![]()
CHAPTER 1
Development of the Heart and Great Vessels
Peter J. Gruber,1 Andy Wessels,2 and Steven W. Kubalak2
1Primary Childrenās Medical Center, Salt Lake City, UT, USA
2Medical University of South Carolina, Charleston, SC, USA
Introduction
Modern cardiac embryology combines molecular and cellular biologic techniques with traditional embryologic morphologic approaches during development. The limited descriptions of human cardiac development are necessarily supplemented by nonhuman models of cardiac development. Avian embryos have traditionally been favored experimental models because of the ease with which they can be observed and manipulated. More recently, the developing mouse has become the preferred model for studying cardiac development because of the strength of genetic and molecular investigative tools available in this species. Where possible, this chapter discusses how results in experimental animal models relate to human cardiac development. Table 1.1 provides a simplified comparison of two widely utilized developmental schemes for developmental staging in chick and mouse embryos [1ā7]. The comparison of multiple species provides an important platform for understanding the development of the human heart and the pathogenesis of human disease.
Table 1.1 Simplified comparison of developmental stages between human, mouse, and chicken embryos.
Formation of Cardiac Precursors
All of the cells that will become part of the heart derive from populations of undifferentiated precursors that will be influenced by external signals into their final developmental pathways. In addition to the intellectual challenge of understanding how these acts of differentiation occur, intense activity in this field is also driven by the possibility of controlling cardiac tissue differentiation to replace diseased myocardium in the postnatal heart.
Repeated cell divisions of the fertilized egg form a cell mass that evolves into two distinct layers of cells. The epiblast layer is separated from a second layer of cells, called the hypoblast in the chick or the primitive endoderm in the mouse and human. The next critical stage of development is gastrulation where widespread cell migration into and reorganization within the blastocoele cavity result in the formation of three germ layers (ectoderm, mesoderm, and endoderm) and the determination of the future body plan of the embryo (Figure 1.1) [7,8].
Gastrulation of precardiac cells is an early event in all species. In the human, gastrulation takes place at the beginning of the third week of development and angioblasts in the cardiogenic region are present shortly thereafter. At the time that precardiac cells gastrulate in chick embryos (HamburgerāHamilton stage 3), the primitive streak is less than 1 mm in length; the portion of the streak through which the precardiac cells ingress extends as a relatively broad swath 0.125ā0.75 mm from the anterior limit of the streak [9]. The most anteriorly gastrulating cells contribute to the most anterior portion of the primitive heart tube.
After cells have undergone gastrulation they enter the undifferentiated mesenchyme. Uncommitted precardiac cells enter the primitive streak only to become specified to their cell type or migratory pathways in the mesoderm after leaving the streak [9]. Subsequently, the precardiac cells will move laterally to join the lateral plate mesoderm at the level of Hensenās node. The lateral plate mesoderm then splits into two layers, a splanchnic layer directly above the endoderm and a somatic layer directly below the ectoderm. The anterior endoderm provides signals to splanchnic mesodermal cells to enter the precardiac lineage. Fibroblast growth factors (FGFs)-1, -2, and -4 and bone morphogenetic protein 2 (BMP-2) are proteins that appear to be critical to this process [10]. However, to date no single gene has been identified whose ablation leads to a specific failure of all myocardial differentiation from precardiac mesoderm. This observation may argue the presence of either a considerable genetic redundancy in precardiac myocyte differentiation or an unsuspected diversity of precardiac myocyte lineages following independent genetic pathways.
Precardiac cells are found in an epithelial sheet at the cranial end of the splanchnic mesoderm and can be identified at this point by a variety of molecular markers such as the transcription factors NKX2-5, MEF2, HAND1, HAND2, GATA4, TBX5, and ISL1 [11ā17]. The region of splanchnic mesoderm expressing precardiac markers is also known as the āheart-forming fieldā and is larger than the region that will actually contribute cells to the heart tube [18]. In rodent embryos, but not chick embryos, precardiac mesodermal cells exhibit spontaneous contractile activity, indicating a relatively advanced state of differentiation towards the cardiac myocyte lineage [19,20].
The precardiac mesodermal cell mass migrates as a single unit rather than as a collection of independent cells. The precardiac mesodermal sheets on each side of the embryo migrate together towards the midline cranial to the anterior intestinal portal. When the most cranial portions of the bilateral precardiac mesoderm masses meet in the midline, the total premyocardial cell population forms a horseshoe-shaped crescent called the first (primary) heart field. The cues that enable and promote movement of these cells are provided by a noncardiac tissue, the endoderm, as demonstrated by experimental removal of the endoderm and/or ectoderm. The extracellular matrix molecule fibronectin may be one of the important components of the endodermal surface to which the precardiac cells are responding [21].
Precursors of the endocardium follow similar migratory pathways as the precardiac cells, but there are important differences. Pre-endocardial cells and pre-endothelial cells are known as angioblasts. The endocardial angioblasts are first detectable in the splanchnic mesoderm. Mesodermal cells are induced to enter the angioblast lineage by signals such as transforming growth factor beta (TGFĪ²) 2ā4 and vascular endothelial growth factor (VEGF) signaling from the endoderm [10]. Endocardial angioblasts migrate anteriorly and to the midline with the premyocardial cell mass, but they do so as individual cells.
Formation of the Tube Heart
As the precardiac cell masses of the first heart field move steadily towards the midline, endocardial cells begin to form a network of tiny channels that will coalesce into a complex endocardial network surrounded by a myocardial mantle [22]. If the mesodermal sheets are prevented from meeting in the midline as a consequence of genetic [15] or mechanical manipulation [23], dual heart tubes will be formed that undergo some degree of further independent development. However, in normal development, the endothelial network quickly transforms into a single endothelial channel within a single myocardial tube (Figure 1.2) [7].
The tube heart at the time of its formation is connected to the foregut along its dorsal surface throughout its length by a structure called the dorsal mesocardium [24]. As looping proceeds, the dorsal mesocardium degenerates until it remains connected only at the atrial and arterial poles of the heart. The disintegration of the central portion of the dorsal mesocardium is a key event for looping to proceed normally, while the arterial and venous attachments provide āanchorsā for the looping heart tube. The mesenchymal portion of the dorsal mesocardium known as the dorsal mesenchymal protrusion [25,26] protrudes into the atrium posteriorly and is a derivative of the second heart field [25]. It is an important contributor to atrioventricular (AV) septation and serves as a conduit for the developing pulmonary veins. The dorsal mesocardium is also a pathway for cellular migrations as development proceeds, including neural crest-derived neural structures [27] and possibly cells derived from the ventral neural tube [28].
Soon after the formation of the cardiac tube the heartbeat is initiated and blood circulation can be observed (embryonic day 8.5 in the mouse and day 20 in the human). With the initiation of circulation the heart becomes the first organ to adopt its essential mature function in the embryo. At this point in cardiac organogenesis, however, the tube heart has not yet obtained its full complement of cell populations necessary for complete cardiac development. Early fate mapping studies [29ā31] showed that the primary heart tube is derived from two bilateral fields of precardiac mesoderm, currently called the first heart field; this precardiac mesoderm was long considered the precursor tissue of the heart. Studies by others [32], however, strongly suggested that growth of the heart tube, specifically at the arterial pole, depended on the addition of cardiac tissue from a secondary pool of progenitor cells. It was not until the early twenty-first century that the nature of this additional cell population was elucidated. The combined studies of various laboratories [25,33ā36] have provided significant new insights into the importance of this additional population of cells, called the second heart field, in the elongation and growth of the heart tube and in the formation of a mature four-chambered heart (Figure 1.3) [7]. Thus, these studies have demonstrated that the secondary heart field contributes at the arterial pole to the outflow tract and right ventricle and at the venous pole to parts of the atria and the dorsal mesenchymal protrusion.
The Tube Heart, Segments, and Segmental Identity
Traditionally, the heart tube has been regarded as containing the precursors of all of the cardiac segments. In reality, at the time the heartbeat is initiated the heart tube primarily consists of future left ventricular tissues [37,38]. Immunohistochemical, in situ hybridization, and cell fate tracing techniques have demonstrated that the outflow tract, the right ventricle [32], the AV junction segment [37], the atria [39], and the sinus venosus are added to the heart as looping proceeds. Indeed, these are the structu...
