Abstract
Parathyroid hormone-related peptide (PTHrP) has been implicated in mediating proliferation and differentiation in various tissues in the developing embryo but its role in early heart development has not been investigated. This study examined the spatial and temporal localization of chicken PTHrP (1-36) and PTHrP-binding sites utilizing immunohistochemistry during cardiac development. A PTHrP antagonist, PTHrP (7-34), was used to ascertain if PTHrP regulates cardiac looping during heart morphogenesis. Stage 10 to 14 chick embryos were immunostained for PTHrP using a PTHrP-specific antibody and PTHrP-binding sites (i.e., target cells) were localized using biotinylated-PTHrP (1-36). From stages 10 to 14 PTHrP immunostaining was greatest along the outer curvatures of the ventricle and atrium compared to their inner curvatures during formation of the initial C- shaped heart. PTHrP-binding sites did not mimic the asymmetry observed with PTHrP immunostaining until stage 12. In the outflow tract, PTHrP immunostaining maintained a high staining intensity throughout all the stages studied while biotinylated-PTHrP binding decreased gradually by stage 14. PTHrP immunostaining decreased between stages 15 to 20 except for in the outflow tract, areas surrounding endocardial-derived cushion cells, and developing trabeculae carnae. Therefore, immunostaining for PTHrP and PTHrP-binding sites was localized within areas thought to undergo cardiac cell proliferation and differentiation. To test for a functional role of PTHrP, a PTHrP antagonist was microinjected into the pericardial cavity prior to cardiac looping and the embryos were allowed to develop to stages 14 to 16. No detectable effect on cardiac morphogenesis was observed.Limitations in the in ovo model used may have resulted in the inability of the PTHrP antagonist to impede or affect cardiac looping at the stages studied but PTHrP immunolocalization and PTHrP-binding site staining did suggest PTHrP may have an important but unknown role in mediating the regional growth and expansion of the looping heart.