fgfr2 to hypoxia in utero increased the incidence and severity of heart defects while Nkx2-5+/− embryos died within 2 days of hypoxic exposure. We identified the molecular consequences of the interaction between Nkx2-5 and short-term gestational hypoxia, which suggest that reduced Nkx2-5 expression and a prolonged hypoxia-inducible factor 1α response together precipitate embryo death. Our study provides insight into the causes of embryo loss and variable penetrance of monogenic CHD, and raises the possibility that cases of foetal death and CHD in humans could be caused by similar gene-environment interactions.

Non-tandem arrangement and divergent transcription of chicken histone genes

Publisher: Springer Science and Business Media LLC

Date: 11-1981

DOI: 10.1038/294049A0

Repair of mirage III aircraft using the BFRP crack-patching technique

Publisher: Elsevier BV

Date: 10-1984

DOI: 10.1016/0167-8442(84)90035-1

Disruption of thermal nociceptive behaviour in mice mutant for the schizophrenia-associated genes NRG1, COMT and DISC1

Publisher: Elsevier BV

Date: 08-2010

DOI: 10.1016/J.BRAINRES.2010.06.027

Abstract: Abnormalities in pain perception, especially altered warmth and heat pain sensitivity, have been reported in schizophrenia. Therefore, genes associated with schizophrenia, including neuregulin-1 (NRG1), catechol-O-methyltranferase (COMT) and disrupted-in-schizophrenia-1 (DISC1), may play a role in modulating the physiological and psychological effects of pain stimuli in such patients. Thermal pain sensitivity was assessed in NRG1, COMT and DISC1 mutant mice, and the anti-nociceptive effects of acute Delta(9)-tetrahydrocannabinol (THC) were compared in NRG1 and COMT mutants. At baseline, deletion of NRG1 and DISC1 each reduced thermal pain sensitivity, while deletion of COMT increased pain sensitivity. Neither NRG1 nor COMT deletion altered the anti-nociceptive effects of acute systemic THC (8.0mg/kg). These results indicate a differential contribution of NRG1 and DISC1 vis-à-vis COMT to the processing of thermal nociceptive stimuli and extend their phenotypic relationship to psychotic illness.

Retinal changes in an ATP-induced model of retinal degeneration

Publisher: Frontiers Media SA

Date: 29-04-2016

DOI: 10.3389/FNANA.2016.00046

Pathophysiological Trends During Withdrawal of Life Support

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 12-2016

DOI: 10.1097/TP.0000000000001396

Myogenic and morphogenetic defects in the heart tubes of murine embryos lacking the homeo box gene Nkx2-5.

Publisher: Cold Spring Harbor Laboratory

Date: 07-1995

DOI: 10.1101/GAD.9.13.1654

Abstract: The murine homeo box gene Nkx2-5 is expressed in precardiac mesoderm and in the myocardium of embryonic and fetal hearts. Targeted interruption of Nkx2-5 resulted in abnormal heart morphogenesis, growth retardation and embryonic lethality at approximately 9-10 days postcoitum (p.c.). Heart tube formation occurred normally in mutant embryos, but looping morphogenesis, a critical determinant of heart form, was not initiated at the linear heart tube stage (8.25-8.5 days p.c.). Commitment to the cardiac muscle lineage, expression of most myofilament genes and myofibrillogenesis were not compromised. However, the myosin light-chain 2V gene (MLC2V) was not expressed in mutant hearts nor in mutant ES cell-derived cardiocytes. MLC2V expression normally occurs only in ventricular cells and is the earliest known molecular marker of ventricular differentiation. The regional expression in mutant hearts of two other ventricular markers, myosin heavy-chain beta and cyclin D2, indicated that not all ventricle-specific gene expression is dependent on Nkx2-5. The data demonstrate that Nkx2-5 is essential for normal heart morphogenesis, myogenesis, and function. Furthermore, this gene is a component of a genetic pathway required for myogenic specialization of the ventricles.

Chamber Formation and Morphogenesis in the Developing Mammalian Heart

Publisher: Elsevier BV

Date: 07-2000

DOI: 10.1006/DBIO.2000.9753

NKX2-5 mutations causative for congenital heart disease retain functionality and are directed to hundreds of targets

Publisher: eLife Sciences Publications, Ltd

Date: 06-07-2015

DOI: 10.7554/ELIFE.06942

Abstract: We take a functional genomics approach to congenital heart disease mechanism. We used DamID to establish a robust set of target genes for NKX2-5 wild type and disease associated NKX2-5 mutations to model loss-of-function in gene regulatory networks. NKX2-5 mutants, including those with a crippled homeodomain, bound hundreds of targets including NKX2-5 wild type targets and a unique set of "off-targets", and retained partial functionality. NKXΔHD, which lacks the homeodomain completely, could heterodimerize with NKX2-5 wild type and its cofactors, including E26 transformation-specific (ETS) family members, through a tyrosine-rich homophilic interaction domain (YRD). Off-targets of NKX2-5 mutants, but not those of an NKX2-5 YRD mutant, showed overrepresentation of ETS binding sites and were occupied by ETS proteins, as determined by DamID. Analysis of kernel transcription factor and ETS targets show that ETS proteins are highly embedded within the cardiac gene regulatory network. Our study reveals binding and activities of NKX2-5 mutations on WT target and off-targets, guided by interactions with their normal cardiac and general cofactors, and suggest a novel type of gain-of-function in congenital heart disease.

ERBB2 triggers mammalian heart regeneration by promoting cardiomyocyte dedifferentiation and proliferation

Publisher: Springer Science and Business Media LLC

Date: 06-04-2015

DOI: 10.1038/NCB3149

Abstract: The murine neonatal heart can regenerate after injury through cardiomyocyte (CM) proliferation, although this capacity markedly diminishes after the first week of life. Neuregulin-1 (NRG1) administration has been proposed as a strategy to promote cardiac regeneration. Here, using loss- and gain-of-function genetic tools, we explore the role of the NRG1 co-receptor ERBB2 in cardiac regeneration. NRG1-induced CM proliferation diminished one week after birth owing to a reduction in ERBB2 expression. CM-specific Erbb2 knockout revealed that ERBB2 is required for CM proliferation at embryonic/neonatal stages. Induction of a constitutively active ERBB2 (caERBB2) in neonatal, juvenile and adult CMs resulted in cardiomegaly, characterized by extensive CM hypertrophy, dedifferentiation and proliferation, differentially mediated by ERK, AKT and GSK3β/β-catenin signalling pathways. Transient induction of caERBB2 following myocardial infarction triggered CM dedifferentiation and proliferation followed by redifferentiation and regeneration. Thus, ERBB2 is both necessary for CM proliferation and sufficient to reactivate postnatal CM proliferative and regenerative potentials.

Hlx homeo box gene is essential for an inductive tissue interaction that drives expansion of embryonic liver and gut.

Publisher: Cold Spring Harbor Laboratory

Date: 1996

DOI: 10.1101/GAD.10.1.70

Abstract: The ergent murine homeo box gene Hlx is expressed in restricted hematopoietic cell types and, during embryogenesis, prominently in visceral mesenchyme of the developing liver, gall bladder, and gut. Targeted disruption of the gene has now established that it plays a key role in visceral organogenesis. Embryos homozygous for the mutation died around embryonic day 15 with anemia and severe hypoplasia of the liver and gut. Liver ontogeny commenced normally with formation of the liver erticulum and differentiation of hepatocytes, but the organ failed to expand and reached only 3% of normal size. The apparent liver hypoplasia was not associated with a notable increase in apoptotic cells. Gut development also began normally, but the intestines failed to undergo extensive elongation and looping and reached only a quarter of normal length. The anemia resulted from a deficiency in the fetal form of hematopoiesis, which occurs in the liver, but no intrinsic defect in Hlx-/- hematopoietic cells was observed in vitro, and liver-derived Hlx-/- hematopoietic stem cells that were transplanted to irradiated normal mice could fully reconstitute hematopoiesis. The impaired fetal hematopoiesis therefore reflects insufficient support function provided by the minute liver. Hlx is normally expressed in visceral mesenchyme lying adjacent to the developing liver and gut epithelia affected by the mutation, but not in the epithelia themselves. Hence, Hlx regulates a mesenchymal-epithelial interaction that drives a vital growth phase in visceral organogenesis. Moreover, because mutation of Hlx blocked liver growth but not its specification, early morphogenesis, or differentiation, development of this organ appears to occur by step-wise inductive interactions under separate genetic control.

The Combinatorial Activities of Nkx2.5 and dHAND Are Essential for Cardiac Ventricle Formation

Publisher: Elsevier BV

Date: 11-2001

DOI: 10.1006/DBIO.2001.0417

Independently evolving chicken histone H2B genes: identification of a ubiquitous H2B-specific 5′ element

Publisher: Oxford University Press (OUP)

Date: 1982

DOI: 10.1093/NAR/10.23.7851

Abstract: The DNA sequence of two chicken histone H2B genes has been determined. Both genes code for the same H2B subtype. Except for conserved "promoter" elements, the sequences 5' to the protein coding regions are completely ergent, indicating that the genes are distantly related and are not evolving in concert. This presents an ideal situation for sequence comparisons. We have discovered a 13 bp, H2B specific homology block, 5' CTCATTTGCATAC 3' located close to the "TATA box". This motif is conserved in all H2B gene leader regions so far sequenced. One of the H2B genes is closely linked, in a ergent arrangement, to an H2A gene, and sequence data suggests that the linked genes share promoter elements.

Chamber Formation and Morphogenesis in the Developing Mammalian Heart

Publisher: Elsevier BV

Date: 09-2000

DOI: 10.1006/DBIO.2000.9859

Characterization of Pitx2c expression in the mouse heart using a reporter transgene

Publisher: Wiley

Date: 18-11-2010

DOI: 10.1002/DVDY.22492

Abstract: To aid in detection and tracking of cells targeted by the left-right (LR) pathway in the heart throughout morphogenesis, expression from a Pitx2c-lacZ transgene (P2Ztg) was analysed in detail. β-galactosidase expression from P2Ztg was robust, allowing reliable visualisation of low-level Pitx2c expression, and was virtually entirely dependent upon NODAL signalling in the heart. P2Ztg showed expression in trabecular and septal, as well as non-trabecular, myocardium, and a strong expression bias in myocardium associated with in idual endocardial cushions of the atrioventricular canal and outflow tract, which are essential for cardiac septation. Expression on the ventral surface of the outflow tract evolved to a specific stripe that could be used to track the future aorta during outflow tract spiralling and remodelling. Our data show that the P2Ztg transgene is a useful resource for detection of molecular disturbances in the LR cascade, as well as morphogenetic defects associated with other cardiac congenital disorders.

Nkx2-5+Islet1+ Mesenchymal Precursors Generate Distinct Spleen Stromal Cell Subsets and Participate in Restoring Stromal Network Integrity

Publisher: Elsevier BV

Date: 04-2013

DOI: 10.1016/J.IMMUNI.2012.12.005

Introduction to the Special Issue on Heart Regeneration and Rejuvenation

Publisher: Elsevier BV

Date: 11-2014

DOI: 10.1016/J.SCR.2014.10.003

Schizophrenia‐related endophenotypes in heterozygous neuregulin‐1 ‘knockout’ mice

Publisher: Wiley

Date: 2010

DOI: 10.1111/J.1460-9568.2009.07069.X

Abstract: Neuregulin-1 (NRG1) has been shown to play a role in glutamatergic neurotransmission and is a risk gene for schizophrenia, in which there is evidence for hypoglutamatergic function. Sensitivity to the behavioural effects of the psychotomimetic N-methyl-D-aspartate receptor antagonists MK-801 and phencyclidine (PCP) was examined in mutant mice with heterozygous deletion of NRG1. Social behaviour (sociability, social novelty preference and dyadic interaction), together with exploratory activity, was assessed following acute or subchronic administration of MK-801 (0.1 and 0.2 mg/kg) or PCP (5 mg/kg). In untreated NRG1 mutants, levels of glutamate, N-acetylaspartate and GABA were determined using high-performance liquid chromatography and regional brain volumes were assessed using magnetic resonance imaging at 7T. NRG1 mutants, particularly males, displayed decreased responsivity to the locomotor-activating effects of acute PCP. Subchronic MK-801 and PCP disrupted sociability and social novelty preference in mutants and wildtypes and reversed the increase in both exploratory activity and social dominance-related behaviours observed in vehicle-treated mutants. No phenotypic differences were demonstrated in N-acetylaspartate, glutamate or GABA levels. The total ventricular and olfactory bulb volume was decreased in mutants. These data indicate a subtle role for NRG1 in modulating several schizophrenia-relevant processes including the effects of psychotomimetic N-methyl-D-aspartate receptor antagonists.

Five years of Stem Cell Research — Reflections on progress in the field

Publisher: Elsevier BV

Date: 11-2012

DOI: 10.1016/J.SCR.2012.10.001

Epithelial to mesenchymal transition as a portal to stem cell characters embedded in gene networks

Publisher: Wiley

Date: 18-09-2012

DOI: 10.1002/BIES.201200089

Abstract: Cells can transit between a range of stable epithelial and mesenchymal states and this has allowed the evolution of complex body forms. Epithelial to mesenchymal transition (EMT) and its reverse, mesenchymal to epithelial transition (MET), occur sequentially in development and organogenesis. EMT often accompanies transitions between stem-like cells and their more differentiated progeny, as occurs at gastrulation, although the relevance of this had not been clarified. New findings from the cancer and cell reprogramming fields suggest that EMT and MET can act as essential portals to stem cell character. Here, we review these findings in the broader context of EMT and MET with emphasis on stem cell biology. Using the heart as an ex le, we also explore the potential role of EMT/MET in organ regeneration. Understanding EMT and MET at a network level will give us new tools to probe stem cell character and enhance tissue repair.

Antisense‐mediated exon skipping: a therapeutic strategy for titin‐based dilated cardiomyopathy

Publisher: EMBO

Date: 10-03-2015

DOI: 10.15252/EMMM.201505047

Abstract: Frameshift mutations in the TTN gene encoding titin are a major cause for inherited forms of dilated cardiomyopathy ( DCM ), a heart disease characterized by ventricular dilatation, systolic dysfunction, and progressive heart failure. To date, there are no specific treatment options for DCM patients but heart transplantation. Here, we show the beneficial potential of reframing titin transcripts by antisense oligonucleotide ( AON )‐mediated exon skipping in human and murine models of DCM carrying a previously identified autosomal‐dominant frameshift mutation in titin exon 326. Correction of TTN reading frame in patient‐specific cardiomyocytes derived from induced pluripotent stem cells rescued defective myofibril assembly and stability and normalized the sarcomeric protein expression. AON treatment in Ttn knock‐in mice improved sarcomere formation and contractile performance in homozygous embryos and prevented the development of the DCM phenotype in heterozygous animals. These results demonstrate that disruption of the titin reading frame due to a truncating DCM mutation can be restored by exon skipping in both patient cardiomyocytes in vitro and mouse heart in vivo , indicating RNA ‐based strategies as a potential treatment option for DCM .

Bioengineered FSTL1 Patches Restore Cardiac Function Following Myocardial Infarction

Publisher: Elsevier BV

Date: 12-2015

DOI: 10.1016/J.MOLMED.2015.10.006

Abstract: Improving the limited ability of the heart to regenerate after infarction is crucial. Researchers now demonstrate that delivery of follistatin-like 1 (FSTL1) into injured hearts via collagen patches stimulates cardiomyocyte proliferation and cardiac functional recovery. These findings highlight the epicardium as a source of novel regenerative factors and biomimetic nanomaterials in cardiac translational medicine.

Author response: Platelet-derived growth factor (PDGF) signaling directs cardiomyocyte movement toward the midline during heart tube assembly

Publisher: eLife Sciences Publications, Ltd

Date: 22-12-2016

DOI: 10.7554/ELIFE.21172.028

Selective Inhibition of Human Group IIA-secreted Phospholipase A2 (hGIIA) Signaling Reveals Arachidonic Acid Metabolism Is Associated with Colocalization of hGIIA to Vimentin in Rheumatoid Synoviocyte

Publisher: Elsevier BV

Date: 05-2013

DOI: 10.1074/JBC.M112.397893

Rotary ATPases: models, machine elements and technical specifications.

Publisher: Informa UK Limited

Date: 2013

DOI: 10.4161/BIOA.23301

Localized Maternal mRNAs in Xenopus laevis Eggs

Publisher: Cold Spring Harbor Laboratory

Date: 1985

DOI: 10.1101/SQB.1985.050.01.005

Abstract: In the title compound, C(14)H(18)N(2)O, the benzimidazole grouping is close to planar, with a maximum deviation of 0.042 Å the six-membered non-aromatic ring adopts an envelope conformation. In the crystal structure, mol-ecules are linked into infinite sheets lying parallel to the bc plane by O-H⋯N and C-H⋯O hydrogen bonds.

Developmental origins and lineage descendants of endogenous adult cardiac progenitor cells

Publisher: Elsevier BV

Date: 11-2014

DOI: 10.1016/J.SCR.2014.09.008

Abstract: Mammalian hearts carry a number of primitive stem cell-like populations, although the magnitude of their contribution to tissue homeostasis and repair remains controversial. Recent CRE recombinase-based lineage tracing experiments suggest only a minor contribution to the formation of new cardiomyocytes from such cells, albeit one that might be augmented therapeutically. As the field explores clinical translation of cardiac stem cells, it will be important to understand the biology of these cells in great detail. In this review we document the various reported stem and progenitor cell populations in mammalian hearts and discuss the current state of knowledge on their origins and lineage capabilities.

Erratum to: ‘CompGO: an R package for comparing and visualizing Gene Ontology enrichment differences between DNA binding experiments’

Publisher: Springer Science and Business Media LLC

Date: 25-04-2016

DOI: 10.1186/S12859-016-1048-Z

Phenotypic effects of repeated psychosocial stress during adolescence in mice mutant for the schizophrenia risk gene neuregulin-1: A putative model of gene × environment interaction

Publisher: Elsevier BV

Date: 05-2012

DOI: 10.1016/J.BBI.2012.02.010

Abstract: There is a paucity of animal models by which the contributions of environmental and genetic factors to the pathobiology of psychosis can be investigated. This study examined the in idual and combined effects of chronic social stress during adolescence and deletion of the schizophrenia risk gene neuregulin-1 (NRG1) on adult mouse phenotype. Mice were exposed to repeated social defeat stress during adolescence and assessed for exploratory behaviour, working memory, sucrose preference, social behaviour and prepulse inhibition in adulthood. Thereafter, in vitro cytokine responses to mitogen stimulation and corticosterone inhibition were assayed in spleen cells, with measurement of cytokine and brain-derived neurotrophic factor (BDNF) mRNA in frontal cortex, hippoc us and striatum. NRG1 mutants exhibited hyperactivity, decreased anxiety, impaired sensorimotor gating and reduced preference for social novelty. The effects of stress on exploratory/anxiety-related parameters, spatial working memory, sucrose preference and basal cytokine levels were modified by NRG1 deletion. Stress also exerted varied effect on spleen cytokine response to concanavalin A and brain cytokine and BDNF mRNA expression in NRG1 mutants. The experience of psychosocial stress during adolescence may trigger further pathobiological features that contribute to the development of schizophrenia, particularly in those with underlying NRG1 gene abnormalities. This model elaborates the importance of gene × environment interactions in the etiology of schizophrenia.

Association of the PHACTR1/EDN1 Genetic Locus With Spontaneous Coronary Artery Dissection

Publisher: Elsevier BV

Date: 2019

DOI: 10.1016/J.JACC.2018.09.085

Abstract: Spontaneous coronary artery dissection (SCAD) is an increasingly recognized cause of acute coronary syndromes (ACS) afflicting predominantly younger to middle-aged women. Observational studies have reported a high prevalence of extracoronary vascular anomalies, especially fibromuscular dysplasia (FMD) and a low prevalence of coincidental cases of atherosclerosis. PHACTR1/EDN1 is a genetic risk locus for several vascular diseases, including FMD and coronary artery disease, with the putative causal noncoding variant at the rs9349379 locus acting as a potential enhancer for the endothelin-1 (EDN1) gene. This study sought to test the association between the rs9349379 genotype and SCAD. Results from case control studies from France, United Kingdom, United States, and Australia were analyzed to test the association with SCAD risk, including age at first event, pregnancy-associated SCAD (P-SCAD), and recurrent SCAD. The previously reported risk allele for FMD (rs9349379-A) was associated with a higher risk of SCAD in all studies. In a meta-analysis of 1,055 SCAD patients and 7,190 controls, the odds ratio (OR) was 1.67 (95% confidence interval [CI]: 1.50 to 1.86) per copy of rs9349379-A. In a subset of 491 SCAD patients, the OR estimate was found to be higher for the association with SCAD in patients without FMD (OR: 1.89 95% CI: 1.53 to 2.33) than in SCAD cases with FMD (OR: 1.60 95% CI: 1.28 to 1.99). There was no effect of genotype on age at first event, P-SCAD, or recurrence. The first genetic risk factor for SCAD was identified in the largest study conducted to date for this condition. This genetic link may contribute to the clinical overlap between SCAD and FMD.

Sexually dimorphic changes in the exploratory and habituation profiles of heterozygous neuregulin-1 knockout mice

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 23-01-2006

DOI: 10.1097/01.WNR.0000192738.31029.0A

Abstract: The neuregulin-1 gene is widely expressed in the central nervous system and is associated with increased risk for schizophrenia. Using an ethologically based approach, the phenotype of neuregulin-1 heterozygous knockout mice was examined by revealing the in idual elements of behaviour in the murine repertoire over the prolonged course of interaction with the environment. During initial exploration, neuregulin-1 mutants displayed a phenotype characterized by increases in locomotion and rearing free, with sex-specific alterations in sifting and grooming. Over subsequent habituation, certain initial effects endured while new phenotypic effects emerged, some of which were again sex-specific. These studies elaborate a pleiotropic role of neuregulin-1 in development, plasticity and function, including sexual dimorphism, by defining the elemental, temporal and sex-specific characteristics of the neuregulin-1 mutant ethogram.

XNKX-2.5, A XENOPUS GENE-RELATED TO NKX-2.5 AND TINMAN - EVIDENCE FOR A CONSERVED ROLE IN CARDIAC DEVELOPMENT

Publisher: Elsevier BV

Date: 03-1994

DOI: 10.1006/DBIO.1994.1089

Abstract: We have isolated a Xenopus homeodomain sequence, XNkx-2.5, which shows significant similarity to mouse Nkx-2.5 and to the Drosophila tinman gene product. In Drosophila, tinman is required for formation of the heart and visceral mesoderm structures. In situ hybridization studies show that XNkx-2.5 is expressed in the heart region during early Xenopus development and later is also expressed in gut tissue. The observed similarity of sequences and expression patterns suggests that the regulatory mechanisms underlying heart formation may be conserved between distant species.

Endothelial to Mesenchymal Transition in Cardiovascular Disease

Publisher: Elsevier BV

Date: 2019

DOI: 10.1016/J.JACC.2018.09.089

Cancer Incidence among Australian Nuclear Industry Workers

Publisher: Wiley

Date: 09-2006

DOI: 10.1539/JOH.48.358

Abstract: To assess whether workers at Lucas Heights Science and Technology Centre (LHSTC) had different levels of cancer incidence from the New South Wales (NSW) population in Australia. A retrospective cohort study was undertaken at LHSTC. Data on 7,076 workers employed between 1957-98 were abstracted from personnel, dosimetry, and medical files. An inception cohort was defined which included 4,523 workers in employment between 1972-96 to examine cancer incidence. Cancer registrations in the inception cohort were identified to 1996 through electronic linkage of records with the NSW and the Australian national registers of cancer incidence. All-cancer incidence in workers at LHSTC was 15% below the NSW rates [SIR=0.85 95% CI=(0.75, 0.95)]. Of 37 specific cancers and groups of cancers examined, statistically significant excesses relative to NSW rates were observed only for pleural cancer incidence [SIR=17.71 95%=(7.96, 39.43)], and for incidence of cancer of the small intestine [SIR=4.34 95% CI=(1.40, 13.46)]. This study gives little evidence of an increased risk of cancers associated with radiation exposure in a cohort of nuclear workers in Australia. The observed increase in the risk of cancer of the pleura was probably due to unmeasured exposures, given the lack of an established association with radiation exposure, and the strong link to asbestos exposure. Findings for cancers of the small intestine were based on small numbers and were likely to be due to chance.

Defining the earliest step of cardiovascular progenitor specification during embryonic stem cell differentiation

Publisher: Rockefeller University Press

Date: 07-03-2011

DOI: 10.1083/JCB.201007063

Abstract: During embryonic development and embryonic stem cell (ESC) differentiation, the different cell lineages of the mature heart arise from two types of multipotent cardiovascular progenitors (MCPs), the first and second heart fields. A key question is whether these two MCP populations arise from differentiation of a common progenitor. In this paper, we engineered Mesp1–green fluorescent protein (GFP) ESCs to isolate early MCPs during ESC differentiation. Mesp1-GFP cells are strongly enriched for MCPs, presenting the ability to differentiate into multiple cardiovascular lineages from both heart fields in vitro and in vivo. Transcriptional profiling of Mesp1-GFP cells uncovered cell surface markers expressed by MCPs allowing their prospective isolation. Mesp1 is required for MCP specification and the expression of key cardiovascular transcription factors. Isl1 is expressed in a subset of early Mesp1-expressing cells independently of Mesp1 and acts together with Mesp1 to promote cardiovascular differentiation. Our study identifies the early MCPs residing at the top of the cellular hierarchy of cardiovascular lineages during ESC differentiation.

Patterning the vertebrate heart

Publisher: Springer Science and Business Media LLC

Date: 07-2002

DOI: 10.1038/NRG843

Cardiovascular Regenerative Medicine: Digging In for the Long Haul

Publisher: Elsevier BV

Date: 12-2007

DOI: 10.1016/J.STEM.2007.11.011

Abstract: Cardiovascular cell-based regenerative medicine has enjoyed a brief but exciting history. In little over a decade, multiple hypotheses have risen and fallen, and this work has now triggered a critical reconsideration of several long-held cardiovascular paradigms. These and other issues were the focus of the second Symposium on Cardiovascular Regenerative Medicine, recently held at the NIH-NHLBI in Bethesda, MD, USA. The meeting served to showcase some of the highlights of the past decade but, at the same time, sharply underlined the enormity of the task ahead. Collectively, a sense emerged that researchers in this field are "digging in for the long haul."

Cardiac looping — an uneasy deal with laterality

Publisher: Elsevier BV

Date: 02-1998

DOI: 10.1006/SCDB.1997.0188

Abstract: Cardiac looping is a key morphogenetic event in vertebrate heart development. In a complex progression, the linear heart tube adopts a sweeping rightward curvature, establishing an architecture for the multichambered organ it soon becomes. Looping is an asymmetric event, its rightward direction being determined by an embryonic left-right axial pathway. The dextral loop has been a vehicle for development of heart complexity during evolution, to the extent that in higher vertebrates and humans, heart structure and function is extremely susceptible to perturbations of laterality, which may be common. The first clues to genetic control of laterality and looping are now emerging. Key issues to resolve include the origin of laterality information in the embryo, the exact nature of the effector molecules that link laterality to looping and the intrinsic mechanisms that drive the looping process itself.

Structural and functional characterization of the mouse Hlx homeobox gene

Publisher: Springer Science and Business Media LLC

Date: 10-2000

DOI: 10.1007/S003350010179

Abstract: Hlx is a mesenchymally expressed homeobox transcription factor gene that is essential for normal intestinal and hepatic development in the mouse. Here we report further characterization of the mouse Hlx gene, including an additional 3.7 kb of 5' sequence as well as the sequence of the three introns. Comparison of the sequence of the mouse Hlx gene 5' to the coding region with that of the human gene revealed multiple regions of high conservation. Neither the mouse nor the human gene contained a TATA box, and ribonuclease protection studies defined heterogeneous transcription start sites for the mouse gene. A number of consensus transcription factor binding sites were conserved between the mouse and human Hlx genes both within and outside of the highly conserved regions. Reporter constructs containing 4.2 or 1.4 kb of mouse 5' sequence showed active expression in cell lines that express Hlx. Further characterization of the mouse Hlx gene will provide insight into the developmental regulation of the mouse digestive system.

Functional Characterization of a Novel Mutation in NKX2-5 Associated With Congenital Heart Disease and Adult-Onset Cardiomyopathy

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 06-2013

DOI: 10.1161/CIRCGENETICS.113.000057

Abstract: The transcription factor NKX2-5 is crucial for heart development, and mutations in this gene have been implicated in erse congenital heart diseases and conduction defects in mouse models and humans. Whether NKX2-5 mutations have a role in adult-onset heart disease is unknown. Mutation screening was performed in 220 probands with adult-onset dilated cardiomyopathy. Six NKX2-5 coding sequence variants were identified, including 3 nonsynonymous variants. A novel heterozygous mutation, I184M, located within the NKX2-5 homeodomain, was identified in 1 family. A subset of family members had congenital heart disease, but there was an unexpectedly high prevalence of dilated cardiomyopathy. Functional analysis of I184M in vitro demonstrated a striking increase in protein expression when transfected into COS-7 cells or HL-1 cardiomyocytes because of reduced degradation by the Ubiquitin-proteasome system. In functional assays, DNA-binding activity of I184M was reduced, resulting in impaired activation of target genes despite increased expression levels of mutant protein. Certain NKX2-5 homeodomain mutations show abnormal protein degradation via the Ubiquitin-proteasome system and partially impaired transcriptional activity. We propose that this class of mutation can impair heart development and mature heart function and contribute to NKX2-5 –related cardiomyopathies with graded severity.

Homeodomain factor Nkx2-5 in heart development and disease

Publisher: Cold Spring Harbor Laboratory

Date: 2002

DOI: 10.1101/SQB.2002.67.107

Heart Development and Regeneration

Publisher: Elsevier

Date: 2010

DOI: 10.1016/C2009-1-62569-7

NK-2Homeobox Genes and Heart Development

Publisher: Elsevier BV

Date: 09-1996

DOI: 10.1006/DBIO.1996.0212

Abstract: Analysis of the phylogenetically ancient NK-2 class of homeobox genes has opened up an entirely new approach to molecular, genetic, and biochemical analysis of early heart development. The Drosophila NK-2 homeobox gene tinman plays an essential role in segregating cardiac and visceral muscle potentiality, as well as that of some somatic muscles, in nascent mesoderm of the fly embryo. In its absence, precursor cells for these muscles do not form. tinman homologues have now been isolated from vertebrate genomes and at least one of them, Nkx2-5, is expressed in heart progenitor cells and is essential for myogenic and morphogenetic differentiation of the mammalian heart. Signaling pathways that establish the tin expression domain also appear to be conserved in vertebrates. These findings suggest that heart development in flies and vertebrates utilize similar genetic pathways and engender optimism that the dissection of mammalian heart development will profoundly profit from the rich genetics of Drosophila. The findings also prompt the questions: are the hearts of vertebrates and invertebrates actually homologous, and how much can we learn from the comparative approach? In the sections below, the structure, regulation, function, and evolution of NK class homeobox genes will be reviewed, emphasizing and contrasting the roles of tinman and Nkx2-5 in heart development.

CompGO: an R package for comparing and visualizing Gene Ontology enrichment differences between DNA binding experiments

Publisher: Springer Science and Business Media LLC

Date: 02-09-2015

DOI: 10.1186/S12859-015-0701-2

MyoD-family inhibitor proteins act as auxiliary subunits of Piezo channels

Publisher: American Association for the Advancement of Science (AAAS)

Date: 18-08-2023

DOI: 10.1126/SCIENCE.ADH8190

Abstract: Piezo channels are critical cellular sensors of mechanical forces. Despite their large size, ubiquitous expression, and irreplaceable roles in an ever-growing list of physiological processes, few Piezo channel–binding proteins have emerged. In this work, we found that MyoD (myoblast determination)–family inhibitor proteins (MDFIC and MDFI) are PIEZO1/2 interacting partners. These transcriptional regulators bind to PIEZO1/2 channels, regulating channel inactivation. Using single-particle cryogenic electron microscopy, we mapped the interaction site in MDFIC to a lipidated, C-terminal helix that inserts laterally into the PIEZO1 pore module. These Piezo-interacting proteins fit all the criteria for auxiliary subunits, contribute to explaining the vastly different gating kinetics of endogenous Piezo channels observed in many cell types, and elucidate mechanisms potentially involved in human lymphatic vascular disease.

musculin: a murine basic helix-loop-helix transcription factor gene expressed in embryonic skeletal muscle

Publisher: Elsevier BV

Date: 08-1998

DOI: 10.1016/S0925-4773(98)00122-1

Abstract: We describe the embryonic expression of musculin, a new murine member of the bHLH family of transcription factors. Musculin protein is closely related to human ABF-1, which is expressed in activated B cells, and to epicardin/capsulin/Pod-1, which is expressed in branchial myoblasts, visceral and urogenital mesoderm and epicardium. In situ hybridisation revealed musculin expression in embryos was largely restricted to the embryonic skeletal muscle lineage. While all skeletal muscles expressed the gene, only a subset of myocytes within each muscle were positive, indicating molecular heterogeneity within fetal muscle.

Single-cell transcriptomics for the assessment of cardiac disease

Publisher: Springer Science and Business Media LLC

Date: 20-12-2022

DOI: 10.1038/S41569-022-00805-7

PDGF signaling directs cardiomyocyte movement toward the midline during heart tube assembly

Publisher: Cold Spring Harbor Laboratory

Date: 21-09-2016

DOI: 10.1101/076703

Abstract: Communication between neighboring tissues plays a central role in guiding organ morphogenesis. During heart tube assembly, interactions with the adjacent endoderm control the medial movement of cardiomyocytes, a process referred to as cardiac fusion. However, the molecular underpinnings of this endodermal-myocardial relationship remain unclear. Here, we show an essential role for platelet-derived growth factor receptor alpha (Pdgfra) in directing cardiac fusion. In both zebrafish and mouse, mutation of pdgfra inhibits cardiac fusion and can lead to cardia bifida. Timelapse analysis of in idual cardiomyocyte trajectories reveals misdirected cells in zebrafish pdgfra mutants, suggesting that PDGF signaling steers cardiomyocytes toward the midline. Intriguingly, the ligand pdgfaa is expressed in the endoderm medial to the pdgfra -expressing myocardial precursors. Ectopic expression of pdgfaa interferes with cardiac fusion, consistent with an instructive role for PDGF signaling. Together, these data uncover a novel mechanism through which endodermal-myocardial communication guides the cell movements that initiate cardiac morphogenesis. Studies in zebrafish and mouse implicate the PDGF signaling pathway in the communication between the endoderm and the myocardium that drives medial myocardial movement and thereby initiates cardiac morphogenesis.

Altered motor activity, exploration and anxiety in heterozygous neuregulin 1 mutant mice: Implications for understanding schizophrenia

Publisher: Wiley

Date: 29-11-2007

DOI: 10.1111/J.1601-183X.2006.00298.X

Cardiac Deletion of Smyd2 Is Dispensable for Mouse Heart Development

Publisher: Public Library of Science (PLoS)

Date: 17-03-2010

DOI: 10.1371/JOURNAL.PONE.0009748

MyoD protein expression in Xenopus embryos closely follows a mesoderm induction-dependent amplification of MyoD transcription and is synchronous across the future somite axis

Publisher: Elsevier BV

Date: 05-1992

DOI: 10.1016/0925-4773(92)90076-V

Abstract: The MyoD-related genes code for key regulators of skeletal muscle commitment and differentiation. In this study, expression of MyoD protein has been examined during Xenopus development. Protein is first detected in presumptive mesoderm at early gastrulation, directly following a dramatic increase in MyoD transcription that occurs in response to mesoderm induction. The pattern of expression resembles the muscle fate map at this time. Protein accumulates synchronously along the future somite axis, with no evidence of a spatial regulation which would explain the anterior osterior wave of myogenic differentiation that follows MyoD expression. During gastrulation, the highest levels of MyoD are in cells next to the developing notochord, suggesting a role for the notochord in induction or maintenance of MyoD expression. After muscle differentiation, MyoD protein is degraded with a half-life of several hours, leading to very low expression in mature somites. These studies support a role for MyoD in induction of muscle mesoderm, but also point to the multi-layered regulation of these events.

Heart field origin of great vessel precursors relies on nkx2.5-mediated vasculogenesis

Publisher: Springer Science and Business Media LLC

Date: 27-10-2013

DOI: 10.1038/NCB2862

Deletion of Nkx2-5 in trabecular myocardium reveals the developmental origins of pathological heterogeneity associated with ventricular non-compaction cardiomyopathy

Publisher: Public Library of Science (PLoS)

Date: 06-07-2018

DOI: 10.1371/JOURNAL.PGEN.1007502

Fibroblast growth factor-mediated proliferation of central nervous system precursors depends on endogenous production of insulin-like growth factor I.

Publisher: Proceedings of the National Academy of Sciences

Date: 15-03-1991

DOI: 10.1073/PNAS.88.6.2199

Abstract: Fibroblast growth factor stimulates proliferation and subsequent differentiation of precursor cells isolated from the neuroepithelium of embryonic day 10 mice in vitro. Here we show that fibroblast growth factor-induced proliferation is dependent on the presence of insulin-like growth factors (IGFs) and that IGF-I is endogenously produced by the neuroepithelial cells. Blocking of endogenous IGF-I activity with anti-IGF-I antibodies results in complete inhibition of fibroblast growth factor-mediated proliferation and in cell death. IGF-I alone acts as a survival agent. These observations correlate with the detection of transcripts for IGF-I and basic fibroblast growth factor in freshly isolated neuroepithelium and are consistent with an autocrine action of these factors in early brain development in vivo.

Tissue-Resident PDGFRα+ Progenitor Cells Contribute to Fibrosis versus Healing in a Context- and Spatiotemporally Dependent Manner

Publisher: Elsevier BV

Date: 2020

DOI: 10.1016/J.CELREP.2019.12.045

Two CCAAT boxes in a novel inverted repeat motif are required for Hlx homeobox gene expression

Publisher: Elsevier BV

Date: 05-2001

DOI: 10.1016/S0167-4781(01)00217-2

Abstract: Hlx is a homeobox transcription factor gene required for normal intestinal and hepatic growth in development. We previously found high sequence identity and 17 conserved consensus cis-regulatory/transcription factor binding elements in the mouse and human Hlx 5' regions. A 594 bp sequence in the Hlx 5' region possessing the same activity in driving luciferase expression as larger Hlx 5' sequences had three segments each necessary but not sufficient for luciferase expression in NIH 3T3 cells (which express Hlx). Nine of the conserved putative regulatory elements are positioned within these segments, including two CCAAT boxes on opposite strands within a conserved 44 bp inverted repeat sequence. To test the hypothesis that these elements are required for promoter activity, we compared the reporter expression activity of segments containing mutations of these elements with activity of the parent Hlx promoter sequence. We found that mutation of either CCAAT box or a conserved AP-2 site resulted in a significant decrease in promoter activity. Restoration of the inverted repeat with complementary mutations of both CCAAT boxes did not restore activity. Further, mutation of other portions of the inverted repeat did not affect promoter activity. Mutation of other elements had no effect on promoter activity.

Analysis of cardiac stem cell self-renewal dynamics in serum-free medium by single cell lineage tracking

Publisher: Elsevier BV

Date: 04-2018

DOI: 10.1016/J.SCR.2018.02.004

Abstract: Cardiac colony forming unit-fibroblasts (cCFU-F) are a population of stromal cells residing within the SCA1

Revitalizing Our View of Heart Development

Publisher: Elsevier BV

Date: 09-2011

DOI: 10.1016/J.DEVCEL.2011.08.011

Essential Role for the Lymphostromal Plasma Membrane Ly-6 Superfamily Molecule Thymic Shared Antigen 1 in Development of the Embryonic Adrenal Gland

Publisher: Informa UK Limited

Date: 02-2002

DOI: 10.1128/MCB.22.3.946-952.2002

Intravital Imaging to Monitor Therapeutic Response in Moving Hypoxic Regions Resistant to PI3K Pathway Targeting in Pancreatic Cancer

Publisher: Elsevier BV

Date: 06-1993

DOI: 10.1016/J.CELREP.2018.05.038

Single-Cell Transcriptome Dynamics of the Autotaxin-Lysophosphatidic Acid Axis During Muscle Regeneration Reveal Proliferative Effects in Mesenchymal Fibro-Adipogenic Progenitors

Publisher: Cold Spring Harbor Laboratory

Date: 03-07-2022

DOI: 10.1101/2022.07.02.498539

Abstract: Lysophosphatidic acid (LPA) is a growth factor-like bioactive phospholipid recognising LPA receptors (LPARs) and mediating signalling pathways that regulate embryonic development, wound healing, carcinogenesis, and fibrosis, via effects on cell migration, proliferation and differentiation. Extracellular LPA is generated from lysophospholipids by the secreted hydrolase - ectonucleotide pyrophosphatase hosphodiesterase 2 (ENPP2 also, AUTOTAXIN / ATX) and metabolised by different membrane-bound phospholipid phosphatases (PLPPs). Here, we use public bulk and singlecell RNA sequencing datasets to explore the gene expression of Lpar 1-6 , Enpp2 , and Plpp genes under skeletal muscle homeostasis and regeneration conditions. We show that the skeletal muscle system dynamically expresses the Enpp2-Lpar-Plpp gene axis, with Lpar1 being the highest expressed member among LPARs. Lpar1 was expressed by mesenchymal fibro-adipogenic progenitors (FAPs) and tenocytes, whereas FAPs mainly expressed Enpp2 . Clustering of FAPs identified populations representing distinct cell states with robust Lpar1 and Enpp2 transcriptome signatures in homeostatic cells expressing higher levels of markers Dpp4 and Hsd11b1 . However, tissue injury induced transient repression of Lpar genes and Enpp2 . The role of LPA in modulating the fate and differentiation of tissue-resident FAPs has not yet been explored. Ex vivo, LPAR1/3 and ENPP2 inhibition significantly decreased the cell-cycle activity of FAPs and impaired fibro-adipogenic differentiation, implicating LPA signalling in the modulation of the proliferative and differentiative fate of FAPs. Together, our results demonstrate the importance of the ENPP2-LPAR-PLPP axis in different muscle cell types and FAP lineage populations in homeostasis and injury, paving the way for further research on the role of this signalling pathway in skeletal muscle homeostasis and regeneration, and that of other organs and tissues, in vivo. Our reanalysis of single-cell transcriptomics revealed the involvement and temporally dynamic expression of the ENPP2-LPAR-PLPP axis in response to skeletal muscle regeneration.

Congenital Asplenia in Mice and Humans with Mutations in a Pbx/Nkx2-5/p15 Module

Publisher: Elsevier BV

Date: 05-2012

DOI: 10.1016/J.DEVCEL.2012.02.009

Muscle costameric protein, Chisel/Smpx, associates with focal adhesion complexes and modulates cell spreading in vitro via a Rac1/p38 pathway

Publisher: Elsevier BV

Date: 07-2005

DOI: 10.1016/J.YEXCR.2005.04.006

Abstract: The murine X-linked gene Chisel (Csl/Smpx) encodes a 9-kDa protein that associates in heart and skeletal muscle cells with the costameric cytoskeleton, implicated in maintaining muscle integrity and responses to biomechanical stress. After expression in C2C12 myoblasts, MYC epitope-tagged Csl co-localized with actin networks at peripheral membranes, and with focal adhesion proteins vinculin, paxillin, integrin beta1, and the small GTPase Rac1. Csl could be co-immunoprecipitated with vinculin from extracts of C2C12 cells and native muscle. MYC-Csl induced cell spreading and lamellipodia formation in C2C12 cells at the expense of filopodia, suggestive of modulation of Rac1 activity. Lamellipodia formation was indeed Rac1-dependent, and in MYC-Csl cells replated on fibronectin, Rac1 activity was increased relative to controls. Expression of MYC-Csl led to an increased association between vinculin and p34, a subunit of the Arp2/3 actin nucleation complex, a Rac1-dependent event. Induced cell spreading was also dependent upon p38 kinases that act downstream of Rac1 to control the actin capping activity of heat shock protein 27. Our data suggest that Csl localizes to the costameric cytoskeleton of muscle cells through an association with focal adhesion proteins, where it may participate in regulation of cytoskeletal dynamics through the Rac1-p38 pathway.

Comparative regenerative mechanisms across different mammalian tissues

Publisher: Springer Science and Business Media LLC

Date: 23-02-2018

DOI: 10.1038/S41536-018-0044-5

Abstract: Stimulating regeneration of complex tissues and organs after injury to effect complete structural and functional repair, is an attractive therapeutic option that would revolutionize clinical medicine. Compared to many metazoan phyla that show extraordinary regenerative capacity, which in some instances persists throughout life, regeneration in mammalians, particularly humans, is limited or absent. Here we consider recent insights in the elucidation of molecular mechanisms of regeneration that have come from studies of tissue homeostasis and injury repair in mammalian tissues that span the spectrum from little or no self-renewal, to those showing active cell turnover throughout life. These studies highlight the ersity of factors that constrain regeneration, including immune responses, extracellular matrix composition, age, injury type, physiological adaptation, and angiogenic and neurogenic capacity. Despite these constraints, much progress has been made in elucidating key molecular mechanisms that may provide therapeutic targets for the development of future regenerative therapies, as well as previously unidentified developmental paradigms and windows-of-opportunity for improved regenerative repair.

Transcription from the intron-containing chicken histone H2A._Fgene is not S-phase regulated

Publisher: Oxford University Press (OUP)

Date: 1989

DOI: 10.1093/NAR/17.4.1745

Abstract: The nucleotide sequence of an 8.2 kb BamHI fragment containing the entire chicken histone H2AF gene has been determined. Unlike the majority of histone genes, the coding region is interrupted by four intervening sequences. While sequencing the 8.2 kb BamHI fragment it was found that the promoter and first exon of an unidentified non-histone gene lies immediately downstream of the H2AF gene. Studies of H2AF gene transcription show that, unlike the major core and H1 histone genes, it is not coupled to DNA synthesis.

The nu gene acts cell-autonomously and is required for differentiation of thymic epithelial progenitors.

Publisher: Proceedings of the National Academy of Sciences

Date: 11-06-1996

DOI: 10.1073/PNAS.93.12.5742

Abstract: The nude mutation (nu) causes athymia and hairlessness, but the molecular mechanisms by which it acts have not been determined. To address the role of nu in thymogenesis, we investigated whether all or part of the nude thymic epithelium could be rescued by the presence of wild-type cells in nude -- wild-type chimeric mice. Detailed immunohistochemical analyses revealed that nude-derived cells could persist in the chimeric thymus but could not contribute to cortical or medullary epithelial networks. Nude-derived cells, present in few clusters in the medulla, expressed markers of a rare subpopulation of adult medullary epithelium. The thymic epithelial rudiment of nude mice strongly expressed these same markers, which may therefore define committed immature thymic epithelial precursor cells. To our knowledge, these data provide the first evidence that the nu gene product acts cell-autonomously and is necessary for the development of all major subpopulations of mature thymic epithelium. We propose that nu acts to regulate growth and/or differentiation, but not determination, of thymic epithelial progenitors.

Expression of Slit and Robo genes in the developing mouse heart

Publisher: Wiley

Date: 12-10-2010

DOI: 10.1002/DVDY.22449

Genetic burden and associations with adverse neurodevelopment in neonates with congenital heart disease

Publisher: Elsevier BV

Date: 07-2018

DOI: 10.1016/J.AHJ.2018.03.021

Abstract: Up to 20% of children with congenital heart disease (CHD) undergoing cardiac surgery develop neurodevelopmental disabilities (NDD), with some studies reporting persistent impairment. Recent large-scale studies have demonstrated shared genetic mechanisms contributing to CHD and NDD. In this study, a targeted approach was applied to assess direct clinical applicability of this information. A gene panel comprising 148 known CHD and/or NDD genes was used to sequence 15 patients with CHD + NDD, 15 patients with CHD, and 15 healthy controls. The number and types of variants between the 3 groups were compared using Poisson log-linear regression, and the SNP-set (Sequence) Kernel Association Test-Optimized was used to conduct single-gene and gene-pathway burden analyses. A significant increase in rare (minor allele frequency < 0.01) and novel variants was identified between the CHD + NDD cohort and controls, P < .001 and P = .001, respectively. There was also a significant increase in rare variants in the CHD cohort compared with controls (P = .04). Rare variant burden analyses implicated pathways associated with "neurotransmitters," "axon guidance," and those incorporating "RASopathy" genes in the development of NDD in CHD patients. These findings suggest that an increase in novel and rare variants in known CHD and/or NDD genes is associated with the development of NDD in patients with CHD. Furthermore, burden analyses point toward rare variant burden specifically in pathways related to brain development and function as contributors to NDD. Although promising variants and pathways were identified, further research, utilizing whole-genome approaches, is required prior to demonstrating clinical utility in this patient group.

Inhibition of Notch2 by Numb/Numblike controls myocardial compaction in the heart

Publisher: Oxford University Press (OUP)

Date: 03-08-2012

DOI: 10.1093/CVR/CVS250

Abstract: The ventricular wall of the heart is composed of trabeculated and compact layers, which are separated by yet unknown processes during embryonic development. Here, we wanted to explore the role of Notch2 and Numb/Numblike for myocardial trabeculation and compaction. We found that Notch2 activity is specifically down-regulated in the compact layer during cardiac development in the mouse. The biological role of Notch2 down-regulation was investigated by the expression of constitutively active Notch2 in the myocardium of transgenic mice, resulting in hypertrabeculation, reduced compaction, and ventricular septum defects. To disclose the mechanism that inhibited Notch2 activity during the formation of myocardial layers, we analysed potential suppressors of Notch signalling. We unveiled that concomitant but not separate ablation of Numb and Numblike in the developing heart leads to increased Notch2 activity along with hypertrabeculation, reduced compaction, and ventricular septum defects, phenocopying effects gained by overexpression of constitutively active Notch2. Expression profiling revealed a strong up-regulation of Bmp10 in Numb/Numblike mutant hearts, which might also interfere with trabeculation and compaction. This study identified potential novel roles of Numb/Numblike in regulating trabeculation and compaction by inhibiting Notch2 and Bmp10 signalling.

Landmarks and Lineages in the Developing Heart

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 05-06-2009

DOI: 10.1161/CIRCRESAHA.109.199729

Differential Binding of an SRF/NK-2/MEF2 Transcription Factor Complex in Normal Versus Neoplastic Smooth Muscle Tissues

Publisher: Elsevier BV

Date: 09-2001

DOI: 10.1074/JBC.M105826200

Absence of yolk sac hematopoiesis from mice with a targeted disruption of the scl gene.

Publisher: Proceedings of the National Academy of Sciences

Date: 18-07-1995

DOI: 10.1073/PNAS.92.15.7075

Abstract: The scl gene encodes a basic-helix-loop-helix transcription factor which was identified through its involvement in chromosomal translocations in T-cell leukemia. To elucidate its physiological role, scl was targeted in embryonic stem cells. Mice heterozygous for the scl null mutation were intercrossed and their offspring were genotyped. Homozygous mutant (scl-/-) pups were not detected in newborn litters, and analysis at earlier time points demonstrated that scl-/- embryos were dying around embryonic day 9.5. The scl-/- embryos were pale, edematous, and markedly growth retarded after embryonic day 8.75. Histological studies showed complete absence of recognizable hematopoiesis in the yolk sac of these embryos. Early organogenesis appeared to be otherwise normal. Culture of yolk sac cells of wild-type, heterozygous, and homozygous littermates confirmed the absence of hematopoietic cells in scl-/- yolk sacs. Reverse transcription PCR was used to examine the transcripts of several genes implicated in early hematopoiesis. Transcripts of GATA-1 and PU.1 transcription factors were absent from RNA from scl-/- yolk sacs and embryos. These results implicate scl as a crucial regulator of early hematopoiesis.

Nkx2-5 Mediates Differential Cardiac Differentiation Through Interaction with Hoxa10

Publisher: Mary Ann Liebert Inc

Date: 08-2013

DOI: 10.1089/SCD.2012.0611

Epicardial Origin of Resident Mesenchymal Stem Cells in the Adult Mammalian Heart

Publisher: MDPI AG

Date: 23-04-2014

DOI: 10.3390/JDB2020117

Author response: Single-cell expression profiling reveals dynamic flux of cardiac stromal, vascular and immune cells in health and injury

Publisher: eLife Sciences Publications, Ltd

Date: 11-03-2019

DOI: 10.7554/ELIFE.43882.061

Disrupted cardiac development but normal hematopoiesis in mice deficient in the second CXCL12/SDF-1 receptor, CXCR7

Publisher: Proceedings of the National Academy of Sciences

Date: 11-09-2007

DOI: 10.1073/PNAS.0702229104

Abstract: Chemotactic cytokines (chemokines) attract immune cells, although their original evolutionary role may relate more closely with embryonic development. We noted differential expression of the chemokine receptor CXCR7 (RDC-1) on marginal zone B cells, a cell type associated with autoimmune diseases. We generated Cxcr7 −/− mice but found that CXCR7 deficiency had little effect on B cell composition. However, most Cxcr7 −/− mice died at birth with ventricular septal defects and semilunar heart valve malformation. Conditional deletion of Cxcr7 in endothelium, using Tie2-Cre transgenic mice, recapitulated this phenotype. Gene profiling of Cxcr7 −/− heart valve leaflets revealed a defect in the expression of factors essential for valve formation, vessel protection, or endothelial cell growth and survival. We confirmed that the principal chemokine ligand for CXCR7 was CXCL12/SDF-1, which also binds CXCR4. CXCL12 did not induce signaling through CXCR7 however, CXCR7 formed functional heterodimers with CXCR4 and enhanced CXCL12-induced signaling. Our results reveal a specialized role for CXCR7 in endothelial biology and valve development and highlight the distinct developmental role of evolutionary conserved chemokine receptors such as CXCR7 and CXCR4.

Widespread expression of MyoD genes in Xenopus embryos is amplified in presumptive muscle as a delayed response to mesoderm induction.

Publisher: Proceedings of the National Academy of Sciences

Date: 15-10-1991

DOI: 10.1073/PNAS.88.20.9198

Abstract: The MyoD gene codes for an important regulatory factor in skeletal myogenesis. To explore the relationship between mesoderm induction in Xenopus embryos and expression of MyoD, I have monitored MyoD mRNA levels in normal embryos and cultured explants by RNase protection. Transcription from the two Xenopus MyoD gene copies is activated weakly across the whole embryo at the midblastula transition, and this activation occurs in the absence of mesoderm induction. In response to induction this basal expression is lified 50- to 100-fold, but in animal-pole explants 6-10 hr elapse before induced mRNAs appear, and this induction requires prior protein synthesis. The promiscuous transcripts disappear from animal explants at a time when induction "competence" is lost, suggesting a link between these events. The data highlight a broad, but transient, permissiveness for MyoD expression in embryos, which is propagated and lified only in presumptive muscle in response to induction. Moreover, muscle-specific MyoD expression is a relatively late (postgastrulation) event in the mesoderm-induction cascade.

Cardiac septal and valvular dysmorphogenesis in mice heterozygous for mutations in the homeobox gene Nkx2-5

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 10-11-2000

DOI: 10.1161/01.RES.87.10.888

Abstract: Abstract —Heterozygous mutations in the cardiac homeobox gene, NKX2-5 , underlie familial cases of atrial septal defect (ASD) with severe atrioventricular conduction block. In this study, mice heterozygous for Nkx2-5 –null alleles were assessed for analogous defects. Although ASD occurred only rarely, atrial septal dysmorphogenesis was evident as increased frequencies of patent foramen ovale and septal aneurysm, and decreased length of the septum primum flap valve. These parameters were compounded by genetic background effects, and in the 129/Sv strain, septal dysmorphogenesis bordered on ASD in 17% of Nkx2-5 heterozygotes. In a proportion of neonatal heterozygotes, as well as in adults with ASD, we found that the size of the foramen ovale was significantly enlarged and altered in shape, potentially exposing the normally thin septum primum to excessive hemodynamic forces. Therefore, defective morphogenesis of the septum secundum may be one contributing factor in the generation of patent foramen ovale, septal aneurysm, and certain ASDs. Mild prolongation of P-R interval in females and an increased frequency of stenotic bicuspid aortic valves were also features of the Nkx2-5 heterozygous phenotype. Our data demonstrate that the complex effects of Nkx2-5 haploinsufficiency in mice are weaker but convergent with those in humans. As in the mouse, the phenotype of human NKX2-5 mutations may be modulated by interacting alleles.

The promises and challenges of exome sequencing in familial, non-syndromic congenital heart disease

Publisher: Elsevier BV

Date: 03-2017

DOI: 10.1016/J.IJCARD.2016.12.024

Abstract: Exome sequencing is an established strategy to identify causal variants in families with two or more members affected by congenital heart disease (CHD). This unbiased approach, in which both rare and common variants are identified, makes it suitable to research complex, heterogeneous diseases such as CHD. Exome sequencing was performed on two affected members of a three generation family with atrial septal defects (ASD), suggesting a dominant inheritance pattern. Variants were filtered using two bioinformatics pipelines and prioritised according to in silico prediction programs. Segregation studies and functional analyses were used to assess co-segregation with disease and effects on protein function, respectively. Following the data and in silico analyses, ten candidate variants were prioritised. Of these, SRPK2 (c.2044C>T[p.Arg682Trp]) and NOTCH1 (c.3835C>T[p.Arg1279Cys]), co-segregated with disease in the family however, previous functional analyses on SRPK2 make this an unlikely candidate. Functional analyses in the variant (c.3835C>T[p.Arg1279Cys]) of the known CHD gene NOTCH1 demonstrated a non-significant decrease in signalling activity. This study demonstrates both the potential, as well as the challenges, of applying exome sequencing to complex diseases such as CHD. While in silico evidence and segregation analyses in the NOTCH1 p.Arg1279Cys variant are highly suggestive of pathogenicity, the minimal change in signalling capacity suggests that other variants may be required for CHD development. This study highlights the difficulties of applying exome sequencing in familial, non-syndromic CHD in the clinical environment and a cautionary note in the interpretation of apparently causal abnormalities in silico without supportive functional data.

The Hlx homeobox transcription factor is required early in enteric nervous system development

Publisher: Springer Science and Business Media LLC

Date: 19-07-2006

DOI: 10.1186/1471-213X-6-33

Abstract: Development of the enteric nervous system (ENS) requires interactions between migrating neural crest cells and the nascent gastrointestinal tract that are dependent upon genes expressed by both cell compartments. Hlx , a homeobox transcription factor gene that is expressed in mouse intestinal and hepatic mesenchyme, is required for normal embryonic growth of intestine and liver, and the Hlx -/- genotype is embryonic lethal. We hypothesized that Hlx is required for ENS development. Enteric neurons were identified in Hlx +/+ and Hlx -/- mouse embryos by immunostaining of embryo sections for the neural markers PGP9.5 and Phox2b, or by staining for β-galactosidase in whole-mount embryos containing the dopamine β-hydroxylase-n LacZ transgene. In Hlx +/+ embryos, neural crest cells/enteric neurons have moved from the stomach into the intestine by E10.5. By contrast, neural crest cells/enteric neurons remain largely restricted to the lateral stomach mesenchyme of Hlx -/- embryos, with only a few scattered neural crest cells/enteric neurons in the intestine between E10.5–16.5. The Hlx homeobox transcription factor is required for early aspects of ENS development.

Epistatic and Independent Effects on Schizophrenia-Related Phenotypes Following Co-disruption of the Risk Factors Neuregulin-1 × DISC1

Publisher: Oxford University Press (OUP)

Date: 09-09-2016

DOI: 10.1093/SCHBUL/SBW120

Abstract: Few studies have addressed likely gene × gene (ie, epistatic) interactions in mediating risk for schizophrenia. Using a preclinical genetic approach, we investigated whether simultaneous disruption of the risk factors Neuregulin-1 (NRG1) and Disrupted-in-schizophrenia 1 (DISC1) would produce a disease-relevant phenotypic profile different from that observed following disruption to either gene alone. NRG1 heterozygotes exhibited hyperactivity and disruption to prepulse inhibition, both reversed by antipsychotic treatment, and accompanied by reduced striatal dopamine D2 receptor protein expression, impaired social cognition, and altered glutamatergic synaptic protein expression in selected brain areas. Single gene DISC1 mutants demonstrated a disruption in social cognition and nest-building, altered brain 5-hydroxytryptamine levels and hippoc al ErbB4 expression, and decreased cortical expression of the schizophrenia-associated microRNA miR-29b. Co-disruption of DISC1 and NRG1, indicative of epistasis, evoked an impairment in sociability and enhanced self-grooming, accompanied by changes in hypothalamic oxytocin/vasopressin gene expression. The findings indicate specific behavioral correlates and underlying cellular pathways downstream of main effects of DNA variation in the schizophrenia-associated genes NRG1 and DISC1.

The small muscle-specific protein CSl modifies cell shape and promotes myocyte fusion in an insulin-like growth factor 1-dependent manner

Publisher: Rockefeller University Press

Date: 21-05-2001

DOI: 10.1083/JCB.153.5.985

Abstract: We have isolated a murine cDNA encoding a 9-kD protein, Chisel (Csl), in a screen for transcriptional targets of the cardiac homeodomain factor Nkx2-5. Csl transcripts were detected in atria and ventricles of the heart and in all skeletal muscles and smooth muscles of the stomach and pulmonary veins. Csl protein was distributed throughout the cytoplasm in fetal muscles, although costameric and M-line localization to the muscle cytoskeleton became obvious after further maturation. Targeted disruption of Csl showed no overt muscle phenotype. However, ectopic expression in C2C12 myoblasts induced formation of lamellipodia in which Csl protein became tethered to membrane ruffles. Migration of these cells was retarded in a monolayer wound repair assay. Csl-expressing myoblasts differentiated and fused normally, although in the presence of insulin-like growth factor (IGF)-1 they showed dramatically enhanced fusion, leading to formation of large dysmorphogenic “myosacs.” The activities of transcription factors nuclear factor of activated T cells (NFAT) and myocyte enhancer–binding factor (MEF)2, were also enhanced in an IGF-1 signaling–dependent manner. The dynamic cytoskeletal localization of Csl and its dominant effects on cell shape and behavior and transcription factor activity suggest that Csl plays a role in the regulatory network through which muscle cells coordinate their structural and functional states during growth, adaptation, and repair.

Chromatin remodelling complex dosage modulates transcription factor function in heart development

Publisher: Springer Science and Business Media LLC

Date: 08-02-2011

DOI: 10.1038/NCOMMS1187

Human cerberus related gene CER1 maps to chromosome 9

Publisher: Elsevier BV

Date: 02-1999

DOI: 10.1006/GENO.1998.5671

Identification and cloning of localized maternal RNAs from xenopus eggs

Publisher: Elsevier BV

Date: 10-1985

DOI: 10.1016/0092-8674(85)90273-9

Abstract: A central question in developmental biology is to explain how cells in different regions of an embryo acquire different developmental fates. We have begun to address this question by investigating whether specific RNAs are localized within a frog egg. Differential screening of a cDNA library shows that most maternal RNAs are uniformly distributed along the animal-vegetal axis. However, we find that a rare class of maternal RNAs is localized. cDNA clones of four localized RNAs have been characterized. Three of these cDNAs are derived from maternal RNAs that are concentrated in the animal hemisphere of unfertilized eggs and remain localized through the early blastula stage. One cDNA is derived from a maternal RNA found almost exclusively in the vegetal hemisphere at both stages. These studies show that some informational molecules, specifically RNAs, are localized in eggs and are inherited by particular blastomeres.

Disruption to social dyadic interactions but not emotional/anxiety-related behaviour in mice with heterozygous ‘knockout’ of the schizophrenia risk gene neuregulin-1

Publisher: Elsevier BV

Date: 02-2008

DOI: 10.1016/J.PNPBP.2007.09.018

Abstract: Clinical genetic studies have implicated neuregulin-1 [NRG1] as a leading susceptibility gene for schizophrenia. NRG1 is known to play a significant role in the developing brain, which is consistent with the prevailing neurodevelopmental model of schizophrenia. Thus, the emotional and social phenotype of adult mice with heterozygous 'knockout' of transmembrane [TM]-domain NRG1 was examined further in both sexes. Emotional/anxiety-related behaviour was assessed using the elevated plus-maze and the light-dark test. Social behaviour was examined in terms of dyadic interactions between NRG1 mutants and an unfamiliar C57BL6 conspecific in a novel environment. There was no effect of NRG1 genotype on performance in either test of emotionality/anxiety. However, previous reports of hyperactivity in NRG1 mutants were confirmed in both paradigms. In the test of social interaction, aggressive following was increased in NRG1 mutants of both sexes, together with an increase in walkovers in female mutants. These findings elaborate the specificity of the NRG1 phenotype for the social rather than the emotional/anxiety-related domain. They indicate that NRG1 is involved in the regulation of reciprocal social interaction behaviour and thus suggest a putative role for NRG1 in a schizophrenia-related endophenotype.

Whole genome sequencing in transposition of the great arteries and associations with clinically relevant heart, brain and laterality genes

Publisher: Elsevier BV

Date: 02-2022

DOI: 10.1016/J.AHJ.2021.10.185

Abstract: The most common cyanotic congenital heart disease (CHD) requiring management as a neonate is transposition of great arteries (TGA). Clinically, up to 50% of TGA patients develop some form of neurodevelopmental disability (NDD), thought to have a significant genetic component. A "ciliopathy" and links with laterality disorders have been proposed. This first report of whole genome sequencing in TGA, sought to identify clinically relevant variants contributing to heart, brain and laterality defects. Initial whole genome sequencing analyses on 100 TGA patients focussed on established disease genes related to CHD (n = 107), NDD (n = 659) and heterotaxy (n = 74). Single variant as well as copy number variant analyses were conducted. Variant pathogenicity was assessed using the American College of Medical Genetics and Genomics-Association for Molecular Pathology guidelines. Fifty-five putatively damaging variants were identified in established disease genes associated with CHD, NDD and heterotaxy however, no clinically relevant variants could be attributed to disease. Notably, case-control analyses identified significantly more predicted-damaging, silent and total variants in TGA cases than healthy controls in established CHD genes (P < .001), NDD genes (P < .001) as well as across the three gene panels (P < .001). We present compelling evidence that the majority of TGA is not caused by monogenic rare variants and is most likely oligogenic and/or polygenic in nature, highlighting the complex genetic architecture and multifactorial influences on this CHD sub-type and its long-term sequelae. Assessment of variant burden in key heart, brain and/or laterality genes may be required to unravel the genetic contributions to TGA and related disabilities.

Vertebrate histone genes: nucleotide sequence of a chicken H2A gene and regulatory flanking sequences

Publisher: Oxford University Press (OUP)

Date: 1981

DOI: 10.1093/NAR/9.13.3119

Abstract: The DNA sequence of a chicken genomal fragment containing a histone H2A gene has been determined. It contains extensive 5' and 3' flanking regions and encodes a protein identical in sequence to the histone H2A protein isolated from chicken erythrocytes. In the 5' flanking region, a possible "TATA box" and three possible "cap sites" can be recognised upstream from the initiation codon. To the 5' side of the "TATA box" is found an unusual sequence of 21 A's interrupted by a central G residue. It occupies the same relative position as the P. miliaris H2A gene-specific 5' dyad symmetry sequence and the "CCAAT box" seen in other eukaryotic polymerase II genes but is clearly different from both. A significant feature of the 3' non-coding region is the presence of a 23 base-pair sequence that is nearly identical to a conserved region found in sea urchin histone genes. The coding region is extremely GC rich, with strong selection for these bases in the third position of codons. Not a single coding triplet ends in U. No intervening sequences were found in this gene.

Cardiomyocytes Replicate and their Numbers Increase in Young Hearts

Publisher: Elsevier BV

Date: 11-2015

DOI: 10.1016/J.CELL.2015.10.038

Murine T-box transcription factor Tbx20 acts as a repressor during heart development, and is essential for adult heart integrity, function and adaptation

Publisher: The Company of Biologists

Date: 15-05-2005

DOI: 10.1242/DEV.01799

Abstract: The genetic hierarchies guiding lineage specification and morphogenesis of the mammalian embryonic heart are poorly understood. We now show by gene targeting that murine T-box transcription factor Tbx20 plays a central role in these pathways, and has important activities in both cardiac development and adult function. Loss of Tbx20 results in death of embryos at mid-gestation with grossly abnormal heart morphogenesis. Underlying these disturbances was a severely compromised cardiac transcriptional program,defects in the molecular pre-pattern, reduced expansion of cardiac progenitors and a block to chamber differentiation. Notably, Tbx20-null embryos showed ectopic activation of Tbx2 across the whole heart myogenic field. Tbx2 encodes a transcriptional repressor normally expressed in non-chamber myocardium, and in the atrioventricular canal it has been proposed to inhibit chamber-specific gene expression through competition with positive factor Tbx5. Our data demonstrate a repressive activity for Tbx20 and place it upstream of Tbx2 in the cardiac genetic program. Thus, hierarchical,repressive interactions between Tbx20 and other T-box genes and factors underlie the primary lineage split into chamber and non-chamber myocardium in the forming heart, an early event upon which all subsequent morphogenesis depends. Additional roles for Tbx20 in adult heart integrity and contractile function were revealed by in-vivo cardiac functional analysis of Tbx20 heterozygous mutant mice. These data suggest that mutations in human cardiac transcription factor genes, possibly including TBX20,underlie both congenital heart disease and adult cardiomyopathies.

RNA toxicity in myotonic muscular dystrophy induces NKX2-5 expression

Publisher: Springer Science and Business Media LLC

Date: 16-12-2007

DOI: 10.1038/NG.2007.28

Complex SUMO-1 Regulation of Cardiac Transcription Factor Nkx2-5

Publisher: Public Library of Science (PLoS)

Date: 12-09-2011

DOI: 10.1371/JOURNAL.PONE.0024812

Mutations in crdiac T-box factor gene TBX20 are associated with diverse cardiac pathologies, including defects of septation and valvulogenesis and cardiomyopathy

Publisher: Elsevier BV

Date: 08-2007

DOI: 10.1086/519530

A Universal and Robust Integrated Platform for the Scalable Production of Human Cardiomyocytes From Pluripotent Stem Cells

Publisher: Oxford University Press (OUP)

Date: 28-10-2015

DOI: 10.5966/SCTM.2014-0275

Abstract: Recent advances in the generation of cardiomyocytes (CMs) from human pluripotent stem cells (hPSCs) and the development of novel cell therapy strategies using hPSC-CMs (e.g., cardiac patches) in conjunction with promising preclinical and clinical studies, have raised new hopes for patients with end-stage cardiovascular disease, which remains the leading cause of morbidity and mortality globally. In this study, a simplified, scalable, robust, and integrated differentiation platform was developed to generate clinical grade hPSC-CMs as cell aggregates under chemically defined culture conditions. This approach resulted in approximately 100% beating CM spheroids with virtually pure (∼90%) functional cardiomyocytes in 10 days from multiple hPSC lines. This universal and robust bioprocessing platform can provide sufficient numbers of hPSC-CMs for companies developing regenerative medicine technologies to rescue, replace, and help repair damaged heart tissues and for pharmaceutical companies developing advanced biologics and drugs for regeneration of lost heart tissue using high-throughput technologies. It is believed that this technology can expedite clinical progress in these areas to achieve a meaningful impact on improving clinical outcomes, cost of care, and quality of life for those patients disabled and experiencing heart disease.

GATA4 mutations in 357 unrelated patients with congenital heart malformation

Publisher: Mary Ann Liebert Inc

Date: 12-2010

DOI: 10.1089/GTMB.2010.0028

Abstract: Congenital heart disease (CHD) represents one of the most common birth defects, but the genetic causes remain largely unknown. Mutations in GATA4, encoding a zinc finger transcription factor with a pivotal role in heart development, have been associated with CHD in several familial cases and a small subset of sporadic patients. To estimate the pathogenetic role of GATA4 in CHD, we screened for mutations in 357 unrelated patients with different congenital heart malformations. In addition to nine synonymous changes, we identified two known (A411V and D425N) and two novel putative mutations (G69D and P163R) in five patients with atrial or ventricular septal defects that were not seen in control subjects. The four mutations did not show altered GATA4 transcriptional activity in synergy with the transcription factors NKX2-5 and TBX20. Our data expand the spectrum of mutations associated with cardiac septal defects but do not support GATA4 mutations as a common cause of CHD.

Compensatory Growth of Healthy Cardiac Cells in the Presence of Diseased Cells Restores Tissue Homeostasis during Heart Development

Publisher: Elsevier BV

Date: 10-2008

DOI: 10.1016/J.DEVCEL.2008.09.005

Abstract: Energy generation by mitochondrial respiration is an absolute requirement for cardiac function. Here, we used a heart-specific conditional knockout approach to inactivate the X-linked gene encoding Holocytochrome c synthase (Hccs), an enzyme responsible for activation of respiratory cytochromes c and c1. Heterozygous knockout female mice were thus mosaic for Hccs function due to random X chromosome inactivation. In contrast to midgestational lethality of Hccs knockout males, heterozygous females appeared normal after birth. Analyses of heterozygous embryos revealed the expected 50:50 ratio of Hccs deficient to normal cardiac cells at midgestation however, diseased tissue contributed progressively less over time and by birth represented only 10% of cardiac tissue volume. This change is accounted for by increased proliferation of remaining healthy cardiac cells resulting in a fully functional heart. These data reveal an impressive regenerative capacity of the fetal heart that can compensate for an effective loss of 50% of cardiac tissue.

T-box transcription factors and their roles in regulatory hierarchies in the developing heart

Publisher: The Company of Biologists

Date: 15-11-2005

DOI: 10.1242/DEV.02099

Abstract: T-box transcription factors are important players in the molecular circuitry that generates lineage ersity and form in the developing embryo. At least seven family members are expressed in the developing mammalian heart,and the human T-box genes TBX1 and TBX5 are mutated in cardiac congenital anomaly syndromes. Here, we review T-box gene function during mammalian heart development in the light of new insights into heart morphogenesis. We see for the first time how hierarchies of transcriptional activation and repression involving multiple T-box factors play out in three-dimensional space to establish the cardiac progenitors fields, to define their subservient lineages, and to generate heart form and function.

Expression of NK-2 class homeobox gene Nkx2–6 in foregut endoderm and heart

Publisher: Elsevier BV

Date: 04-1998

DOI: 10.1016/S0925-4773(98)00037-9

Abstract: NK-2 class homeobox genes are candidate patterning and lineage regulators in erse organisms. We report here the embryonic expression pattern of murine member, Nkx2-6. In keeping with its vertebrate relatives, Nkx2-6 was transcribed in ventrolateral embryonic structures. Expression was first detected at E8.0 in endodermal walls of the foregut pocket, tissue destined to become pharyngeal floor. From E8.0-10.5, transcripts were concentrated in pharyngeal pouches and juxtaposed arch ectoderm and mesoderm, as well as in more caudal gut segments. Expression was also seen at opposite poles of the developing heart from E8-8.5 in posterior myocardial progenitors, then sinus venosa and dorsal pericardium, and from E9.5 in outflow tract myocardium.

Sierra: discovery of differential transcript usage from polyA-captured single-cell RNA-seq data

Publisher: Cold Spring Harbor Laboratory

Date: 06-12-2022

DOI: 10.1101/867309

Abstract: High-throughput single-cell RNA-seq (scRNA-seq) is a powerful tool for studying gene expression in single cells. Most current scRNA-seq bioinformatics tools focus on analysing overall expression levels, largely ignoring alternative mRNA isoform expression. We present a computational pipeline, Sierra, that readily detects differential transcript usage from data generated by commonly used polyA-captured scRNA-seq technology. We validate Sierra by comparing cardiac scRNA-seq cell-types to bulk RNA-seq of matched populations, finding significant overlap in differential transcripts. Sierra detects differential transcript usage across human peripheral blood mononuclear cells and the Tabula Muris, and 3’UTR shortening in cardiac fibroblasts. Sierra is available at github.com/VCCRI/Sierra .

Fibroblast growth factor 10 gene regulation in the second heart field by Tbx1, Nkx2-5, and Islet1 reveals a genetic switch for down-regulation in the myocardium

Publisher: Proceedings of the National Academy of Sciences

Date: 04-10-2012

DOI: 10.1073/PNAS.1215360109

Abstract: During cardiogenesis, Fibroblast Growth Factor ( Fgf10 ) is expressed in the anterior second heart field. Together with Fibroblast growth factor 8 (Fgf8), Fgf10 promotes the proliferation of these cardiac progenitor cells that form the arterial pole of the heart. We have identified a 1.7-kb region in the first intron of Fgf10 that is necessary and sufficient to direct transgene expression in this cardiac context. The 1.7-kb sequence is directly controlled by T-box transcription factor 1 (Tbx1) in anterior second heart field cells that contribute to the outflow tract. It also responds to both NK2 transcription factor related, locus 5 (Nkx2-5) and ISL1 transcription factor, LIM/homeodomain (Islet1), acting through overlapping sites. Mutation of these sites reduces transgene expression in the anterior second heart field where the Fgf10 regulatory element is activated by Islet1 via direct binding in vivo. Analysis of the response to Nkx2-5 loss- and Isl1 gain-of-function genetic backgrounds indicates that the observed up-regulation of its activity in Nkx2-5 mutant hearts, reflecting that of Fgf10 , is due to the absence of Nkx2-5 repression and to up-regulation of Isl1 , normally repressed in the myocardium by Nkx2-5. ChIP experiments show strong binding of Nkx2-5 in differentiated myocardium. Molecular and genetic analysis of the Fgf10 cardiac element therefore reveals how key cardiac transcription factors orchestrate gene expression in the anterior second heart field and how genes, such as Fgf10 , normally expressed in the progenitor cell population, are repressed when these cells enter the heart and differentiate into myocardium. Our findings provide a paradigm for transcriptional mechanisms that underlie the changes in regulatory networks during the transition from progenitor state to that of the differentiated tissue.

Transcriptional heterogeneity of fibroblasts is a hallmark of the aging heart

Publisher: American Society for Clinical Investigation

Date: 14-11-2019

DOI: 10.1172/JCI.INSIGHT.131092

A gain-of-function TBX20 mutation causes congenital atrial septal defects, patent foramen ovale and cardiac valve defects

Publisher: BMJ

Date: 16-09-2009

DOI: 10.1136/JMG.2009.069997

Homeodomain Factor Nkx2-3 Controls Regional Expression of Leukocyte Homing Coreceptor MAdCAM-1 in Specialized Endothelial Cells of the Viscera

Publisher: Elsevier BV

Date: 11-2000

DOI: 10.1006/DBIO.2000.9958

Links in the Left/Right Axial Pathway

Publisher: Elsevier BV

Date: 08-1998

DOI: 10.1016/S0092-8674(00)81468-3

Abstract: We now have a sketch of the vertebrate L/R pathway from egg to organ. There is a lot to learn and a lot to explain, but the beauty of the pathway is already evident. We also have an inkling of its fragility. Set against the complex laterality disorders seen in humans, efforts to understand this pathway should continue to challenge our intellectual and experimental dexterity.

Targeted next-generation sequencing identifies pathogenic variants in familial congenital heart disease

Publisher: Elsevier BV

Date: 12-2014

DOI: 10.1016/J.JACC.2014.09.048

Abstract: Many genes have been implicated in the development of congenital heart disease (CHD). Next-generation sequencing offers opportunities for genetic testing but is often complicated by logistic and interpretative hurdles. This study sought to apply next-generation sequencing technology to CHD families with multiple affected members using a purpose-designed gene panel to assess diagnostic potential for future clinical applications. We designed a targeted next-generation sequencing gene panel for 57 genes previously implicated in CHD. Probands were screened in 16 families with strong CHD histories and in 15 control subjects. Variants affecting protein-coding regions were classified in silico using prediction programs and filtered according to predicted mode of inheritance, minor allele frequencies, and presence in databases such as dbSNP (Single Nucleotide Polymorphism Database) and ESP (Exome Sequencing Project). Disease segregation studies were conducted in variants identified in CHD cases predicted to be deleterious and with minor allele frequencies <0.1%. Thirteen potential disease-causing variants were identified in 9 families. Of these, 5 variants segregated with disease phenotype, revealing a likely molecular diagnosis in 31% of this cohort. Significant increases in the number of "indels, nonsense, and splice" variants, as well as variants classified as "probably damaging" were identified in CHD cases but not in control subjects. Also, there was a significant increase in the total number of "rare" and "low" frequency variants (minor allele frequencies <0.05) in the CHD cases. When multiple relatives are affected by CHD, a gene panel-based approach may identify its cause in up to 31% of families. Identifying causal variants has implications for clinical care and future family planning.

Precardiac deletion of Numb and Numblike reveals renewal of cardiac progenitors

Publisher: eLife Sciences Publications, Ltd

Date: 24-04-2014

DOI: 10.7554/ELIFE.02164

Abstract: Cardiac progenitor cells (CPCs) must control their number and fate to sustain the rapid heart growth during development, yet the intrinsic factors and environment governing these processes remain unclear. Here, we show that deletion of the ancient cell-fate regulator Numb (Nb) and its homologue Numblike (Nbl) depletes CPCs in second pharyngeal arches (PA2s) and is associated with an atrophic heart. With histological, flow cytometric and functional analyses, we find that CPCs remain undifferentiated and expansive in the PA2, but differentiate into cardiac cells as they exit the arch. Tracing of Nb- and Nbl-deficient CPCs by lineage-specific mosaicism reveals that the CPCs normally populate in the PA2, but lose their expansion potential in the PA2. These findings demonstrate that Nb and Nbl are intrinsic factors crucial for the renewal of CPCs in the PA2 and that the PA2 serves as a microenvironment for their expansion.

Platelet-derived growth factor-AB improves scar mechanics and vascularity after myocardial infarction

Publisher: American Association for the Advancement of Science (AAAS)

Date: 2020

DOI: 10.1126/SCITRANSLMED.AAY2140

Abstract: Recombinant human platelet-derived growth factor-AB improves cardiac function and survival after myocardial infarction in a porcine model.

Foxh1 Is Essential for Development of the Anterior Heart Field

Publisher: Elsevier BV

Date: 09-2004

DOI: 10.1016/J.DEVCEL.2004.07.023

Cardiac homeobox gene NKX2-5mutations and congenital heart disease

Publisher: Elsevier BV

Date: 06-2003

DOI: 10.1016/S0735-1097(03)00420-0

Abstract: We sought to examine the importance of mutations in the cardiac transcription factor gene NKX2-5 in patients with an atrial septal defect (ASD), patent foramen ovale (PFO), or hypoplastic left heart syndrome (HLHS). Mutations in NKX2-5 have been found in families showing secundum ASD and atrioventricular (AV) conduction block and in some in iduals with tetralogy of Fallot. The prevalence of NKX2-5 mutations in sporadic cases of ASD/PFO and other forms of congenital heart disease is unknown. A cohort of 146 in iduals with secundum ASD, PFO complicated by paradoxical embolism, or HLHS were evaluated. Patients with ASD or PFO were ascertained irrespective of family history or associated cardiac abnormalities. The coding region of the NKX2-5 locus was lified by polymerase chain reaction and sequenced. Among 102 ASD and 25 PFO patients screened, 13 patients (10%) had a positive family history and 5 patients (4%) had AV conduction block. We found one previously documented NKX2-5 missense mutation, T178M, in members of a family with ASD without AV conduction block. One NKX2-5 mutation-positive child from this family had HLHS, although no mutations were subsequently found in 18 patients with sporadic or familial HLHS. In a second ASD family without AV conduction block, we found a missense change, E21Q, previously reported as pathogenic. Because this change did not segregate with disease status, we propose that it is a non-disease-causing polymorphism. Our findings suggest that NKX2-5 mutations are a relatively infrequent cause of sporadic ASD and HLHS. Screening for NKX2-5 mutations may be warranted in in iduals with ASD and a positive family history, irrespective of the presence or absence of AV conduction block.

Cardiogenic Genes Expressed in Cardiac Fibroblasts Contribute to Heart Development and Repair

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 25-04-2014

DOI: 10.1161/CIRCRESAHA.114.302530

Abstract: Cardiac fibroblasts are critical to proper heart function through multiple interactions with the myocardial compartment, but appreciation of their contribution has suffered from incomplete characterization and lack of cell-specific markers. To generate an unbiased comparative gene expression profile of the cardiac fibroblast pool, identify and characterize the role of key genes in cardiac fibroblast function, and determine their contribution to myocardial development and regeneration. High-throughput cell surface and intracellular profiling of cardiac and tail fibroblasts identified canonical mesenchymal stem cell and a surprising number of cardiogenic genes, some expressed at higher levels than in whole heart. While genetically marked fibroblasts contributed heterogeneously to interstitial but not cardiomyocyte compartments in infarcted hearts, fibroblast-restricted depletion of one highly expressed cardiogenic marker, T-box 20, caused marked myocardial dysmorphology and perturbations in scar formation on myocardial infarction. The surprising transcriptional identity of cardiac fibroblasts, the adoption of cardiogenic gene programs, and direct contribution to cardiac development and repair provoke alternative interpretations for studies on more specialized cardiac progenitors, offering a novel perspective for reinterpreting cardiac regenerative therapies.

Murine cerberus homologue Cer1 maps to chromosome 4

Publisher: Elsevier BV

Date: 04-1998

DOI: 10.1006/GENO.1997.5249

Abstract: Fosfomycin is currently a viable option against urinary tract infections, particularly against extended-spectrum β-lactamases (ESBL)-producing

A tyrosine-rich domain within homeodomain transcription factor Nkx2-5 is an essential element in the early cardiac transcriptional regulatory machinery

Publisher: The Company of Biologists

Date: 04-2006

DOI: 10.1242/DEV.02305

Abstract: Homeodomain factor Nkx2-5 is a central component of the transcription factor network that guides cardiac development in humans, mutations in NKX2.5 lead to congenital heart disease (CHD). We have genetically defined a novel conserved tyrosine-rich domain (YRD) within Nkx2-5 that has co-evolved with its homeodomain. Mutation of the YRD did not affect DNA binding and only slightly diminished transcriptional activity of Nkx2-5 in a context-specific manner in vitro. However, the YRD was absolutely essential for the function of Nkx2-5 in cardiogenesis during ES cell differentiation and in the developing embryo. Furthermore, heterozygous mutation of all nine tyrosines to alanine created an allele with a strong dominant-negative-like activity in vivo: ES cell↔embryo chimaeras bearing the heterozygous mutation died before term with cardiac malformations similar to the more severe anomalies seen in NKX2.5 mutant families. These studies suggest a functional interdependence between the NK2 class homeodomain and YRD in cardiac development and evolution, and establish a new model for analysis of Nkx2-5 function in CHD.

Arrhythmia induced by spatiotemporal overexpression of calreticulin in the heart

Publisher: Elsevier BV

Date: 07-2007

DOI: 10.1016/J.YMGME.2007.02.003

Abstract: Calreticulin (CRT) is a Ca(2+)-binding protein of the endoplasmic reticulum essential for cardiac development. For further investigation of the functional mechanism of calreticulin, we generated transgenic mice with spatiotemporal overexpression of calreticulin using a cre-loxP system. To elucidate the role of the protein in cardiogenesis, we adopted Nkx2.5-cre mice for heart specific overexpression. The overexpression of calreticulin was associated with arrhythmia, chamber dilation and sudden death, as observed in 6- to 10-week-old mice. Furthermore, transgenic mice displayed marked edema at 7-weeks of age. RT-PCR analysis revealed that the expression of hyperpolerization-activated cyclic nucleotide-gated channel1 (HCN1), an essential component for cardiac pace maker activity, had receded in the heart of transgenic mice. In addition, the protein level of connexin40 (Cx40), connexin43 (Cx43), components of gap junction, and myocyte-enhancer factor (MEF) 2C, a cardiac-specific transcriptional factor, were reduced in the transgenic mice hearts. These findings suggest that calreticulin affects cardiac arrhythmia with disruption of cardiac signaling, such as the HCN family members, and with low levels of Cx40 and Cx43. Overepression of calreticulin also leads to a decreased protein level of MEF2C and this may cause changes in cardiac structure. Our findings support calreticulin being critical for normal heart function and structure. These mice are a useful model for the study of endoplasmic reticulum proteins, such as calreticulin, in various tissues.

H2A.F: an extremely variant histone H2A sequence expressed in the chicken embryo.

Publisher: Proceedings of the National Academy of Sciences

Date: 05-1983

DOI: 10.1073/PNAS.80.10.2819

Abstract: A cDNA clone bank has been constructed from chicken embryonic RNA. Clones hybridizing poorly to embryonic histone gene probes were selected as possible variant gene transcripts. The DNA sequence of one cDNA predicts an extremely variant H2A protein (H2A.F), which is 40% ergent from the most abundant H2A protein in chicken erythrocyte chromatin. The H2A.F gene is not highly conserved across large species barriers, but in the chicken there may be a family of linked genes. The H2A.F mRNA is approximately equal to 820 base pairs in length and, unlike most other histone mRNAs, is polyadenylylated. Significantly, the H2A.F transcript shows a limited tissue distribution in the chicken embryo.

Victor Chang Cardiac Research Institute

Location: Australia

View Organisation

Discovery Projects - Grant ID: DP0988507

Start Date: 2009

End Date: 12-2011

Amount: $460,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP0212000

Start Date: 2002

End Date: 12-2004

Amount: $231,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP0346726

Start Date: 2003

End Date: 12-2005

Amount: $240,000.00

Funder: Australian Research Council

Special Research Initiatives - Grant ID: SR0354500

Start Date: 2003

End Date: 12-2003

Amount: $10,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP160104858

Start Date: 2016

End Date: 12-2018

Amount: $467,400.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP190101475

Start Date: 2019

End Date: 12-2021

Amount: $545,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP210102134

Start Date: 08-2021

End Date: 12-2024

Amount: $804,100.00

Funder: Australian Research Council

Special Research Initiatives - Grant ID: SR0354622

Start Date: 11-2003

End Date: 03-2005

Amount: $20,000.00

Funder: Australian Research Council

Research Networks - Grant ID: RN0457079

Start Date: 09-2004

End Date: 08-2009

Amount: $1,500,000.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE110100092

Start Date: 04-2011

End Date: 03-2013

Amount: $300,000.00

Funder: Australian Research Council

Special Research Initiatives - Grant ID: SR1101002

Start Date: 07-2011

End Date: 12-2019

Amount: $21,000,000.00

Funder: Australian Research Council