Abstract 1459: Tbx5 Regulates Progenitor Cell Migration During Cardiac Vasculogenesis

Mutations in TBX5 cause Holt-Oram syndrome (HOS) characterized by congenital heart malformation and upper limb deformity. Human TBX5 is expressed not only in a chamber specific myocardial distribution but also in coronary blood vessels. We have shown that mouse and chick Tbx5 are expressed in the proepicardial organ (PEO), a transitory structure from which epicardium and coronary vessels arise. We therefore investigated Tbx5 function during coronary vasculogenesis by manipulating Tbx5 gene dosage at critical points during chick and mouse cardioogenesis. Overexpression or inactivation of Tbx5 in chick PEO and chick epicardial cells leads to impaired migration and blocks incorporation of epicardial derived-cells (EPDCs) into coronary vessels. These data suggest Tbx5 acts during migration of PEO cells to the myocardium and migration of EPDCs into the subepicardium and myocardium. Epicardial restricted inactivation of Tbx5 using Wt1-cre and Tbx5 lox/lox mice reveals impaired migration (E10.5–11.5) of cytokeratin-positive PEO cells to the heart tube. However, at E14.5, mutant hearts display evidence of an epicardial layer suggesting delayed PE cell migration. However, unlike the flattened layer of epicardial cells in wildtype hearts, mutant hearts exhibit epicardial cells with irregular polygonal shapes and epicardial cyst-like structures lined by delaminated epicardium and filled with erythrocytes. At E17.5, we observe partial detachment of epicardium and nests of multiple contiguous subepicardial vessels without penetration into the myocardium. These observations suggest impaired migration of EPDCs into myocardium with consequent abnormally exuberant vasculogenesis restricted to the subepicardium. PECAM-1 expression analyses in E17.5 mutant embryos and neonatal mutant mice reveal marked reduction of myocardial coronary endothelial cells as well as defective coronary arteries and veins. These defects are associated with an upregulation of HIF1a in the myocardium, and, using an oxygen sensing dye, we observe relative tissue hypoxia in mutant myocardium. Our findings support a conserved requirement for Tbx5 in progenitor cell migration for the establishment of the coronary vasculature.