There is substantial nutritional and oxygen need for the embryo proportional to its astounding developing pace. Thus, it is not surprising that heart is the first organ begins to function.1 This process works fast enough to give rise a functioning heart by the fourth week of development. Vascular system development co-occurs with heart formation to take on the distribution of these necessities. At the third week of embryogenesis, specialized cell lineages that are destined to become heart and outflow tracts organize to form two angioblastic cords.2 Each cord is continuous with dorsal aorta cranially, and with vittelo-umbilical vein caudally.3 These cords become primitive tubes with tubular formation. Primitive tubes, the first evidence of a developing heart at the level of tissue organization, fuse along and create a single tubular heart primordium. Hyaluronate rich and cell free extracellular matrix, sometimes named “cardiac jelly,” fills tubular heart. This primordial tubular structure continues to elongate and enlarge. At certain time, it bends over itself and brings the arterial (cranial) and venous (caudal) poles close to each other. This process is called looping, which is, in part, caused by unequal evolvement speed of the two tubular ends. Also, inward bending of splanchnic mesoderm helps this process.4,5 Several genes and pathways have been identified by means of animal models and gene knock-out research that play crucial roles in tubular fusion and looping processes.
Initially, the heart is attached to the dorsal part of the developing embryo. However, this mesenteric attachment disappears soon and heart becomes suspended between its two ends: arterial outflow end is anchored by pharyngeal arches and venous pole is by the septum transversum.
PARTITIONING: ATRIOVENTRICULAR CANAL AND ATRIA
Inside the developing tubular heart, at the dorsal and ventral aspects of the future atrioventricular (AV) canal, tissue accumulations are being noticed. These aggregations are called as endocardial cushions. By continuous 2proliferation from dorsal and ventral walls, eventually they merge into each other in the middle of the canal. In doing so, AV canal is separated into left and right portions. Endocardial cushions further transform into valves and into membranous segment of the septum by proliferation of specialized cell lineages under the complex process of transcription factor signaling.
At the beginning, atrium is a large unseptated cavity. This common primordial atrium is separated by two different septa into right and left atria in successive and overlapping processes. These septa are called septum primum and septum secundum, named in the order of their appearance. Septum primum, a membranous septum, partially divides the common primordial atrium into right and left chambers. The incomplete division at the endocardial cushion region creates an opening called as foramen primum. While foramen primum gradually closes down, small holes are formed as a consequence of regional apoptosis upon completion of septal formation. These holes merge to become foramen secundum. Soon after the completion of septum primum, septum secundum begins to develop. At this stage, septum primum begins to disappear. Septum secundum contains more muscle fiber. It grows very close to the septum primum, in a position that almost attached to it. The nonoverlapped portion of both septa (between the disappearing primum and newly developed secundum) is left open and called as foramen ovale. Through these foramens, blood with higher oxygen saturation bypasses fetal lungs and flows from right to left atrium. Caudal remnant of septum primum serves as a valve and it can be appreciated during fetal echocardiography while swinging into left atrium. Significant changes in cardiovascular dynamics occur at the time of birth that cause foramen ovale to close in approximately 96 hours.
During atrial partitioning, transformation and incorporation of sinus venosus into atrium occurs. Auricles are formed as ridged muscular pockets in each atrium. Later, endocardial cells transform into mesenchymal cells to invade extracellular matrix. Then endocardial cushions contribute to the formation of valves and membranous part of interventricular septum (IVS).
VENTRICULAR PARTITIONING
Ventricular septal formation occurs at the seventh week of embryogenesis. However, heart begins to contract prior to this point. Adult IVS has muscular and membranous segments. Muscular segment begins to appear as a ridge in ventricular floor. As discussed previously, it grows toward endocardial cushions and together they divide common ventricle into right and left ventricles. At very early stages of the first trimester, a small opening in IVS near the endocardial cushions lets interventricular blood passage until the seventh week of gestation. Membranous part of the IVS that originates from endocardial cushions merges with expanding muscular IVS and this small opening is closed. 3Prior to this final process, ventricular separation is completed. Importantly, this segment of IVS shows continuation with the conotruncal septum that lies between aorta and main pulmonary artery. In clinical practice, visualization of membranous IVS may sometimes be insufficient secondary to echolucent artifacts during ultrasound evaluation of the fetal heart. Hence, the suspicion of a ventricular septal defect during fetal echocardiography in four chamber view can be ruled out by visualization of normal left ventricular outflow tract.
SINUS VENOSUS
Venous entrance of primordial atria is noticed as a bulbous enlargement of the looped primordial heart and it is called “sinus venosus.” Its opening soon takes part in the right atrium after partitioning of common atrial primordium. Sinus venosus collects venous drainage from bilateral vitellin, umbilical and cardinal veins. As embryo develops and venous return increases, two bulges appear in sinus venosus and they are called right and left horns of sinus venosus. With changing blood flow dynamics along with complex transcriptional regulation, the right horn develops into superior vena cava (SVC) and the left one into coronary sinus.3 Other than their contribution to systemic venous return, sinus horns also involve in sinoatrial node development.6
DEVELOPMENT OF OUTFLOW TRACT
Bulbus Cordis and Truncus Arteriosus
As sinus venosus plays significant role in atrial partitioning at the venous pole, bulbus cordis has similar anatomic importance for outflow tract development at the arterial pole. Aortic sac, which gives rise to the aortic arches superiorly, connects to the bulbus cordis of developing tubular heart. This bulbar structure is continued with truncus arteriosus (common trunk), which is a primordial origin of aorta and main pulmonary artery.
The septum that separates truncus arteriosus into aorta and main pulmonary artery (conotruncal septum) is originated from mesenchymal tissues in bulbus cordis. Two regional proliferations begin to appear and they are called as ridges (bulbar and truncal or conotruncal ridges). With continuous proliferation and extension, they meet in the middle to create two lumens. Cardiac neural crest cells also involve in this process along with other regional cell lineages. The term of “conotruncal defects” refers to the developmental abnormalities of conus (the segment of bulbus arteriosus close to the developing heart) and truncus (the farther segment of bulbus cordis). An inverted rotation of conotruncus would lead to the transposition of great arteries, one of the common cyanotic congenital heart defects.7
In the proximal part of outflow tract, endocardial cushions meet in the middle and fuse to separate the common trunk. Conversely, distal outflow tract separation is achieved by the migration of cardiac neural crest cell 4population.8 During internal and longitudinal separation of common trunk into aorta and pulmonary trunk, the whole outflow structure twists around itself. This spiraling process eventually gives itself a unique orientation that can be appreciated during fetal echocardiography, in three-vessel view. As embryonic development continues, bulbus cordis is eventually incorporated in ventricles.
DEVELOPMENT OF CARDIAC VALVES
Aortic and pulmonary semilunar valves are the first set of valves to form. They develop by the protrusion of subendocardial tissues located in respective regions. This process begins after the complete longitudinal separation of truncus arteriosus. Distal outflow tract and intercalated cushions set the stage for ventriculoarterial valve development. Knock-out animal model studies have shown that Tbx1 gene is crucial for the fusion of these outflow cushions and for the formation of separate valves.9 Aortic and pulmonary valves each consist of three leaflets.
Initially endocardial cushions serve as primitive valves for blood to flow in one direction.10 Soon, mitral and tricuspid valves develop from regional tissue swellings at endocardial cushions. Mitral and tricuspid valves have two and three cusps, hence the name bicuspid and tricuspid valves, respectively. These leaflets are derived from endothelial layer of primordial heart tube where specific cell population undergoes mesenchymal transformation. Studies have shown that epithelial layer also contribute to AV valvar development.9 Papillary muscles are thought to be originated by further condensation of local trabeculations. Papillary muscles attach to the cusps of AV valves via the chordae tendineae.
DEVELOPMENT OF CARDIAC CONDUCTION SYSTEM
Since all cardiac myocytes of the developing heart have pacemaker abilities initially, contractions begin in tubular primordium even before looping process sets up. Usually, a regular rhythm is established during the fifth week of development with ventricular contractions following atrial contractions.11,12 However, earlier to this point, common primordial atrium originated peristalsis helps blood to move forward at around 23rd day of embryogenesis.2,3 Shortly, sinus venosus takes over this task until specialized cell lineages of noncontractile myocytes accrete and begin to function as sinoatrial node in the right atrial primordium. This specific cell group connects with separately developing AV node to create a bundle. AV bundle fibers grow and divide into two ventricular branches just after their entrance to ventricles. Sinoatrial node is located in the right atrium and AV node is in close proximity to endocardial cushions, at the right side of the interatrial septum. Even finer and wider distribution 5throughout the ventricles is achieved by penetrating His bundle and Purkinje fibers that are in part originated from cardiogenic mesoderm and epicardial surface.3–5
DEVELOPMENT OF FETAL VASCULAR SYSTEM
The venous return to future atria occurs from three sources: common cardinal, vitellin and umbilical veins. Vitelline veins regress after embryonic period.11 Cardinal veins collect blood from developing body of the embryo. They are formed inferiorly and superiorly in bilateral fashion. When they join each other, they form right and left common cardinal veins. Part of the left common cardinal vein persists and forms coronary sinus. It receives drainage from most epicardial ventricular veins and empties directly into the right atrium. However, coronary vasculature is one of the last developing structures of embryonic heart.13
The brachiocephalic vein is formed by the anastomosis of right and left superior cardinal veins. SVC and internal jugular veins are formed by superior cardinal veins also. Inferior vena cava (IVC), common iliac veins, renal, suprarenal, gonadal, azygos and hemiazygos veins develop from the respective segments of subcardinal and cardinal veins during their sequential appearance and disappearance. Dysgenesis or abnormalities in these sequential events may lead to vascular developmental anomalies such as persistent SVC, segmental atresia of IVC and double IVC or double SVC.
As the third source of venous return to primordial atrium, umbilical veins provide highly oxygenated blood for developing embryo. Umbilical vein and the IVC are connected via ductus venosus. With adequate pressure, a great volume of blood with high nutrient and oxygen content bypasses the liver and reaches to IVC. A sphincter mechanism in ductus venosus helps to regulate the amount of blood flow directed to IVC and fetal liver. Also, IVC collects blood from caudal part of the embryo. One of the paired umbilical veins disappears in the early first trimester.
The development of aortic root and the ascending aorta has been discussed in the outflow partitioning topic. Another bypass mechanism, ductus arteriosus connects pulmonary trunk to aorta, redirects the majority of right ventricular output into systemic circulation. Aortic arches stem from aortic sac and they supply pharyngeal arches. Body of aorta develops from fused pair of two dorsal vessel primordia. It successively gives many branches along the body of developing embryo such as intercostal, vertebral, lumbar and common iliac arteries. Internal iliac arteries arise from aorta via umbilical arteries. Even yolk sac and chorion are supplied by aorta.
In the embryonic-fetal life, a small percentage of blood flow is directed toward lungs. Accordingly, there is a small amount of venous return from lungs to primordial left atrium carried by pulmonary veins. The origin of the pulmonary vein is incorporated into dorsal atrial wall during atrial 6partitioning; pulmonary vein primordium has been showed to contain traces of atrial myoblasts. Soon, four separate pulmonary veins develop in the mediastinal mesenchyme.
CARDIAC MORPHOGENESIS
From early progenitor cells to its final three-dimensional form, regulation of cardiac development requires a complex regulatory process involving multiple signaling pathways and many transcription factors. First cells that leave the primitive streak to become heart are splanchnopleuric mesodermal cells. Surrounding local endodermal signaling systems, especially bone morphogenic protein (BMP) and WNT lead the synthesis of important transcription factors. TBX5, GATA4, BAF60c, along with other transcription factors stimulate myocardial progenitor cells to coalesce into a region called “primary heart field.”14,15 They further organize into endocardial-angioblastic strands. These bilateral tubular strands soon merge each other and form single tubular heart primordia as discussed previously. From this stage, extracardiac pharyngeal mesodermal progenitor cells (the secondary heart field) involve in and boost cardiac development along with proliferating cardiac progenitor cells. Elongation and looping are achieved by active proliferation and differentiation of early myocardial progenitor cells at the two poles of tubular heart primordium.14 Then, by means of retinoic acid effect, a specific cell population in the posterior primary heart field begins to proliferate and transform for atrial development. The lack of retinoic acid exposure in the anterior primary heart field leads to ventricular rather than atrial development by default.10 In other words; cells in the primary heart field give rise to the development of left and right atria and the left ventricle. Conversely, the cell population in the secondary heart field develop into right ventricle and outflow tract.7
While WNT, fibroblast growth factor (FGF) and Hedgehog signaling pathways are crucial for proliferative elongation and looping, inhibitor signals (such as BMP signaling at the arterial pole) are also important in cardiac morphogenesis by causing regional discordant growth.14
During the looping process, polarity of the developing heart becomes right to left.4,5 A considerable amount of transcription factors play in the field in different moments and interacts with each other for tubular heart formation, looping, elongation, ballooning and condensation processes. As an example, transcription factors Hand1, Hand2 and Pitx2 have been shown to play significant role in looping.4,5,10 Also, certain left–right patterning proteins (Nodal and Lefty-2) have been demonstrated to play a role in polarity formation. However, the underlying mechanism for the rightward looping of primordial tubular heart is poorly understood.14
Ballooning morphogenesis is an attractive model in describing early embryonic cardiac developmental stages. Atrial and ventricular primordia 7along with endocardial cushions develop as sequentially appearing proliferation niduses at respective regions. T-box family of transcription factors plays significant role in the orchestration of this region specific proliferation process.14 Also, Neuroregulin1-ErbB signaling system is believed to be involved in the formation of trabeculae and Purkinje fibers.16
Recent advances have also shed light on coronary vasculature development pathways. Coronary development involves communication between the epicardium, the subepicardial mesenchyme and the myocardium. This communication includes signaling systems such as WNT and Hedgehog, growth factors and signal molecules such as vascular endothelial growth factor, FGF, transforming growth factor-β and erythropoietin.17
The mechanisms behind many congenital cardiovascular malformations are complex. There are modifier genes, dosage sensitive genes and many transcription factors that work in several cardiac morphogenetic pathways. In many instances, it is more complicated than a simple gene mutation. As an example, majority of 22q11.2 microdeletion syndrome cases have conotruncal defects, some carry mutations in a dosage sensitive gene TBX1, others suspected to have modifier molecule abnormalities affecting sonic hedgehog and retinoic acid signaling pathways.18 Also, environmental (epigenetic) factors may contribute to the development of certain cardiovascular malformations, e.g. high glucose levels may affect secondary heart field and neural crest in pregestational diabetes.19,20
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- Nakajima Y. Mechanism responsible for D-transposition of the great arteries: is this part of the spectrum of right isomerism? Congenit Anom (Kyoto). 2016;56(5):196–202.
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- Lai D, Liu X, Forrai A, et al. Neuregulin 1 sustains the gene regulatory network in both trabecular and nontrabecular myocardium. Circ Res. 2010;107(6):715–27.
- Olivey HE, Svensson EC. Epicardial-myocardial signaling directing coronary vasculogenesis. Circ Res. 2010;106(5):818–32.
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- Roest PA, van Iperen L, Vis S, et al. Exposure of neural crest cells to elevated glucose leads to congenital heart defects, an effect that can be prevented by N-acetylcysteine. Birth Defects Res A Clin Mol Teratol. 2007;79(3):231–5.
- Molin DG, Roest PA, Nordstrand H, et al. Disturbed morphogenesis of cardiac outflow tract and increased rate of aortic arch anomalies in the offspring of diabetic rats. Birth Defects Res A Clin Mol Teratol. 2004;70(12):927–38.