The cardiovascular system is the major transport system of the body, conducting oxygen, cells, nutrients, and waste products to their final destinations. The heart is the first organ to develop; severe defects in heart development are one major cause of embryonic lethality. In the human embryo, the heart begins beating at ~21 days post-conception. The heart pumps throughout its morphogenesis, and its constituent cells are exposed to multiple mechanical stimuli, including wall shear stress, transmural pressure, and stretch. The development of the heart proceeds via sequential processes of cardiomyocyte specification (day 15 in the human embryo; HH stages 5–9 in the chicken (Hamburger and Hamilton, 1951
)), formation and looping of the heart tube (days 20–28 in the human; HH stages 10–24 in the chicken), development and growth of the chambers (days 28–32 in the human; HH stages 19–24 in the chicken), and development of cardiac cushions, valves, and septae (days 32-birth in the human; HH stages 25–34 in the chicken)(Bruneau, 2008
; Manner, 2009
The mechanical forces accompanying heart development have been most closely examined in chicken and zebrafish embryos. Cardiac looping morphogenesis in the chicken closely resembles that in the human embryo (Kirby, 2007
). During looping, the initially straight heart tube undergoes bending and torsion to form the basic topographical pattern of the multi-chambered heart (). During the first stage of looping, c-looping, the heart tube bends ventrally and twists toward the right (dextrally) to form a c-shaped tube. During s-looping, the c-shaped tube shortens and twists into an s-shaped tube. Bending can occur normally even in embryonic chicken hearts isolated in vitro (Manning and McLachlan, 1990
), and is thought to result from actomyosin-mediated changes in cell shape (Latacha et al., 2005
). As the heart tube bends, myocardial cells flatten at the outer convex curvature, whereas others elongate and thicken at the inner concave curvature (Manasek et al., 1972
). Inhibiting actin polymerization blocks bending in vivo and in isolated hearts (Latacha et al., 2005
), consistent with a cell shape-driven process.
Morphological changes during looping of the heart tube. The straight heart tube bends and twists dextrally to form a c-shaped tube. The tube then shortens and twists into an s-shaped tube.
Changes in cell shape also appear to be responsible for the ballooning process that sculpts the chambers of the heart. Ballooning involves expansion and bulging of the linear walls of the looped heart tube into bean-shaped chambers, again with a convex outer curvature and a concave inner curvature. High-resolution imaging of the developing zebrafish heart revealed that cells of the outer curvature flatten and elongate relative to those of the inner curvature, which remain cuboidal (Auman et al., 2007
). Physical forces resulting from normal blood flow were shown to be required for cell shape change, as disrupting flow either genetically or pharmacologically inhibited the flattening and elongation of cells in the outer curvature (Auman et al., 2007
). It is still unclear how the cells are sensing these forces and transducing them into alterations in cell shape.
Forces from blood flow also regulate development of the heart valves. After looping is completed in the zebrafish embryo, cardiac cushions form at the atrioventricular (AV) boundary, the separation between the ventricle and atrium chambers (Eisenberg and Markwald, 1995
). Communication between the myocardial and endocardial layers at the AV boundary leads to an EMT of the endocardial cells, forming cushions that then differentiate into the flaps of the AV valve. Quantitative in vivo imaging of hemodynamics in the developing zebrafish heart revealed the presence of high-shear (>1 dyn/cm2
) vortical flow during and after valvulogenesis (Hove et al., 2003
). Valves fail to form in mutant embryos that lack a heartbeat (Bartman et al., 2004
), and mechanically blocking blood flow disrupted looping of the heart tube and valve formation (Hove et al., 2003
). Fraser’s group recently showed that disturbed flow patterns, rather than shear stress per se, are responsible for activating the flow-responsive genes that direct valvulogenesis (Vermot et al., 2009
). Specifically, reversing flows activated Kruppel-like factor (Klf)-2a in the endothelium in valve-forming regions, and morpholino-mediated knockdown of Klf2a disrupted heart valve formation (Vermot et al., 2009
). How endocardial cells sense and respond to disturbed flow is unclear, but extensive analysis of hemodynamic signaling in the endothelium of adult vessels yields several promising clues. Oscillatory flow patterns with recirculation tend to form at branch points within the vascular bed and distal to stenoses, and these hemodynamic forces have been implicated in the local development of atherosclerosis (White and Frangos, 2007
). Endothelial cells in regions of disturbed flow in vivo and in culture exhibit increased proliferation, apoptosis, junctional permeability, and oxidative stress (Hahn and Schwartz, 2008
) resulting from sustained production of reactive oxygen species (ROS), which themselves act as mediators of EMT (Radisky et al., 2005
). It remains to be determined whether and how disturbed flows signal to endocardium through ROS.
Formation of the heart valves proceeds similarly in amniotes. In the chicken embryo, the AV and outflow tract cushions start to emerge as expansions of cardiac jelly, the ECM in between the myocardium and endocardium that is rich in hyaluronan and chondroitin sulfate proteoglycans. The myocardium then induces an EMT of the endocardium, causing the latter cell population to migrate into the cushions and form the mitral and tricuspid valves as well as the atrial and ventricular septae (Martinsen, 2005
). The Rho GTPase effector, Rho kinase (ROCK), is expressed in the migrating population and treatment with the ROCK inhibitor Y27632 blocks valve formation in the chicken embryonic heart (Sakabe et al., 2006
). Furthermore, the basic helix-loop-helix (bHLH) transcription factor Twist1 is expressed in the mesenchyme of the developing heart cushions (Ma et al., 2005
), and induces proliferation and migration of endocardial cushion cells (Shelton and Yutzey, 2008
). Mechanical stress activates ROCK (Chiquet et al., 2009
; Wozniak and Chen, 2008
) and induces expression of Twist in Drosophila embryos (Desprat et al., 2008a
; Farge, 2003
), suggesting that hemodynamic forces may induce EMT and heart valve formation through Rho GTPases and Twist.
Congenital heart defects affect approximately one percent of newborn infants (Hoffman, 1995a
), making them the most common type of birth defect and a major cause of infant mortality in the first year of life. Abnormalities in the early stages of heart development result in embryonic lethality (Hoffman, 1995a
), so the majority of heart defects in infants that make it to term result from abnormal development of the valves and septae (Hoffman, 1995b
). Members of the Notch family have emerged as key regulators of valve development, and disruptions in Notch or Jagged signaling can lead to bicuspid aortic valve (BAV) (Garg et al., 2005
), a defect in which the aortic valve has two leaflets instead of three. Notch is thought to act by promoting EMT in the cushions during valve formation (Timmerman et al., 2004
). It will be interesting to determine the links, if any, between mechanical stress, Notch, and EMT in cardiac morphogenesis.