Outflow Tract Of Heart Research Articles

Abstract 96: Epigenetic And Transcriptional Regulation Of Induced Pluripotent Stem Cells In Patients With Hypoplastic Left Heart Syndrome

Background: Although a number of studies have uncovered heterozygous mutations in cardiac regulatory genes caused hypoplastic left heart syndrome (HLHS), the identified genetic variants may not be directly correlated with the disease development. The aim of this study is to determine the epigenetic and transcriptional network responsible for myocardial patterning and morphogenesis during cardiac development in HLHS by using patient-derived induced pluripotent stem (iPS) cells. Methods: Five-independent iPS cell lines were generated from HLHS and biventricle (BV) heart-derived cardiac progenitor cells (CPCs). Global gene expression analysis, real-time RT-PCR, mutation analysis, ChIP assay, and cardiac-specific gene promoter activities were examined during differentiation. Results: We found one synonymous single nucleotide polymorphism in NKX2-5 and five in NOTCH1, respectively. Cardiac transcriptional factors such as NKX2-5, HAND1, and NOTCH/HEY, those are known to drive primary heart field and outflow tract development, were significantly reduced in HLHS-derived iPS cells after differentiation compared with BV- and control 201B7 iPS-derived cardiomyocytes. ChIP assay showed that a marked decrease in dimethylated histone H3-lysine 4 and acetylated histone H3 was found within the NKX2-5 promoter regions, whereas a significantly increased trimethylated H3-lysine 27 was identified in differentiated HLHS-derived iPS cells. To specify the target transcripts responsible for cardiac development of HLHS, cardiac troponin-T and natriuretic peptide A promoter analyses were performed. We found that both promoter activities were significantly suppressed in HLHS-derived CPCs and iPS cells compared with those from BV hearts. These repressed promoter activities could be fully restored by transient transfection of NKX2-5, HAND1, and NOTCH1 genes into these stem/progenitor cells by synergistic manner. Conclusions: These findings suggest that patient-specific iPS cells provide a potential to dissecting the complex cardiac malformations in human. The epigenetic and transcriptional regulation of NKX2-5, HAND1, and NOTCH1 may mutually participate in the development of myocardial growth, patterning, and morphogenesis in HLHS.

Driving vascular endothelial cell fate of human multipotent Isl1+ heart progenitors with VEGF modified mRNA

Distinct families of multipotent heart progenitors play a central role in the generation of diverse cardiac, smooth muscle and endothelial cell lineages during mammalian cardiogenesis. The identification of precise paracrine signals that drive the cell-fate decision of these multipotent progenitors, and the development of novel approaches to deliver these signals in vivo, are critical steps towards unlocking their regenerative therapeutic potential. Herein, we have identified a family of human cardiac endothelial intermediates located in outflow tract of the early human fetal hearts (OFT-ECs), characterized by coexpression of Isl1 and CD144/vWF. By comparing angiocrine factors expressed by the human OFT-ECs and non-cardiac ECs, vascular endothelial growth factor (VEGF)-A was identified as the most abundantly expressed factor, and clonal assays documented its ability to drive endothelial specification of human embryonic stem cell (ESC)-derived Isl1+ progenitors in a VEGF receptor-dependent manner. Human Isl1-ECs (endothelial cells differentiated from hESC-derived ISL1+ progenitors) resemble OFT-ECs in terms of expression of the cardiac endothelial progenitor- and endocardial cell-specific genes, confirming their organ specificity. To determine whether VEGF-A might serve as an in vivo cell-fate switch for human ESC-derived Isl1-ECs, we established a novel approach using chemically modified mRNA as a platform for transient, yet highly efficient expression of paracrine factors in cardiovascular progenitors. Overexpression of VEGF-A promotes not only the endothelial specification but also engraftment, proliferation and survival (reduced apoptosis) of the human Isl1+ progenitors in vivo. The large-scale derivation of cardiac-specific human Isl1-ECs from human pluripotent stem cells, coupled with the ability to drive endothelial specification, engraftment, and survival following transplantation, suggest a novel strategy for vascular regeneration in the heart.

A Rare Case of Left Ventricular Outflow Tract Mass

Thrombus formation in left ventricular outflow tract (LVOT) of a normal heart is a very rare occurrence. A 23-year-old male who presented with syncope, on evaluation found to have obstructing mass in the LVOT. His heart was otherwise normal. His investigations were not contributory except for significant eosinophilia. Due to recurrence of syncope he underwent emergency surgery for extraction of the mass, which on histopathological examination was found to be organizing thrombus. His eosinophil count normalized after the surgery. Tests for hypercoaguable states and investigations for known cause of eosinophilia were normal. There was no recurrence of thrombus or eosinophilia at 6 months after surgery. He was diagnosed to have obstructive LVOT thrombus in a normal heart secondary to transient eosinophilia. Presentation of this interesting case with literature on left ventricular thrombus and eosinophilia is discussed.

Evolution of the in‐series circulation in tetrapods: form follows function

The ancestral in‐parallel circulation of tetrapods combines a single ventricle and two systemic aortae, which allow central vascular shunting. Loss of the shunt and evolution of the in‐series circulatory design, as exemplified by extant mammals and birds, involved independent acquisition of two anatomic features: a complete interventricular septum, and a single aorta. Extent and location of the interventricular septum is dictated by the expression gradient of Tbx transcription factors in the myocardium. Loss of an aorta, however, has yet to be addressed by developmental studies. Our experimental work on the American alligator suggests that the heart's outflow tract is remodelled by changes in blood flow distribution in response to environmental stressors. Flow capacity of the left aorta (a conduit for the pulmonary bypass shunt) was significantly reduced (−27%) in alligator hatchlings chronically exposed for three months to atmospheric hypoxia (12%O2), when compared to conspecifics reared in normoxia (21%O2) or hyperoxia (30%O2). In contrast, outflow tract capacities did not differ in juvenile alligators subject to 15 months of different exercise regimina (sedentary, running or swimming). We suggest that a global hypoxic episode, rather than increased activity, may have been the driving pressure behind loss of the shunt and origin of the in‐series circulation. Funded by NSF IOB0445680 and IOS922756 to JWH.

Jun Is Required in Isl1-Expressing Progenitor Cells for Cardiovascular Development

Jun is a highly conserved member of the multimeric activator protein 1 transcription factor complex and plays an important role in human cancer where it is known to be critical for proliferation, cell cycle regulation, differentiation, and cell death. All of these biological functions are also crucial for embryonic development. Although all Jun null mouse embryos die at mid-gestation with persistent truncus arteriosus, a severe cardiac outflow tract defect also seen in human congenital heart disease, the developmental mechanisms are poorly understood. Here we show that murine Jun is expressed in a restricted pattern in several cell populations important for cardiovascular development, including the second heart field, pharyngeal endoderm, outflow tract and atrioventricular endocardial cushions and post-migratory neural crest derivatives. Several genes, including Isl1, molecularly mark the second heart field. Isl1 lineages include myocardium, smooth muscle, neural crest, endocardium, and endothelium. We demonstrate that conditional knockout mouse embryos lacking Jun in Isl1-expressing progenitors display ventricular septal defects, double outlet right ventricle, semilunar valve hyperplasia and aortic arch artery patterning defects. In contrast, we show that conditional deletion of Jun in Tie2-expressing endothelial and endocardial precursors does not result in aortic arch artery patterning defects or embryonic death, but does result in ventricular septal defects and a low incidence of semilunar valve defects, atrioventricular valve defects and double outlet right ventricle. Our results demonstrate that Jun is required in Isl1-expressing progenitors and, to a lesser extent, in endothelial cells and endothelial-derived endocardium for cardiovascular development but is dispensable in both cell types for embryonic survival. These data provide a cellular framework for understanding the role of Jun in the pathogenesis of congenital heart disease.

Hyperglycemia Slows Embryonic Growth and Suppresses Cell Cycle via Cyclin D1 and p21

In pregnant women, the diabetic condition results in a three- to fivefold increased risk for fetal cardiac malformations as a result of elevated glucose concentrations and the resultant osmotic stress in the developing embryo and fetus. Heart development before septation in the chick embryo was studied under two hyperglycemic conditions. Pulsed hyperglycemia induced by daily administration of glucose during 3 days of development caused daily spikes in plasma glucose concentration. In a second model, sustained hyperglycemia was induced with a single injection of glucose into the yolk on day 0. The sustained model raised the average plasma glucose concentration from 70 mg/dL to 180 mg/dL and led to decreased gene expression of glucose transporter GLUT1. Both models of hyperglycemia reduced embryo size, increased mortality, and delayed development. Within the heart outflow tract, reduced proliferation of myocardial and endocardial cells resulted from the sustained hyperglycemia and hyperosmolarity. The cell cycle inhibitor p21 was significantly increased, whereas cyclin D1, a cell cycle promoter, decreased in sustained hyperglycemia compared with controls. The evidence suggests that hyperglycemia-induced developmental delays are associated with slowed cell cycle progression, leading to reduced cellular proliferation. The suppression of critical developmental steps may underlie the cardiac defects observed during late gestation under hyperglycemic conditions.

Embryonic development of the bulbus arteriosus of the primitive heart of jawed vertebrates

Recent work has shown that the outflow tract (OFT) of the chondrichthyan heart does not consist of a single component, the conus arteriosus, as has been classically accepted, but two, the myocardial conus arteriosus and the non-myocardial bulbus arteriosus. The aim of this study was to describe the embryonic development of the bulbus, under the assumption that the heart of extant chondrichthyans represents the primitive condition of this organ in jawed vertebrates. The study was carried out in Atlantic sawtail catshark embryos belonging to developmental stages 20–34 of Ballard et al. (1993). They were examined using histochemical and immunohistochemical techniques for light and fluorescence microscopy. The entire OFT displays a myocardial character during early development. At stage 28, a new, non-myocardial segment forms at the distal end of the OFT. This segment, which is the precursor of the bulbus arteriosus, shows positive reactivity for 4,5-diaminofluorescein 2-diacetate throughout development from stage 31. Smooth muscle α-actin-positive cells appear in the prospective bulbus at stage 33; they markedly increase in number during stage 34. Subsequently, they extend backwards to overlap with the conal myocardium, giving rise to a myocardium and smooth muscle interface that has been described elsewhere as the middle component of the vertebrate OFT. Our embryological findings in a elasmobranch species are consistent with the notion that a second heart field has existed since the beginning of the jawed vertebrate radiation and that the bulbus arteriosus of chondrichthyans is a second heart field-derived structure.

Extracting cardiac shapes and motion of the chick embryo heart outflow tract from four-dimensional optical coherence tomography images

Recent advances in optical coherence tomography (OCT), and the development of image reconstruction algorithms, enabled four-dimensional (4-D) (three-dimensional imaging over time) imaging of the embryonic heart. To further analyze and quantify the dynamics of cardiac beating, segmentation procedures that can extract the shape of the heart and its motion are needed. Most previous studies analyzed cardiac image sequences using manually extracted shapes and measurements. However, this is time consuming and subject to inter-operator variability. Automated or semi-automated analyses of 4-D cardiac OCT images, although very desirable, are also extremely challenging. This work proposes a robust algorithm to semi automatically detect and track cardiac tissue layers from 4-D OCT images of early (tubular) embryonic hearts. Our algorithm uses a two-dimensional (2-D) deformable double-line model (DLM) to detect target cardiac tissues. The detection algorithm uses a maximum-likelihood estimator and was successfully applied to 4-D in vivo OCT images of the heart outflow tract of day three chicken embryos. The extracted shapes captured the dynamics of the chick embryonic heart outflow tract wall, enabling further analysis of cardiac motion.

In vivofunctional imaging of blood flow and wall strain rate in outflow tract of embryonic chick heart using ultrafast spectral domain optical coherence tomography

During cardiac development, the cardiac wall and flowing blood are two important cardiac tissues that constantly interact with each other. This dynamic interaction defines appropriate biomechanical environment to which the embryonic heart is exposed. Quantitative assessment of the dynamic parameters of wall tissues and blood flow is required to further our understanding of cardiac development. We report the use of an ultrafast 1310-nm dual-camera spectral domain optical coherence tomography (SDOCT) system to characterize/image, in parallel, the dynamic radial strain rate of the myocardial wall and the Doppler velocity of the underlying flowing blood within an in vivo beating chick embryo. The OCT system operates at 184-kHz line scan rate, providing the flexibility of imaging the fast blood flow and the slow tissue deformation within one scan. The ability to simultaneously characterize tissue motion and blood flow provides a useful approach to better understand cardiac dynamics during early developmental stages.

Biomechanics of the Chick Embryonic Heart Outflow Tract at HH18 Using 4D Optical Coherence Tomography Imaging and Computational Modeling

During developmental stages, biomechanical stimuli on cardiac cells modulate genetic programs, and deviations from normal stimuli can lead to cardiac defects. Therefore, it is important to characterize normal cardiac biomechanical stimuli during early developmental stages. Using the chicken embryo model of cardiac development, we focused on characterizing biomechanical stimuli on the Hamburger–Hamilton (HH) 18 chick cardiac outflow tract (OFT), the distal portion of the heart from which a large portion of defects observed in humans originate. To characterize biomechanical stimuli in the OFT, we used a combination of in vivo optical coherence tomography (OCT) imaging, physiological measurements and computational fluid dynamics (CFD) modeling. We found that, at HH18, the proximal portion of the OFT wall undergoes larger circumferential strains than its distal portion, while the distal portion of the OFT wall undergoes larger wall stresses. Maximal wall shear stresses were generally found on the surface of endocardial cushions, which are protrusions of extracellular matrix onto the OFT lumen that later during development give rise to cardiac septa and valves. The non-uniform spatial and temporal distributions of stresses and strains in the OFT walls provide biomechanical cues to cardiac cells that likely aid in the extensive differential growth and remodeling patterns observed during normal development.

Maternal occupation and the risk of major birth defects: A follow-up analysis from the National Birth Defects Prevention Study

This study further examined the association between selected maternal occupations and a variety of birth defects identified from prior analysis and explored the effect of work hours and number of jobs held and potential interaction between folic acid and occupation. Data from a population-based, multi-center case-control study was used. Analyses included 45 major defects and specific sub-occupations under five occupational groups: healthcare workers, cleaners, scientists, teachers and personal service workers. Both logistic regression and Bayesian models (to minimize type-1 errors) were used, adjusted for potential confounders. Effect modification by folic acid was also assessed. More than any other occupation, nine different defects were positively associated with maids or janitors [odds ratio (OR) range: 1.72-3.99]. Positive associations were also seen between the following maternal occupations and defects in their children (OR range: 1.35-3.48): chemists/conotruncal heart and neural tube defects (NTDs), engineers/conotruncal defects, preschool teachers/cataracts and cleft lip with/without cleft palate (CL/P), entertainers/athletes/gastroschisis, and nurses/hydrocephalus and left ventricular outflow tract heart defects. Non-preschool teachers had significantly lower odds of oral clefts and gastroschisis in their offspring (OR range: 0.53-0.76). There was a suggestion that maternal folic acid use modified the effects with occupations including lowering the risk of NTDs and CL/P. No consistent patterns were found between maternal work hours or multiple jobs by occupation and the risk of birth defects. Overall, mothers working as maids, janitors, biologists, chemists, engineers, nurses, entertainers, child care workers and preschool teachers had increased risks of several malformations and non-preschool teachers had a lower risk of some defects. Maternal folic acid use reduced the odds of NTDs and CL/P among those with certain occupations. This hypothesis-generating study will provide clues for future studies with better exposure data.

Absolute Blood Flow Measurement of early Stage Chicken Embryo Heart Using High Speed 4D Optical Coherence Tomography

The measurement of blood-plasma absolute velocity distributions with spatial and temporal resolution in vivo is important for research on early stage embryo heart development. We introduce a novel method to measure absolute blood flow velocity based on high speed spectral domain Optical Coherence Tomography (OCT) and applied it to measure velocities on the heart outflow tract (OFT) of a chicken embryo (stage HH18).

Computational Fluid Dynamics of Developing Avian Outflow Tract Heart Valves

Hemodynamic forces play an important role in sculpting the embryonic heart and its valves. Alteration of blood flow patterns through the hearts of embryonic animal models lead to malformations that resemble some clinical congenital heart defects, but the precise mechanisms are poorly understood. Quantitative understanding of the local fluid forces acting in the heart has been elusive because of the extremely small and rapidly changing anatomy. In this study, we combine multiple imaging modalities with computational simulation to rigorously quantify the hemodynamic environment within the developing outflow tract (OFT) and its eventual aortic and pulmonary valves. In vivo Doppler ultrasound generated velocity profiles were applied to Micro-Computed Tomography generated 3D OFT lumen geometries from Hamburger-Hamilton (HH) stage 16-30 chick embryos. Computational fluid dynamics simulation initial conditions were iterated until local flow profiles converged with in vivo Doppler flow measurements. Results suggested that flow in the early tubular OFT (HH16 and HH23) was best approximated by Poiseuille flow, while later embryonic OFT septation (HH27, HH30) was mimicked by plug flow conditions. Peak wall shear stress (WSS) values increased from 18.16 dynes/cm(2) at HH16 to 671.24 dynes/cm(2) at HH30. Spatiotemporally averaged WSS values also showed a monotonic increase from 3.03 dynes/cm(2) at HH16 to 136.50 dynes/cm(2) at HH30. Simulated velocity streamlines in the early heart suggest a lack of mixing, which differed from classical ink injections. Changes in local flow patterns preceded and correlated with key morphogenetic events such as OFT septation and valve formation. This novel method to quantify local dynamic hemodynamics parameters affords insight into sculpting role of blood flow in the embryonic heart and provides a quantitative baseline dataset for future research.

Homocysteine Exposure Affects Early Hemodynamic Parameters of Embryonic Chicken Heart Function

Maternal hyperhomocysteinemia has been associated with an increased risk of newborns with a congenital heart defect. This has been substantiated in the chicken embryo, as congenital heart defects have been induced after homocysteine treatment. Comparable heart defects are observed in venous clipping studies, a model of altered embryonic blood flow. Because of this overlap in heart defects, our aim was to test the hypothesis that homocysteine would cause alterations in embryonic heart function that precede the structural malformations previously described. Therefore, Doppler flow velocity waveforms were recorded in both primitive ventricles and the outflow tract of the embryonic heart of homocysteine treated and control chicken embryos at embryonic day 3.5. Homocysteine treatment consisted of 50 μL 0.05 M L-homocysteine thiolactone at 24, 48, and 72 hr. Homocysteine-treated embryos displayed significantly lower mean heart rates of 134 (SD 22) bpm, compared to 150 (14) bpm in control embryos. Homocysteine treatment caused an inhibiting effect on hemodynamic parameters, and altered heart function was presented by a shift in the proportions of the different wave times in percentage of total cycle time. Homocysteine induces changes in hemodynamic parameters of early embryonic chicken heart function. These changes may precede morphological changes and contribute to the development of CHD defects through alterations in shear stress and shear stress related genes, as seen before in venous clipping studies.

344 Molecular patterning of the cardiac outflow tract and coronary arteries of the mouse heart

344 Molecular patterning of the cardiac outflow tract and coronary arteries of the mouse heart

Cardiac outflow and wall motion in hypothermic chick embryos

Cardiac outflow in the early developmental stage of a chick embryo is known to be highly variable depending on environmental temperature. To investigate the effects of environmental hypothermia on the blood flow in the outflow tract (OFT) of chick embryonic hearts, microscopic flow images were consecutively captured from chick embryos at HH stage 17 (2.5days of incubation) at room temperature. Instantaneous velocity field information of blood flow in OFT was obtained using a micro-particle image velocimetry technique. The cyclic variations of the OFT vessel diameter and wall thickness were simultaneously measured. The experimental results show that environmental hypothermia causes bradycardia with a decrease in peak velocity during systole and the occurrence of backflow during diastole in the OFT. These abnormal phenomena seem to be attributed to the suppression of myocardial wall motion under hypothermic conditions.

Quantifying blood flow and wall shear stresses in the outflow tract of chick embryonic hearts

Wall shear stresses (WSS) exerted by blood flow on cardiac tissues modulate growth and development of the heart. To study the role of hemodynamic conditions on cardiac morphogenesis, here, we present a methodology that combines imaging and finite element modeling to quantitate the in vivo blood flow dynamics and WSS in the cardiac outflow tract (OFT) of early chicken embryos (day 3 out of 21-day incubation). We found a distinct blood flow field and heterogeneous distribution of WSS in the chicken embryonic heart OFT, which have physiological implications for OFT morphogenesis.

Lipomatosis of interventricular septum and both ventricles: an extremely rare pattern

Fatty masses of the heart are relatively uncommon. This report is about an extremely rare case of cardiac lipomatosis that implicates both ventricles and inter-atrial septum. The patient was a 27-year-old female who was accidentally diagnosed during a routine physical examination. Trans-thoracic echocardiography (TTE) discovered a large homogenous bi-lobed, non-encapsulated echogenic mass. Magnetic resonance imaging (MRI) showed soft tissue abnormal enhancement in the anterior left base of the heart and sub-aortic outflow tract. A trans-jugular echomyocardial biopsy was done because of a lack of a definite diagnosis by non-invasive techniques. Pathological study showed benign fatty infiltration suggestive for benign lipomatous hypertrophy.

Common cause of cardiogenic syncope and emergency treatment

Cardiogenic syncope is the relatively common type of syncope in children. The state of illness is more serious at the onset,the prognosis of partial patients is very dangerous,even it can cause cardiac sudden death, so clinical doctors pay close attention to cardiogenic syncope. The common pathogeny of cardiogenic syncope in children includes serious arrhythmia, obstruction of the outflow tract of heart, restricted diastole of heart or weak systole of heart. Before the particular cause of disease is determined,some examinations and laboratory tests should be made. In this article, the writer introduces detailed methods of the emergent treatment. Key words: Cardiogenic syncope; Etiology; Treatment

P08.07: Ectopia cordis: association with amniotic band syndrome in seven cases

Objectives: To identify the outflow tracts of the fetal heart using the STIC method (spatiotemporal image correlation) in the 11–13 week scan. Methods: We performed 30 ultrasound examinations of pregnant women between 11 and 13 weeks from november 2009 to february 2010 at the Center for Fetal Medicine of Campinas (CENTRUS), according to the screening protocol in the first trimester traced by the Fetal Medicine Foundation using the HD11 (Philips). After the morphological analysis it was assessed the fetal heart following the methodology described by Paladini in 2008. We identified the heart through the four-chamber view and after activation of the color Doppler the STIC method was performed. It was then carried out post-processing, making it possible to identify the cardiac atria and the position of the aorta and pulmonary arteries, and the relationships among these structures. Results: A total of 30 women between 11 and 13 weeks. Six women (20.0%) had a gestational age of 11 weeks, 13 (43.3%) 12 weeks and the remaining 11 (36.7%) women had 13 weeks of gestation. The crow rump length averaged 64.3 mm (47.3 to 82.3 mm), with standard deviation of 9.4 mm. The mean nuchal translucency thickness was 1.9 mm (1.2 to 5.0 mm) with a standard deviation of 0.7 mm. Examination showed no regurgitation of the tricuspid valve. The nasal bone was present in 25 (83.3%) samples. The visualization of the outflow tract and their relationship to the cardiac atria occurred in 19 (63.3%) women. Inappropriate positioning of the fetus (7 cases), Doppler acquisition of poor quality (2 cases) and the impossibility of assessing the aorta (2 cases) were the reasons for not identifying the correct exit routes. Conclusions: The application of ultrasound examination suggested by the methodology described by Paladini proved to be feasible within the screening period of the first trimester, antecipating the identification of the outflow tract of the fetal heart.