A universal and early step in vertebrate cardiogenesis is the formation of a linear heart tube, from which the mature chambered organ is derived. Whilst considerable progress has been made in understanding subsequent steps in chamber formation and the contribution of cells that initially lie outside the heart tube (Boogerd et al., 2009; Buckingham et al., 2005
), much less is known about the morphogenetic events that regulate tube formation itself.
The contractile, myocardial cells of the heart tube are derived from bilateral, anterior lateral plate (splanchnic) mesodermal domains that converge on the ventral midline. In amphibian embryos, such domains have been termed the heart field, referring to classical embryological studies demonstrating the regulative capacity of such tissue in its contribution to the embryonic heart (reviewed in Mohun et al., 2003
). In Xenopus laevis
, myocardial precursors within the heart field express markers of striated muscle differentiation (stage 27) prior to fusion of the bilateral domains, some 9 h before formation of the heart tube (stage 32) (Latinkic et al., 2004; Mohun et al., 2000
). Furthermore, as the bilateral precursors approach the ventral midline, they overlap with a second and distinct population of cells, the embryonic macrophages. This myeloid domain is also mesodermal in origin and initially encompasses the position on the ventral midline where the heart will ultimately form (Smith et al., 2002
). Whilst macrophages begin to disperse from this site of origin prior to differentiation of cardiac precursors, they remain prevalent in the heart-forming region, filling the trough-shape made as the myocardium begins to fold into a tube (stage 29) and only become less numerous within the heart as a distinct endocardial population becomes evident (stages 30–31).
Such a first wave of migratory, “primitive” macrophages is also universal in development, providing an innate immune function to embryos. In addition to this role, there is also evidence that macrophages may play an important role in tissue and organ morphogenesis, although the precise nature of their contribution is poorly understood (reviewed in Ovchinnikov, 2008
). For example, the osteopetrotic (op
) mouse that lacks the principal growth factor for macrophages, Csf1
, displays numerous organ defects that can be attributed to a lack of macrophage participation within developing tissues (Banaei-Bouchareb et al., 2004; Van Nguyen and Pollard, 2002
). Macrophages are certainly responsible for phagocytic clearance of apoptotic corpses during embryogenesis (Henson and Hume, 2006
) and one suggestion is that they can provide growth support, or a “trophic role” during formation of complex organ structures (Pollard, 2009
). Consistent with this, transcriptional profiling indicates that macrophages synthesize a wide array of secreted proteins, including cytokines, growth factors and VEGFs (Rae et al., 2007
), which could be deposited in discrete embryonic locations. The clearest picture of migratory macrophage involvement in early embryogenesis has been described in Drosophila
larvae that possess equivalent cells called plasmatocytes (a class of hemocyte). In pvr
mutant larvae that lack the sole VEGF/PDGF receptor in flies, plasmatocyte migration and viability are critically impaired (Bruckner et al., 2004
). The lack of macrophage function is lethal, with a substantial loss of macrophage ECM deposition causing widespread morphogenesis defects including abnormalities to the ventral CNS (Olofsson and Page, 2005
The overlapping location of myeloid and cardiac domains immediately prior to and during heart tube formation in Xenopus embryos raises the possibility that macrophages may play some role in early cardiac morphogenesis. To test this possibility, we have used morpholino oligonucleotide-mediated gene knockdown to interfere with macrophage differentiation or function and examined the effect on heart formation. Our results demonstrate that amphibian heart morphogenesis is indeed critically dependent upon the presence and activity of the primitive macrophage population.