Cell-ECM interactions play distinct and essential roles at multiple stages during the development of the aortic arch arteries

Rationale: Defects in the morphogenesis of the aortic arch arteries (AAAs) are among the most severe congenital birth defects. Understanding genes and mechanisms regulating AAA formation and remodeling will provide important insights into the etiology and potential treatments of congenital heart disease. Objective: Cell-ECM interactions play essential roles in the AAA morphogenesis; however, their specific functions are not well-understood. Previously, we demonstrated that integrin a5b1 and fibronectin (Fn1) expressed in the Isl1 lineage and its derivatives regulate the formation of the pharyngeal arch arteries (PAAs), the vessels giving rise to the AAAs. The objective of these studies was to investigate the mechanisms by which integrin a5b1 and Fn1 regulate AAA morphogenesis. Methods and Results: Using temporal lineage tracing, we found that endothelial progenitors of the AAA endothelium arise early during the development of the second heart field (SHF) and that the 4th PAAs contain the highest percentage of the SHF-derived ECs (ECs). To understand the role of cell-extracellular matrix (ECM) interactions in AAA development, we deleted either integrin a5 or its major extracellular ligand Fn1 in the Isl1 lineage. We used whole-mount confocal imaging to define the complex spatial and temporal EC dynamics during PAA formation at the quantitative level and assessed how cell-ECM interactions modulated these dynamics. Our analyses demonstrated that integrin a5b1 and Fn1 mediate AAA morphogenesis by regulating the accrual of SHF-derived endothelium into the 4th pharyngeal arches and the remodeling of the 4th pharyngeal arch EC plexus into the PAAs. Following PAA formation, integrin a5b1 is essential for the activation of Notch in the neural crest-derived cells surrounding the 4th PAAs and for the differentiation of the neural crest cells into vascular smooth muscle cells. Conclusions: Our data demonstrate that cell-ECM interactions regulated by integrin a5b1 and Fn1 function reiteratively during AAA development to mediate the multi-step process of AAA morphogenesis.