Risk factors associated with the development of double-inlet ventricle congenital heart disease.
Birth defects research
Beyond Gene Panels: Whole Exome Sequencing for Diagnosis of Congenital Heart Disease.
Circulation. Genomic and precision medicine
2018; 11 (3): e002097
A Temporal Chromatin Signature in Human Embryonic Stem Cells Identifies Regulators of Cardiac Development
2012; 151 (1): 221-232
BACKGROUND: Congenital heart disease (CHD) is the most common birth defect group and a significant contributor to neonatal and infant death. CHD with single ventricle anatomy, including hypoplastic left heart syndrome (HLHS), tricuspid atresia (TA), and various double-inlet ventricle (DIV) malformations, is the most complex with the highest mortality. Prenatal risk factors associated with HLHS have been studied, but such data for DIV are lacking.METHODS: We analyzed DIV cases and nonmalformed controls in the National Birth Defects Prevention Study, a case-control, multicenter population-based study of birth defects. Random forest analysis identified potential predictor variables for DIV, which were included in multivariable models to estimate effect magnitude and directionality.RESULTS: Random forest analysis identified pre-pregnancy diabetes, history of maternal insulin use, maternal total lipid intake, paternal race, and intake of several foods and nutrients as potential predictors of DIV. Logistic regression confirmed pre-pregnancy diabetes, maternal insulin use, and paternal race as risk factors for having a child with DIV. Additionally, higher maternal total fat intake was associated with a reduced risk.CONCLUSIONS: Maternal pre-pregnancy diabetes and history of insulin use were associated with an increased risk of having an infant with DIV, while maternal lipid intake had an inverse association. These novel data provide multiple metabolic pathways for investigation to identify better the developmental etiologies of DIV and suggest that public health interventions targeting diabetes prevention and management in women of childbearing age could reduce CHD risk.
View details for PubMedID 30920163
Endogenous Wnt/beta-Catenin Signaling Is Required for Cardiac Differentiation in Human Embryonic Stem Cells
2010; 5 (6)
Directed differentiation of human embryonic stem cells (ESCs) into cardiovascular cells provides a model for studying molecular mechanisms of human cardiovascular development. Although it is known that chromatin modification patterns in ESCs differ markedly from those in lineage-committed progenitors and differentiated cells, the temporal dynamics of chromatin alterations during differentiation along a defined lineage have not been studied. We show that differentiation of human ESCs into cardiovascular cells is accompanied by programmed temporal alterations in chromatin structure that distinguish key regulators of cardiovascular development from other genes. We used this temporal chromatin signature to identify regulators of cardiac development, including the homeobox gene MEIS2. Using the zebrafish model, we demonstrate that MEIS2 is critical for proper heart tube formation and subsequent cardiac looping. Temporal chromatin signatures should be broadly applicable to other models of stem cell differentiation to identify regulators and provide key insights into major developmental decisions.
View details for DOI 10.1016/j.cell.2012.08.027
View details for Web of Science ID 000309544200022
View details for PubMedID 22981225
View details for PubMedCentralID PMC3462257
Multi-disciplinary evaluation of a 5-month-old with hypertrophic cardiomyopathy related to a functional adrenocortical tumor.
Journal of pediatric endocrinology & metabolism : JPEM
Wnt/beta-catenin signaling is an important regulator of differentiation and morphogenesis that can also control stem cell fates. Our group has developed an efficient protocol to generate cardiomyocytes from human embryonic stem (ES) cells via induction with activin A and BMP4.We tested the hypothesis that Wnt/beta-catenin signals control both early mesoderm induction and later cardiac differentiation in this system. Addition of exogenous Wnt3a at the time of induction enhanced cardiac differentiation, while early inhibition of endogenous Wnt/beta-catenin signaling with Dkk1 inhibited cardiac differentiation, as indicated by quantitative RT-PCR analysis for beta-myosin heavy chain (beta-MHC), cardiac troponin T (cTnT), Nkx2.5, and flow cytometry analysis for sarcomeric myosin heavy chain (sMHC). Conversely, late antagonism of endogenously produced Wnts enhanced cardiogenesis, indicating a biphasic role for the pathway in human cardiac differentiation. Using quantitative RT-PCR, we show that canonical Wnt ligand expression is induced by activin A/BMP4 treatment, and the extent of early Wnt ligand expression can predict the subsequent efficiency of cardiogenesis. Measurement of Brachyury expression showed that addition of Wnt3a enhances mesoderm induction, whereas blockade of endogenously produced Wnts markedly inhibits mesoderm formation. Finally, we show that Wnt/beta-catenin signaling is required for Smad1 activation by BMP4.Our data indicate that induction of mesoderm and subsequent cardiac differentiation from human ES cells requires fine-tuned cross talk between activin A/BMP4 and Wnt/beta-catenin pathways. Controlling these pathways permits efficient generation of cardiomyocytes for basic studies or cardiac repair applications.
View details for DOI 10.1371/journal.pone.0011134
View details for Web of Science ID 000278775900023
View details for PubMedID 20559569
View details for PubMedCentralID PMC2886114
Cardiac Regeneration Lessons From Development
2017; 120 (6): 941-959
Background Hypertrophic cardiomyopathy (HCM) in childhood is a rare diagnosis, and associations with adrenocortical tumors (ACTs) have been rarely reported in the pediatric literature. Case Presentation We present a case of a 5-month-old who presented with HCM and during the evaluation for hypertension was found to have elevated glucocorticoids, mineralocorticoids, androgens and urine metanephrines. During preoperative evaluation, he developed shock followed by cardiogenic collapse requiring extracorporeal membrane oxygenation (ECMO); however, he did not survive. Pathology revealed an ACT with hormone production that contributed to his demise. Conclusion Adrenocortical tumors associated with hypertrophic cardiomyopathy can be life-threatening. We discuss the complex interplay of unrestricted cortical hormone production in the setting of hypertrophic cardiomyopathy that may lead to rapid decline and poor clinical outcomes.
View details for PubMedID 30352041
Nkx2.5+ Cardiomyoblasts Contribute to Cardiomyogenesis in the Neonatal Heart.
2017; 7 (1): 12590
Palliative surgery for congenital heart disease has allowed patients with previously lethal heart malformations to survive and, in most cases, to thrive. However, these procedures often place pressure and volume loads on the heart, and over time, these chronic loads can cause heart failure. Current therapeutic options for initial surgery and chronic heart failure that results from failed palliation are limited, in part, by the mammalian heart's low inherent capacity to form new cardiomyocytes. Surmounting the heart regeneration barrier would transform the treatment of congenital, as well as acquired, heart disease and likewise would enable development of personalized, in vitro cardiac disease models. Although these remain distant goals, studies of heart development are illuminating the path forward and suggest unique opportunities for heart regeneration, particularly in fetal and neonatal periods. Here, we review major lessons from heart development that inform current and future studies directed at enhancing cardiac regeneration.
View details for DOI 10.1161/CIRCRESAHA.116.309040
View details for Web of Science ID 000397330700007
View details for PubMedID 28302741
Comparison of Human Embryonic Stem Cell-Derived Cardiomyocytes, Cardiovascular Progenitors, and Bone Marrow Mononuclear Cells for Cardiac Repair
STEM CELL REPORTS
2015; 5 (5): 753-762
During normal lifespan, the mammalian heart undergoes limited renewal of cardiomyocytes. While the exact mechanism for this renewal remains unclear, two possibilities have been proposed: differentiated myocyte replication and progenitor/immature cell differentiation. This study aimed to characterize a population of cardiomyocyte precursors in the neonatal heart and to determine their requirement for cardiac development. By tracking the expression of an embryonic Nkx2.5 cardiac enhancer, we identified cardiomyoblasts capable of differentiation into striated cardiomyocytes in vitro. Genome-wide expression profile of neonatal Nkx2.5+ cardiomyoblasts showed the absence of sarcomeric gene and the presence of cardiac transcription factors. To determine the lineage contribution of the Nkx2.5+ cardiomyoblasts, we generated a doxycycline suppressible Cre transgenic mouse under the regulation of the Nkx2.5 enhancer and showed that neonatal Nkx2.5+ cardiomyoblasts mature into cardiomyocytes in vivo. Ablation of neonatal cardiomyoblasts resulted in ventricular hypertrophy and dilation, supporting a functional requirement of the Nkx2.5+ cardiomyoblasts. This study provides direct lineage tracing evidence that a cardiomyoblast population contributes to cardiogenesis in the neonatal heart. The cell population identified here may serve as a promising therapeutic for pediatric cardiac regeneration.
View details for PubMedID 28974782
Mechanical Stress Promotes Maturation of Human Myocardium From Pluripotent Stem Cell-Derived Progenitors
2015; 33 (7): 2148-2157
Cardiomyocytes derived from human embryonic stem cells (hESC-CMs) can improve the contractility of injured hearts.We hypothesized that mesodermal cardiovascular progenitors (hESC-CVPs), capable of generating vascular cells in addition to cardiomyocytes, would provide superior repair by contributing to multiple components of myocardium. We performed a head-to-head comparison of hESC-CMs and hESC-CVPs and compared these with the most commonly used clinical cell type, human bone marrow mononuclear cells (hBMMNCs). In a nude rat model of myocardial infarction, hESC-CMs and hESC-CVPs generated comparable grafts. Both similarly improved systolic function and ventricular dilation. Furthermore, only rare human vessels formed from hESC-CVPs. hBM-MNCs attenuated ventricular dilation and enhanced host vascularization without engrafting long-term or improving contractility. Thus, hESC-CMs and CVPs show similar efficacy for cardiac repair, and both are more efficient than hBM-MNCs. However, hESC-CVPs do not form larger grafts or more significant numbers of human vessels in the infarcted heart.
View details for DOI 10.1016/j.stemcr.2015.09.011
View details for Web of Science ID 000364991000008
View details for PubMedID 26607951
View details for PubMedCentralID PMC4649260
Molecular Regulation of Cardiomyocyte Differentiation
2015; 116 (2): 341-353
Recent advances in pluripotent stem cell biology and directed differentiation have identified a population of human cardiovascular progenitors that give rise to cardiomyocytes, smooth muscle, and endothelial cells. Because the heart develops from progenitors in 3D under constant mechanical load, we sought to test the effects of a 3D microenvironment and mechanical stress on differentiation and maturation of human cardiovascular progenitors into myocardial tissue. Progenitors were derived from embryonic stem cells, cast into collagen hydrogels, and left unstressed or subjected to static or cyclic mechanical stress. Compared to 2D culture, the unstressed 3D environment increased cardiomyocyte numbers and decreased smooth muscle numbers. Additionally, 3D culture suppressed smooth muscle α-actin content, suggesting diminished cell maturation. Cyclic stress-conditioning increased expression of several cardiac markers, including β-myosin heavy chain and cardiac troponin T, and the tissue showed enhanced calcium dynamics and force production. There was no effect of mechanical loading on cardiomyocyte or smooth muscle specification. Thus, 3D growth conditions favor cardiac differentiation from cardiovascular progenitors, whereas 2D conditions promote smooth muscle differentiation. Mechanical loading promotes cardiomyocyte structural and functional maturation. Culture in 3-D facilitates understanding how cues such as mechanical stress affect the differentiation and morphogenesis of distinct cardiovascular cell populations into organized, functional human cardiovascular tissue. Stem Cells 2015;33:2148-2157.
View details for DOI 10.1002/stem.2036
View details for Web of Science ID 000356668200007
View details for PubMedID 25865043
View details for PubMedCentralID PMC4478130
Engineered Biomaterials Control Differentiation and Proliferation of Human-Embryonic-Stem-Cell-Derived Cardiomyocytes via Timed Notch Activation
STEM CELL REPORTS
2014; 2 (3): 271-281
The heart is the first organ to form during embryonic development. Given the complex nature of cardiac differentiation and morphogenesis, it is not surprising that some form of congenital heart disease is present in ≈1 percent of newborns. The molecular determinants of heart development have received much attention over the past several decades. This has been driven in large part by an interest in understanding the causes of congenital heart disease coupled with the potential of using knowledge from developmental biology to generate functional cells and tissues that could be used for regenerative medicine purposes. In this review, we highlight the critical signaling pathways and transcription factor networks that regulate cardiomyocyte lineage specification in both in vivo and in vitro models. Special focus will be given to epigenetic regulators that drive the commitment of cardiomyogenic cells from nascent mesoderm and their differentiation into chamber-specific myocytes, as well as regulation of myocardial trabeculation.
View details for DOI 10.1161/CIRCRESAHA.116.302752
View details for Web of Science ID 000347939000019
View details for PubMedCentralID PMC4299877
Transmembrane protein 88: a Wnt regulatory protein that specifies cardiomyocyte development
2013; 140 (18): 3799-3808
For cell-based treatments of myocardial infarction, a better understanding of key developmental signaling pathways and more robust techniques for producing cardiomyocytes are required. Manipulation of Notch signaling has promise as it plays an important role during cardiovascular development, but previous studies presented conflicting results that Notch activation both positively and negatively regulates cardiogenesis. We developed surface- and microparticle-based Notch-signaling biomaterials that function in a time-specific activation-tunable manner, enabling precise investigation of Notch activation at specific developmental stages. Using our technologies, a biphasic effect of Notch activation on cardiac differentiation was found: early activation in undifferentiated human embryonic stem cells (hESCs) promotes ectodermal differentiation, activation in specified cardiovascular progenitor cells increases cardiac differentiation. Signaling also induces cardiomyocyte proliferation, and repeated doses of Notch-signaling microparticles further enhance cardiomyocyte population size. These results highlight the diverse effects of Notch activation during cardiac development and provide approaches for generating large quantities of cardiomyocytes.
View details for DOI 10.1016/j.stemcr.2014.01.011
View details for Web of Science ID 000336647700004
View details for PubMedID 24672751
View details for PubMedCentralID PMC3964284
Developmental Fate and Cellular Maturity Encoded in Human Regulatory DNA Landscapes
2013; 154 (4): 888-903
Genetic regulation of the cell fate transition from lateral plate mesoderm to the specification of cardiomyocytes requires suppression of Wnt/β-catenin signaling, but the mechanism for this is not well understood. By analyzing gene expression and chromatin dynamics during directed differentiation of human embryonic stem cells (hESCs), we identified a suppressor of Wnt/β-catenin signaling, transmembrane protein 88 (TMEM88), as a potential regulator of cardiovascular progenitor cell (CVP) specification. During the transition from mesoderm to the CVP, TMEM88 has a chromatin signature of genes that mediate cell fate decisions, and its expression is highly upregulated in advance of key cardiac transcription factors in vitro and in vivo. In early zebrafish embryos, tmem88a is expressed broadly in the lateral plate mesoderm, including the bilateral heart fields. Short hairpin RNA targeting of TMEM88 during hESC cardiac differentiation increases Wnt/β-catenin signaling, confirming its role as a suppressor of this pathway. TMEM88 knockdown has no effect on NKX2.5 or GATA4 expression, but 80% of genes most highly induced during CVP development have reduced expression, suggesting adoption of a new cell fate. In support of this, analysis of later stage cell differentiation showed that TMEM88 knockdown inhibits cardiomyocyte differentiation and promotes endothelial differentiation. Taken together, TMEM88 is crucial for heart development and acts downstream of GATA factors in the pre-cardiac mesoderm to specify lineage commitment of cardiomyocyte development through inhibition of Wnt/β-catenin signaling.
View details for DOI 10.1242/dev.094789
View details for Web of Science ID 000323698100009
View details for PubMedID 23924634
View details for PubMedCentralID PMC3754478
Cardiogenesis From Human Embryonic Stem Cells - Mechanisms and Applications
2010; 74 (12): 2517-2526
Cellular-state information between generations of developing cells may be propagated via regulatory regions. We report consistent patterns of gain and loss of DNase I-hypersensitive sites (DHSs) as cells progress from embryonic stem cells (ESCs) to terminal fates. DHS patterns alone convey rich information about cell fate and lineage relationships distinct from information conveyed by gene expression. Developing cells share a proportion of their DHS landscapes with ESCs; that proportion decreases continuously in each cell type as differentiation progresses, providing a quantitative benchmark of developmental maturity. Developmentally stable DHSs densely encode binding sites for transcription factors involved in autoregulatory feedback circuits. In contrast to normal cells, cancer cells extensively reactivate silenced ESC DHSs and those from developmental programs external to the cell lineage from which the malignancy derives. Our results point to changes in regulatory DNA landscapes as quantitative indicators of cell-fate transitions, lineage relationships, and dysfunction.
View details for DOI 10.1016/j.cell.2013.07.020
View details for Web of Science ID 000323202500018
View details for PubMedID 23953118
View details for PubMedCentralID PMC3962256
Chromatin remodeling during mouse and human embryonic stem cell differentiation
2008; 237 (5): 1389-1398
Over the past decade, the ability to culture and differentiate human embryonic stem cells (ESCs) has offered researchers a novel therapeutic that may, for the first time, repair regions of the damaged heart. Studies of cardiac development in lower organisms have led to identification of the transforming growth factor-β superfamily (eg, activin A and bone morphogenic protein 4) and the Wnt/β-catenin pathway as key inducers of mesoderm and cardiovascular differentiation. These factors act in a context-specific manner (eg, Wnt/β-catenin is required initially to form mesoderm but must be antagonized thereafter to make cardiac muscle). Different lines of ESCs produce different levels of agonists and antagonists for these pathways, but with careful optimization, highly enriched populations of immature cardiomyocytes can be generated. These cardiomyocytes survive transplantation to infarcted hearts of experimental animals, where they create new human myocardial tissue and improve heart function. The grafts generated by cell transplantation have been small, however, leading to an exploration of tissue engineering as an alternate strategy. Engineered tissue generated from preparations of human cardiomyocytes survives poorly after transplantation, most likely because of ischemia. Creation of pre-organized vascular networks in the tissue markedly enhances survival, with human capillaries anastomosed to the host coronary circulation. Thus, pathways controlling formation of the human cardiovascular system are emerging, yielding the building blocks for tissue regeneration that may address the root causes of heart failure.
View details for DOI 10.1253/circj.CJ-10-0958
View details for Web of Science ID 000284861400002
View details for PubMedID 21084757
View details for PubMedCentralID PMC3938118
Embryonic stem cell (ESC) differentiation is an excellent model to study chromatin changes at developmentally regulated loci. Differentiating mouse and human ESCs increase genome-wide acetylation (euchromatic) and tri-methylation (heterochromatic) of lysine 9 on histone H3. The Oct4 locus is euchromatic when expressed in undifferentiated ESCs and heterochromatic after differentiation. Brachyury T, a mesoderm-specific transcription factor, is not yet expressed in undifferentiated cells, where its locus has "bivalent" tri-methyl lysine 4 and lysine 27 modifications. During directed differentiation to pre-cardiac mesoderm, the activated brachyury locus has high levels of tri-methyl lysine 4 (euchromatin), switching to heterochromatin after gene silencing. Thus, ESC differentiation is accompanied by genome-wide commitment to euchromatin or heterochromatin. Undifferentiated hESCs bivalently modify the brachyury locus, activate it to euchromatin during mesoderm induction, and subsequently repress it to heterochromatin, demonstrating, to our knowledge, the first analysis of chromatin dynamics at a locus essential for mesoderm and endoderm differentiation.
View details for DOI 10.1002/dvdy.21545
View details for Web of Science ID 000255842900015
View details for PubMedID 18425849
View details for PubMedCentralID PMC3075915