The above biomechanical studies and in vivo examples strongly suggest that heart function affects development of form. Possible mechanisms for this affect include (1) mechanical motion of the heart directly pushes cells and tissues into their locations, and/or (2) mechanical function affects signaling pathways and, therefore, the cellular program of cells and their developmental fate. The later could occur through at least two different mechanisms.
First, shear stress on endocardial cells might induce local signaling changes, as has been demonstrated with endothelial cells in both in vitro and in vivo studies (e.g., Icardo, 1989; Helmlinger et al., 1991, 1995). Some of the molecules that may sense this shear stress include the integrins (Ingber, 1999), platelet endothelial cell adhesion molecule-1 (PECAM-1; Fujiwara et al., 2001), VE-cadherin (Shay-Salit et al., 2002), ion channels (Barakat, 1999), vascular endothelial growth factor (VEGF) receptor 2 (Shay-Salit et al., 2002), and G proteins (Gudi et al., 1996). Several studies, including some using microarray technology, have identified some of those genes downstream of the stress signals (Resnick et al., 1997; Chien et al., 1998; Malek et al., 1999; Gimbrone et al., 2000; McCormick et al., 2003; Black, 2000), and it is possible that some of these genes may affect cardiac morphogenesis. Development of the cushions and valves seems particularly sensitive to changes in heart function, and molecules known to be expressed in the endocardium and responsible for endocardial cushion/valve morphogenesis include ES/130 (Ramsdell et al., 1998), transforming growth factor (TGF) -β3 (Nakajima et al., 1994, 1998; Brown et al., 1996; Ramsdell and Markwald, 1997; Boyer et al., 1999), VEGF (Dor et al., 2003), nuclear factor of activated T cell (NFAT) c1 (de la Pompa et al., 1998; Ranger et al., 1998), Notch and hesr2 (Kokubo et al., 2004), Wnt/β-catenin (Hurlstone et al., 2003; Liebner, 2004), bone morphogenetic protein/TGF-β (Nakajima et al., 1998), ErbB, and NF1/Ras (all nicely reviewed in Armstrong and Bischoff, 2004). It remains to be seen which, if any, of these endothelial/endocardial signaling pathways are perturbed in instances of poor heart function.
Second, changes in myocardial function may affect gene expression within that layer, and these molecular changes may affect both myocardial and endocardial cell development. An abundant literature describes muscle genes whose expression is affected by muscle function (reviewed in Seidman and Seidman, 2001), although this literature focuses on the remodeling events that take place in adult hearts. However, some of the genes expressed in these altered conditions are those used for heart morphogenesis in the embryo. For example, eHand and dHand, two prominent players in chamber formation, experience dynamic changes in their level of expression in cardiomyopathies (Ritter et al., 1999; Natarajan et al., 2001). Other molecules that regulate morphogenesis and demonstrate changes in expression or posttranslational regulation in response to mechanical forces include β-catenin (Masuelli et al., 2003) and the calcineurin/NFAT pathway (Molkentin et al., 1998). Because the calcineurin/NFATc pathway is affected by changes in heart function in adult animals, is also involved in endocardial cushion and valve morphogenesis (Uhing et al., 1993; de la Pompa et al., 1998; Ranger et al., 1998; Graef et al., 2001; Chang et al., 2004), and because changes in heart function affects development of the valves (Bartman et al., 2004), this pathway becomes an extremely attractive candidate for linking embryonic heart function with cushion and valve formation. Studies that have used cyclosporin A (CsA) to suppress NFAT signaling have demonstrated similar cushion/valve defects to the NFATc1 null mice (Uhing et al., 1993; Graef et al., 2001). However, cyclosporin treatment is known to affect contractile activity (Abbott et al., 1998; Janssen et al., 2000), which may indicate that the defects seen with CsA treatment of mouse embryos may be secondary to changes in heart function. In chick embryos treated with CsA, defects were seen in ventricular wall morphology, heart looping, and formation of the endocardial cushions (Liberatore and Yutzey, 2004). Presumably, heart function was poor in these embryos due to the changes in ventricular wall morphology, but this was not clearly demonstrated.
One recent study that attempted to tease apart the issues of NFAT's role in function and form in utero ultrasound biomicroscopy to study NFATc1−/− embryos and control littermates sequentially from E10.5 to E14.5 (Phoon et al., 2004). These authors showed that null mice developed defects in diastolic dysfunction and valvular regurgitation, despite maintaining normal contractile (systolic) function. Further studies such as these will be required to prove or disprove the relationship between NFAT's role in heart function and its role in heart morphogenesis.