Evolutionary Aspects of Chamber Formation and Septation.

The formed hearts of vertebrates are widely different in anatomy and performance, yet their embryonic hearts are surprisingly similar. Developmental and molecular biology are making great advances in reconciling these differences by revealing an evolutionarily conserved building plan to the vertebrate heart. This suggests that perspectives from evolution may improve our understanding of the formation of the human heart. Here, we exemplify this approach by discussing atrial and ventricular septation and the associated processes of remodeling of the atrioventricular junction and formation of the atrioventricular insulating plane.

Equal force generation potential of trabecular and compact wall ventricular cardiomyocytes.

Trabecular myocardium makes up most of the ventricular wall of the human embryo. A process of compaction in the fetal period presumably changes ventricular wall morphology by converting ostensibly weaker trabecular myocardium into stronger compact myocardium. Using developmental series of embryonic and fetal humans, mice and chickens, we show ventricular morphogenesis is driven by differential rates of growth of trabecular and compact layers rather than a process of compaction. In mouse, fetal cardiomyocytes are relatively weak but adult cardiomyocytes from the trabecular and compact layer show an equally large force generating capacity. In fetal and adult humans, trabecular and compact myocardium are not different in abundance of immunohistochemically detected vascular, mitochondrial and sarcomeric proteins. Similar findings are made in human excessive trabeculation, a congenital malformation. In conclusion, trabecular and compact myocardium is equally equipped for force production and their proportions are determined by differential growth rates rather than by compaction.

Congenital systemic arteriovenous fistulas: Interventional strategies and embryological perspectives.

Background: Data on congenital systemic arteriovenous fistulas are largely based on individual case reports. A true systemic arteriovenous fistula needs to be differentiated from other vascular malformations like capillary or venous hemangiomas, which are far more common. Objectives: We sought to identify the varied symptoms, diagnostic challenges, describe interventional treatment options, and postulate an embryological basis for this uncommonly described entity. Methods: This is a descriptive study of a cohort of systemic arteriovenous fistulas seen in the department of pediatric cardiology at a tertiary cardiac institute from 2010 to 2020, with prospective medium-term follow-up. A total of seven cases were identified. The diagnosis was confirmed by computed tomographic imaging, magnetic resonance angiography, or conventional angiography. Results: All were successfully closed using duct occluders or embolization coils with no recurrence in six cases over a median duration of follow-up of 48 months (interquartile range: 16; 36-52 months). Four of the seven cases underwent follow-up imaging using echocardiography or ultrasound. Conclusion: The incidence of congenital systemic arteriovenous fistulas is low and accounted for 0.009% of pediatric outpatients seen over 10 years at our institute. The spectrum of clinical presentation varies from an innocuous swelling or a pulsating mass to frank heart failure. Strong clinical suspicion and advanced imaging modalities have helped identify some hitherto undescribed connections. Large malformations with multiple communications may persist or recur despite transcatheter closure.

Fetal Tricuspid Valve Agenesis/Atresia: Testing Predictions of the Embryonic Etiology.

Tricuspid valve agenesis/atresia (TVA) is a congenital cardiac malformation where the tricuspid valve is not formed. It is hypothesized that TVA results from a failure of the normal rightward expansion of the atrioventricular canal (AVC). We tested predictions of this hypothesis by morphometric analyses of the AVC in fetal hearts. We used high-resolution MRI and ultrasonography on a post-mortem fetal heart with TVA and with tricuspid valve stenosis (TVS) to validate the position of measurement landmarks that were to be applied to clinical echocardiograms. This revealed a much deeper right atrioventricular sulcus in TVA than in TVS. Subsequently, serial echocardiograms of in utero fetuses between 12 and 38 weeks of gestation were included (n = 23 TVA, n = 16 TVS, and n = 74 controls) to establish changes in AVC width and ventricular dimensions over time. Ventricular length and width and estimated fetal weight all increased significantly with age, irrespective of diagnosis. Heart rate did not differ between groups. However, in the second trimester, in TVA, the ratio of AVC to ventricular width was significantly lower compared to TVS and controls. This finding supports the hypothesis that TVA is due to a failed rightward expansion of the AVC. Notably, we found in the third trimester that the AVC to ventricular width normalized in TVA fetuses as their mitral valve area was greater than in controls. Hence, TVA associates with a quantifiable under-development of the AVC. This under-development is obscured in the third trimester, likely because of adaptational growth that allows for increased stroke volume of the left ventricle.

Virtual and augmented reality: New tools for visualizing, analyzing, and communicating complex morphology.

Virtual and augmented reality (VR/AR) are new technologies with the power to revolutionize the study of morphology. Modern imaging approaches such as computed tomography, laser scanning, and photogrammetry have opened up a new digital world, enabling researchers to share and analyze morphological data electronically and in great detail. Because this digital data exists on a computer screen, however, it can remain difficult to understand and unintuitive to interact with. VR/AR technologies bridge the analog-to-digital divide by presenting 3D data to users in a very similar way to how they would interact with actual anatomy, while also providing a more immersive experience and greater possibilities for exploration. This manuscript describes VR/AR hardware, software, and techniques, and is designed to give practicing morphologists and educators a primer on using these technologies in their research, pedagogy, and communication to a wide variety of audiences. We also include a series of case studies from the presentations and workshop given at the 2019 International Congress of Vertebrate Morphology, and suggest best practices for the use of VR/AR in comparative morphology.

Quantified growth of the human embryonic heart.

The size and growth patterns of the components of the human embryonic heart have remained largely undefined. To provide these data, three-dimensional heart models were generated from immunohistochemically stained sections of ten human embryonic hearts ranging from Carnegie stage 10 to 23. Fifty-eight key structures were annotated and volumetrically assessed. Sizes of the septal foramina and atrioventricular canal opening were also measured. The heart grows exponentially throughout embryonic development. There was consistently less left than right atrial myocardium, and less right than left ventricular myocardium. We observed a later onset of trabeculation in the left atrium compared to the right. Morphometry showed that the rightward expansion of the atrioventricular canal starts in week 5. The septal foramina are less than 0.1 mm2 and are, therefore, much smaller than postnatal septal defects. This chronological, graphical atlas of the growth patterns of cardiac components in the human embryo provides quantified references for normal heart development. Thereby, this atlas may support early detection of cardiac malformations in the foetus.This article has an associated First Person interview with the first author of the paper.

An Appreciation of Anatomy in the Molecular World.

Robert H. Anderson is one of the most important and accomplished cardiac anatomists of the last decades, having made major contributions to our understanding of the anatomy of normal hearts and the pathologies of acquired and congenital heart diseases. While cardiac anatomy as a research discipline has become largely subservient to molecular biology, anatomists like Professor Anderson demonstrate anatomy has much to offer. Here, we provide cases of early anatomical insights on the heart that were rediscovered, and expanded on, by molecular techniques: migration of neural crest cells to the heart was deduced from histological observations (1908) and independently shown again with experimental interventions; pharyngeal mesoderm is added to the embryonic heart (1973) in what is now defined as the molecularly distinguishable second heart field; chambers develop from the heart tube as regional pouches in what is now considered the ballooning model by the molecular identification of regional differentiation and proliferation. The anatomical discovery of the conduction system by Purkinje, His, Tawara, Keith, and Flack is a special case because the main findings were never neglected in later molecular studies. Professor Anderson has successfully demonstrated that sound knowledge of anatomy is indispensable for proper understanding of cardiac development.

Identification of the building blocks of ventricular septation in monitor lizards (Varanidae).

Among lizards, only monitor lizards (Varanidae) have a functionally divided cardiac ventricle. The division results from the combined function of three partial septa, which may be homologous to the ventricular septum of mammals and archosaurs. We show in developing monitors that two septa, the 'muscular ridge' and 'bulbuslamelle', express the evolutionarily conserved transcription factors Tbx5, Irx1 and Irx2, orthologues of which mark the mammalian ventricular septum. Compaction of embryonic trabeculae contributes to the formation of these septa. The septa are positioned, however, to the right of the atrioventricular junction and they do not participate in the separation of incoming atrial blood streams. That separation is accomplished by the 'vertical septum', which expresses Tbx3 and Tbx5 and orchestrates the formation of the electrical conduction axis embedded in the ventricular septum. These expression patterns are more pronounced in monitors than in other lizards, and are associated with a deep electrical activation near the vertical septum, in contrast to the primitive base-to-apex activation of other lizards. We conclude that evolutionarily conserved transcriptional programmes may underlie the formation of the ventricular septa of monitors.

Sinus venosus incorporation: contentious issues and operational criteria for developmental and evolutionary studies.

The sinus venosus is a cardiac chamber upstream of the right atrium that harbours the dominant cardiac pacemaker. During human heart development, the sinus venosus becomes incorporated into the right atrium. However, from the literature it is not possible to deduce the characteristics and importance of this process of incorporation, due to inconsistent terminology and definitions in the description of multiple lines of evidence. We reviewed the literature regarding the incorporation of the sinus venosus and included novel electrophysiological data. Most mammals that have an incorporated sinus venosus show a loss of a functional valve guard of the superior caval vein together with a loss of the electrical sinuatrial delay between the sinus venosus and the right atrium. However, these processes are not necessarily intertwined and in a few species only the sinuatrial delay may be lost. Sinus venosus incorporation can be characterised as the loss of the sinuatrial delay of which the anatomical and molecular underpinnings are not yet understood.

Evolution and Development of the Atrial Septum.

The complete division of the atrial cavity by a septum, resulting in a left and right atrium, is found in many amphibians and all amniotes (reptiles, birds, and mammals). Surprisingly, it is only in eutherian, or placental, mammals that full atrial septation necessitates addition from a second septum. The high incidence of incomplete closure of the atrial septum in human, so-called probe patency, suggests this manner of closure is inefficient. We review the evolution and development of the atrial septum to understand the peculiar means of forming the atrial septum in eutherian mammals. The most primitive atrial septum is found in lungfishes and comprises a myocardial component with a mesenchymal cap on its leading edge, reminiscent to the primary atrial septum of embryonic mammals before closure of the primary foramen. In reptiles, birds, and mammals, the primary foramen is closed by the mesenchymal tissues of the atrioventricular cushions, the dorsal mesenchymal protrusion, and the mesenchymal cap. These tissues are also found in lungfishes. The closure of the primary foramen is preceded by the development of secondary perforations in the septal myocardium. In all amniotes, with the exception of eutherian mammals, the secondary perforations do not coalesce to a secondary foramen. Instead, the secondary perforations persist and are sealed by myocardial and endocardial growth after birth or hatching. We suggest that the error-prone secondary foramen allows large volumes of oxygen-rich blood to reach the cardiac left side, needed to sustain the growth of the extraordinary large offspring that characterizes eutherian mammals. Anat Rec, 302:32-48, 2019. © 2018 The Authors. The Anatomical Record published by Wiley Periodicals, Inc. on behalf of American Association of Anatomists.

Morpho-functional characterization of the systemic venous pole of the reptile heart.

Mammals evolved from reptile-like ancestors, and while the mammalian heart is driven by a distinct sinus node, a sinus node is not apparent in reptiles. We characterized the myocardial systemic venous pole, the sinus venosus, in reptiles to identify the dominant pacemaker and to assess whether the sinus venosus remodels and adopts an atrium-like phenotype as observed in mammals. Anolis lizards had an extensive sinus venosus of myocardium expressing Tbx18. A small sub-population of cells encircling the sinuatrial junction expressed Isl1, Bmp2, Tbx3, and Hcn4, homologues of genes marking the mammalian sinus node. Electrical mapping showed that hearts of Anolis lizards and Python snakes were driven from the sinuatrial junction. The electrical impulse was delayed between the sinus venosus and the right atrium, allowing the sinus venosus to contract and aid right atrial filling. In proximity of the systemic veins, the Anolis sinus venosus expressed markers of the atrial phenotype Nkx2-5 and Gja5. In conclusion, the reptile heart is driven by a pacemaker region with an expression signature similar to that of the immature sinus node of mammals. Unlike mammals, reptiles maintain a sinuatrial delay of the impulse, allowing the partly atrialized sinus venosus to function as a chamber.

Excessive trabeculations in noncompaction do not have the embryonic identity.

BACKGROUND: Ventricular noncompaction is characterized by excessive trabeculations and is associated with heart failure. The lesion is hypothesized to result from failed compaction and thus retention of embryonic trabeculations. Here, we assess for the first time the identity of trabeculations in noncompaction to test whether noncompacted hearts show retention of embryonic trabeculations. METHODS: Using immunohistochemistry, we analyzed cardiac sections of the heart of a control embryo, 3 cases of fetal noncompaction (a set of twins and an unrelated fetus) and 3 fetal hearts without noncompaction. RESULTS: In the embryo, the ventricular trabeculations strongly expressed ANF/NPPA whereas the compact wall did not. In the noncompaction hearts, trabeculations constituted an excessively thick layer. In noncompaction and control fetal hearts alike, however, only a miniscule subset of sub-endocardial myocardium of the trabeculations most proximal to the central ventricular lumen exhibited strong expression of ANF/NPPA, representing Purkinje myocardium. The trabeculations of both fetal control and noncompaction hearts were ANF-negative and orders of magnitude wider than those of the embryo. Both the compact and noncompaction trabeculated myocardium were rich in coronary vasculature. Like embryonic trabeculations, the ANF+ Purkinje myocardium had little if any vasculature. CONCLUSION: The excessive trabeculations in noncompaction do not have the embryonic identity and noncompaction is probably not the result of failed compaction. We propose the lesion results from the compact wall growing into the ventricular lumen in a trabecular fashion.

An interactive three-dimensional digital atlas and quantitative database of human development.

Current knowledge about human development is based on the description of a limited number of embryonic specimens published in original articles and textbooks, often more than 100 years ago. It is exceedingly difficult to verify this knowledge, given the restricted availability of human embryos. We created a three-dimensional digital atlas and database spanning the first 2 months of human development, based on analysis of nearly 15,000 histological sections of the renowned Carnegie Collection of human embryonic specimens. We identified and labeled up to 150 organs and structures per specimen and made three-dimensional models to quantify growth, establish changes in the position of organs, and clarify current ambiguities. The atlas provides an educational and reference resource for studies on early human development, growth, and congenital malformations.

The hypertrabeculated (noncompacted) left ventricle is different from the ventricle of embryos and ectothermic vertebrates.

Ventricular hypertrabeculation (noncompaction) is a poorly characterized condition associated with heart failure. The condition is widely assumed to be the retention of the trabeculated ventricular design of the embryo and ectothermic (cold-blooded) vertebrates. This assumption appears simplistic and counterfactual. Here, we measured a set of anatomical parameters in hypertrabeculation in man and in the ventricles of embryos and animals. We compared humans with left ventricular hypertrabeculation (N=21) with humans with structurally normal left ventricles (N=54). We measured ejection fraction and ventricular trabeculation using cardiovascular MRI. Ventricular trabeculation was further measured in series of embryonic human and 9 animal species, and in hearts of 15 adult animal species using MRI, CT, or histology. In human, hypertrabeculated left ventricles were significantly different from structurally normal left ventricles by all structural measures and ejection fraction. They were far less trabeculated than human embryonic hearts (15-40% trabeculated volume versus 55-80%). Early in development all vertebrate embryos acquired a ventricle with approximately 80% trabeculations, but only ectotherms retained the 80% trabeculation throughout development. Endothermic (warm-blooded) animals including human slowly matured in fetal and postnatal stages towards ventricles with little trabeculations, generally less than 30%. Further, the trabeculations of all embryos and adult ectotherms were very thin, less than 50 µm wide, whereas the trabeculations in adult endotherms and in the setting of hypertrabeculation were wider by orders of magnitude. It is concluded in contrast to a prevailing assumption, the hypertrabeculated left ventricle is not like the ventricle of the embryo or of adult ectotherms. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel.

Development of the human heart.

Molecular and genetic studies around the turn of this century have revolutionized the field of cardiac development. We now know that the primary heart tube, as seen in the early embryo contains little more than the precursors for the left ventricle, whereas the precursor cells for the remainder of the cardiac components are continuously added, to both the venous and arterial pole of the heart tube, from a single center of growth outside the heart. While the primary heart tube is growing by addition of cells, it does not show significant cell proliferation, until chamber differentiation and expansion starts locally in the tube, by which the chambers balloon from the primary heart tube. The transcriptional repressors Tbx2 and Tbx3 locally repress the chamber-specific program of gene expression, by which these regions are allowed to differentiate into the distinct components of the conduction system. Molecular genetic lineage analyses have been extremely valuable to assess the distinct developmental origin of the various component parts of the heart, which currently can be unambiguously identified by their unique molecular phenotype. Despite the enormous advances in our knowledge on cardiac development, even the most common congenital cardiac malformations are only poorly understood. The challenge of the newly developed molecular genetic techniques is to unveil the basic gene regulatory networks underlying cardiac morphogenesis.

Development of the human aortic arch system captured in an interactive three-dimensional reference model.

Variations and mutations in the human genome, such as 22q11.2 microdeletion, can increase the risk for congenital defects, including aortic arch malformations. Animal models are increasingly expanding our molecular and genetic insights into aortic arch development. However, in order to justify animal-to-human extrapolations, a human morphological, and molecular reference model would be of great value, but is currently lacking. Here, we present interactive three-dimensional reconstructions of the developing human aortic arch system, supplemented with the protein distribution of developmental markers for patterning and growth, including T-box transcription factor TBX1, a major candidate for the phenotypes found in patients with the 22q11.2 microdeletion. These reconstructions and expression data facilitate unbiased interpretations, and reveal previously unappreciated aspects of human aortic arch development. Based on our reconstructions and on reported congenital anomalies of the pulmonary trunk and tributaries, we postulate that the pulmonary arteries originate from the aortic sac, rather than from the sixth pharyngeal arch arteries. Similar to mouse, TBX1 is expressed in pharyngeal mesenchyme and epithelia. The endothelium of the pharyngeal arch arteries is largely negative for TBX1 and family member TBX2 but expresses neural crest marker AP2α, which gradually decreases with ongoing development of vascular smooth muscle. At early stages, the pharyngeal arch arteries, aortic sac, and the dorsal aortae in particular were largely negative for proliferation marker Ki67, potentially an important parameter during aortic arch system remodeling. Together, our data support current animal-to-human extrapolations and future genetic and molecular analyses using animal models of congenital heart disease. © 2013 Wiley Periodicals, Inc.

Structure and function of the hearts of lizards and snakes.

With approximately 7000 species, snakes and lizards, collectively known as squamates, are by far the most species-rich group of reptiles. It was from reptile-like ancestors that mammals and birds evolved and squamates can be viewed as phylogenetically positioned between them and fishes. Hence, their hearts have been studied for more than a century yielding insights into the group itself and into the independent evolution of the fully divided four-chambered hearts of mammals and birds. Structurally the heart is complex and debates persist on rudimentary issues such as identifying structures critical to understanding ventricle function. In seeking to resolve these controversies we have generated three-dimensional (3D) models in portable digital format (pdf) of the anaconda and anole lizard hearts ('typical' squamate hearts) and the uniquely specialized python heart with comprehensive annotations of structures and cavities. We review the anatomy and physiology of squamate hearts in general and emphasize the unique features of pythonid and varanid lizard hearts that endow them with mammal-like blood pressures. Excluding pythons and varanid lizards it is concluded that the squamate heart has a highly consistent design including a disproportionately large right side (systemic venous) probably due to prevailing pulmonary bypass (intraventricular shunting). Unfortunately, investigations on rudimentary features are sparse. We therefore point out gaps in our knowledge, such as the size and functional importance of the coronary vasculature and of the first cardiac chamber, the sinus venosus, and highlight areas with implications for vertebrate cardiac evolution.