Pod1/Tcf21 is regulated by retinoic acid signaling and inhibits differentiation of epicardium-derived cells into smooth muscle in the developing heart
ABSTRACT Epicardium-derived cells (EPDCs) invade the myocardium and differentiate into fibroblasts and vascular smooth muscle (SM) cells, which support the coronary vessels. The transcription factor Pod1 (Tcf21) is expressed in subpopulations of the epicardium and EPDCs in chicken and mouse embryonic hearts, and the transcription factors WT1, NFATC1, and Tbx18 are expressed in overlapping and distinct subsets of Pod1-expressing cells. Expression of Pod1 and WT1, but not Tbx18 or NFATC1, is activated with all-trans-retinoic acid (RA) treatment of isolated chick EPDCs in culture. In intact chicken hearts, RA inhibition leads to decreased Pod1 expression while RA treatment inhibits SM differentiation. The requirements for Pod1 in differentiation of EPDCs in the developing heart were examined in mice lacking Pod1. Loss of Pod1 in mice leads to epicardial blistering, increased SM differentiation on the surface of the heart, and a paucity of interstitial fibroblasts, with neonatal lethality. Epicardial epithelial-to-mesenchymal transition (EMT) and endothelial differentiation of coronary vessels are relatively unaffected. On the surface of the myocardium, expression of multiple SM markers is increased in Pod1-deficient EPDCs, demonstrating premature SM differentiation. Increased SM differentiation also is observed in Pod1-deficient lung mesenchyme. Together, these data demonstrate a critical role for Pod1 in controlling mesenchymal progenitor cell differentiation into SM and fibroblast lineages during cardiac development.
SourceAvailable from: Shigeru Kuratani[Show abstract] [Hide abstract]
ABSTRACT: During cardiogenesis the epicardium, covering the surface of the myocardial tube, has been ascribed several functions essential for normal heart development of vertebrates from lampreys to mammals. We investigated a novel function of the epicardium in ventricular development in species with partial and complete septation. These species include reptiles, birds and mammals. Adult turtles, lizards and snakes have a complex ventricle with three cava, partially separated by the horizontal and vertical septa. The crocodilians, birds and mammals with origins some 100 million years apart, however, have a left and right ventricle that are completely separated, being a clear example of convergent evolution. In specific embryonic stages these species show similarities in development, prompting us to investigate the mechanisms underlying epicardial involvement. The primitive ventricle of early embryos becomes septated by folding and fusion of the anterior ventricular wall, trapping epicardium in its core. This folding septum develops as the horizontal septum in reptiles and the anterior part of the interventricular septum in the other taxa. The mechanism of folding is confirmed using DiI tattoos of the ventricular surface. Trapping of epicardium-derived cells is studied by transplanting embryonic quail pro-epicardial organ into chicken hosts. The effect of decreased epicardium involvement is studied in knock-out mice, and pro-epicardium ablated chicken, resulting in diminished and even absent septum formation. Proper folding followed by diminished ventricular fusion may explain the deep interventricular cleft observed in elephants. The vertical septum, although indistinct in most reptiles except in crocodilians and pythonidsis apparently homologous to the inlet septum. Eventually the various septal components merge to form the completely septated heart. In our attempt to discover homologies between the various septum components we aim to elucidate the evolution and development of this part of the vertebrate heart as well as understand the etiology of septal defects in human congenital heart malformations.PLoS ONE 09/2014; 9(9):e106569. DOI:10.1371/journal.pone.0106569 · 3.53 Impact Factor
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ABSTRACT: Rationale: Holt-Oram syndrome (HOS) is an autosomal dominant heart-hand syndrome caused by mutations in the TBX5 gene. Overexpression of Tbx5 in the chick proepicardial organ (PEO) impaired coronary blood vessel formation. However, the potential activity of Tbx5 in the epicardium itself, and Tbx5's role in mammalian coronary vasculogenesis, remains largely unknown. Objective: To evaluate the consequences of altered Tbx5 gene dosage during PEO and epicardial development in the embryonic chick and mouse. Methods and Results: Retroviral-mediated knockdown or upregulation of Tbx5 expression in the embryonic chick PEO as well as proepicardial-specific deletion of Tbx5 in the embryonic mouse (Tbx5(epi-/-)) impaired normal PEO cell development, inhibited epicardial and coronary blood vessel formation and altered developmental gene expression. The generation of epicardial-derived cells (EPDCs) and their migration into the myocardium was impaired between embryonic day (E) 13.5-15.5 in mutant hearts due to delayed epicardial attachment to the myocardium and subepicardial accumulation of EPDCs. This caused defective coronary vasculogenesis associated with impaired vascular smooth muscle cell recruitment, and reduced invasion of cardiac fibroblasts and endothelial cells into myocardium. In contrast to wildtype hearts that exhibited an elaborate ventricular vascular network, Tbx5(epi-/-) hearts displayed a marked decrease in vascular density that was associated with myocardial hypoxia as exemplified by HIF1α upregulation and increased binding of Hypoxyprobe-1. Tbx5(epi-/-) mice with such myocardial hypoxia exhibited reduced exercise capacity compared to wildtype mice. Conclusions: Our findings support a conserved Tbx5 dose-dependent requirement for both proepicardial and epicardial progenitor cell development in chick and mouse coronary vascular formation.Circulation Research 09/2014; 115(10). DOI:10.1161/CIRCRESAHA.115.304379 · 11.09 Impact Factor
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ABSTRACT: The molecular mechanism that regulates epicardial development has yet to be understood. In this study, we explored the function of CDX1, a Caudal-related family member, in epicardial epithelial-to-mesenchymal transition (EMT) and in the migration and the differentiation of epicardium-derived progenitors into vascular smooth muscle cells. We detected a transient expression of CDX1 in murine embryonic hearts at 11.5 days post coitum (dpc). Using a doxycycline-inducible CDX1 mouse model, primary epicardium, and ex vivo heart culture, we further demonstrated that ectopic expression of CDX1 promoted epicardial EMT. In addition, a low-dose CDX1 induction led to enhanced migration and differentiation of epicardium-derived cells into α-SMA+ vascular smooth muscles. In contrast, either continued high-level induction of CDX1 or CDX1 deficiency attenuated the ability of epicardium-derived cells to migrate and to mature into smooth muscles induced by TGF-β1. Further RNA-seq analyses showed that CDX1 induction altered the transcript levels of genes involved in neuronal development, angiogenesis, and cell adhesions required for EMT. Our data have revealed a previously undefined role of CDX1 during epicardial development, and suggest that transient expression of CDX1 promotes epicardial EMT, whereas subsequent down-regulation of CDX1 after 11.5 dpc in mice is necessary for further subepicardial invasion of EPDCs and contribution to coronary vascular endothelium or smooth muscle cells.PLoS ONE 07/2014; 9(7):e103271. DOI:10.1371/journal.pone.0103271 · 3.53 Impact Factor