Epicardial cell shape and maturation are regulated by Wt1 via transcriptional control of Bmp4

Abstract

The epicardium plays a crucial role in embryonic heart development and adult heart repair; however, the molecular events underlying its maturation remain unknown. Wt1, one of the main markers of the embryonic epicardium, is essential for epicardial development and function. Here, we analyse the transcriptomic profile of epicardialenriched cells at different stages of development and from control and epicardial-specific Wt1 knockout (Wt1KO) mice. Transcriptomic and cell morphology analyses of epicardial cells from epicardial-specific Wt1KO mice revealed a defect in the maturation process of the mutant epicardium, including sustained upregulation of Bmp4 expression and the inability of mutant epicardial cells to transition into a mature squamous phenotype. We identified Bmp4 as a transcriptional target of Wt1, thus providing a molecular basis for the retention of the cuboidal cell shape observed in the Wt1KO epicardium. Accordingly, inhibition of the Bmp4 signalling pathway both ex vivo and in vivo rescued the cuboidal phenotype of the mutant epicardium. Our findings indicate the importance of the cuboidal-to-squamous transition in epicardial maturation, a process regulated by Wt1.

Epicardial TGFβ and BMP Signaling in Cardiac Regeneration: What Lesson Can We Learn from the Developing Heart?

“…In contrast, in these embryos with epicardial specific knock-down of ALK3, neither the development of the pro-epicardium, the formation of the epicardium, nor the migration of EPDCs into the myocardium were affected [108]. Recently, BMP signaling was described to be involved in epicardial maturation [42]. Epicardial maturation is the process that starts after the epicardium has formed, during which the cells turn from a cuboidal into a squamous epithelial cell layer, characterized by flattening of the cells and elongation of the nuclei [42].…”

“…In WT1KO embryos, this process is impaired, resulting in persistence of cuboidal cells, which coincided with a higher expression of epicardial BMP4 compared to wildtype embryos. Addition of BMP inhibitor LDN-193189 rescued the phenotype of WT1KO embryos, revealing that BMP signaling needs to be absent for proper epicardial maturation [42] (see Figure 3).…”

“…From E9.5 onwards, cells from the pro-epicardium (PE), a cell cluster located at the venous inflow tract, migrate towards the heart and proliferate to cover the bare myocardium with a third cardiac layer: the epicardium [41]. By E11.5, the epicardial cells have enveloped the heart and at this point display a cuboidal epithelial phenotype [42], characterized by the expression of cell adhesion molecules such as E-cadherin and β-catenin, that ascertain integrity of the layer and the maintenance of an apical-basal polarity [43]. Furthermore, the epicardial cells express Wilms' tumor 1 (WT1) and retinaldehyde dehydrogenase 2 (RALDH2) [44], proteins that are often used as markers for activated epicardium.…”

“…Recently, BMP signaling was described to be involved in epicardial maturation [42]. Epicardial maturation is the process that starts after the epicardium has formed, during which the cells turn from a cuboidal into a squamous epithelial cell layer, characterized by flattening of the cells and elongation of the nuclei [42]. In WT1KO embryos, this process is impaired, resulting in persistence of cuboidal cells, which coincided with a higher expression of epicardial BMP4 compared to wildtype embryos.…”

“…Another opportunity to activate the epicardium of the injured heart could be via BMP4 signaling. During development, BMP4 was shown to be vital for epicardial maturation [42]. Thus far, the exact nature and function of epicardial maturation is not extensively described, but the fact that two of the main epicardial specific proteins, WT1 [42] and TCF21 [147], are proposed to be involved in this process indicates that it is an essential part of epicardial development.…”