Cardiac progenitor cells

The heart is composed of diverse myogenic and non-myogenic cell lineages: cardiomyocytes, conduction system cells, endocardial endothelial cells, fibroblasts, smooth muscle cells, valvular components and connective tissue. During heart development, differentiation of these multiple cardiac lineages is spatially and temporally controlled, resulting in the coordinated formation of the distinct tissue components of the heart (Harvey, 2002). In this regard, tissue ablation, genetic ablation, and lineage labeling experiments have demonstrated the dynamic nature of heart tube formation, and the existence of two myocardial sources in the mouse embryo (Buckingham et al., 2005; Evans et al., 2010; Kelly, 2005). According to this model, growth of the embryonic heart occurs by progressive addition of progenitor cells of the second heart field (SHF), situated in pharyngeal mesoderm, to the poles of the heart tube, itself derived from the cardiac crescent or first heart field.Cells from the SHF give rise to a large part of the heart including atrial, atrio-ventricular, right ventricular and outflow tract (OFT) myocardium (Buckingham et al., 2005; Evans et al., 2010). Discovery of the SHF has had a major impact on our understanding of heart development and it is now well established that perturbation of SHF development results in a spectrum of common congenital heart malformations (Rochais et al., 2009; Vincent and Buckingham, 2010).

Acide Rétino´que pendant l'organogénèse précoce

Figure 1: Retinoic acid signalling during early organogenesis.

Retinoic acid (RA) generated by retinaldehyde deshydrogen-ase 2 (Raldh2) in the lateral mesoderm travels anteriorly where it provides a signal that establishes the posterior border of the heart field. RA may function as a repressor of cardiac progenitors.

The correct establishment of anterior-posterior polarity in the vertebrate embryonic heart tube during embryogenesis is crucial for the proper morphogenesis of the mature heart. Molecular details of this process are poorly understood. There is now considerable evidence that retinoic acid (RA) is a morphogen that communicates anterior-posterior polarity to the heart (reviewed in Zaffran & Niederreither, 2015). Using several approaches to examine the contribution of the SHF to pharyngeal mesoderm, atria and OFT in RA-deficient mouse embryos, we have recently shown that RA is required to restrict the SHF posteriorly (Ryckebusch et al., 2008). Mouse embryos lacking RALDH2 exhibit an increase in Fgf8 and Isl1 cardiac expression posteriorly (Fig 1). This study suggests that RA regulates SHF organisation and furthermore indicate that this occurs within a precise time frame. More recently, we have revealed Hox gene expression in the SHF. Our analysis shown that expression of Hox genes in the SHF defines new cardiac progenitor sub-domains. A genetic lineage tracing analysis shown that cardiac Hox-positive cells contribute to atria and inferior wall of the outlfow tract, which subsequently gives rise to sub-pulmonary myocardium (Fig 2). Reducion or excess of RA demonstrate that Hox-positive SHF cells are RA sensitive.

hox

Figure 2: Model for cardiac contributions of progenitor cells expressing Hox genes in the second heart field.

Frontal view is shown for embryonic day 7.5 (E7.5) and lateral view for E8.5. Early Hoxb1/a1/a3 expressing cells characterize distinct sub-domains along the antero-posterior axis in the SHF. Later, these cardiac progenitor cells contribute to both atria and the inferior wall of the OFT, which subsequently gives rise to myocardium at the base of pulmonary trunk. Ao, aorta; CC, cardiac crescent; ep, epicardium; ht, heart tube; LA, left atria; Pt, pulmonary trunk; RA, right atria; r4, rhombomere 4.

Congenital heart defects

Congenital heart defects are the most common class of birth defect and about 30% affects the conotruncal region (also called the outflow tract (OFT)), which gives rise to the connection between the ventricles and the great arteries. We have recently shown that Hoxa1 and Hoxb1 are expressed in a sub-population of the SHF contributing to the OFT (Bertand et al., 2011). We now report that Hoxa1 and Hoxb1 are required for correct OFT development (Roux et al., 2015). Indeed, Hoxb1 deficiency results in a shorter OFT and ventricular septal defects (VSD). Lack of Hoxb1 perturbs second heart field development through premature myocardial differentiation. Hence, the positioning and remodeling of the mutant OFT is disrupted. Hoxa1-/- embryos, in contrast, have low percentage of VSD and normal SHF development. However, compound Hoxa1-/-;Hoxb1+/- embryos display OFT defects associated with premature SHF differentiation, demonstrating redundant roles of these factors during OFT development (Fig 3). Therefore, our work identifies a novel role for anterior Hox genes in cardiac progenitor cells that contribute to the formation of the OFT (also called conotruncal region).

model

Figure 3: Model showing premature differentiation of cardiac progenitor cells in the SHF of WT, Hoxb1-/- and compound Hoxa1-/-;Hoxb1+/- embryos

The common clinical features associated with DiGeorge/velo-cardio-facial syndrome (DGS/VCFS) are congenital cardiovascular malformations, thymic and parathyroid aplasia or hypoplasia and craniofacial defects. However, despite the vast majority of patients diagnosed with DGS/VCFS having the same region of 22q11.2 deleted, there is wide phenotypic variation. The causes of this phenotype variability remain unknown. Genetic studies have identified the transcription factor encoding gene Tbx1 as a major candidate gene for DGS/VCFS. Tbx1 heterozygous mutant mice display absence or hypoplasia of the 4th pharyngeal arch artery (PAA) at E10.5 and associated great artery anomalies at later stages of development. Retinoic acid (RA) interacts with Tbx1, as Tbx1 expression was increased in Raldh2-/- deficient embryos and reduction of RA synthesis modulates development of the 4th PAA in Tbx1+/- embryos by accelerating the recovery of the arterial growth delay (Ryckebusch et al., 2010). RA deficiency therefore induces earlier recovery of DiGeorge-related aortic arch defects, supporting the hypothesis that differences in levels of embryonic RA may contribute to the phenotypic variability observed in DGS/VCFS patients (Fig 4). Genes involved in RA synthesis, metabolism and signaling should be considered candidate modifiers of the DGS/VCFS phenotype associated with 22q11deletion.

graphic

Figure 4: Percentage of Raldh2+/-, Tbx1+/- and Raldh2+/-

Tbx1+/- embryos with 4th PAAs or 4th derived defects at E10.5, E11.5 and fetal stages. The percentage of embryos that overcomes the primary 4th PAA defects at E11.5 is significantly increased in compound heterozygous mutants compare to single Tbx1+/- (*p inferior to 0.01). Surprisingly, the difference observed at E11.5 is decreased at later stages when a majority of Tbx1+/- embryos overcome the primary 4th PAA defect.

These works were supported by the AFM-Telethon (grants 13517/14134 and NMH-Decrypt), the FRM, and the ANR (ANR-07-MRAR-003 and ANR-13-BSV2-0003)

retour en haut de page