BMP-mediated inhibition of FGF signaling promotes cardiomyocyte differentiation of anterior heart field progenitors.

The anterior heart field (AHF) encompasses a niche in which mesoderm-derived cardiac progenitors maintain their multipotent and undifferentiated nature in response to signals from surrounding tissues. Here, we investigate the signaling mechanism that promotes the shift from proliferating cardiac progenitors to differentiating cardiomyocytes in chick embryos. Genomic and systems biology approaches, as well as perturbations of signaling molecules, in vitro and in vivo, reveal tight crosstalk between the bone morphogenetic protein (BMP) and fibroblast growth factor (FGF) signaling pathways within the AHF niche: BMP4 promotes myofibrillar gene expression and cardiomyocyte contraction by blocking FGF signaling. Furthermore, inhibition of the FGF-ERK pathway is both sufficient and necessary for these processes, suggesting that FGF signaling blocks premature differentiation of cardiac progenitors in the AHF. We further revealed that BMP4 induced a set of neural crest-related genes, including MSX1. Overexpression of Msx1 was sufficient to repress FGF gene expression and cell proliferation, thereby promoting cardiomyocyte differentiation. Finally, we show that BMP-induced cardiomyocyte differentiation is diminished following cranial neural crest ablation, underscoring the key roles of these cells in the regulation of AHF cell differentiation. Hence, BMP and FGF signaling pathways act via inter- and intra-regulatory loops in multiple tissues, to coordinate the balance between proliferation and differentiation of cardiac progenitors.

Distinct origins and genetic programs of head muscle satellite cells.

Adult skeletal muscle possesses a remarkable regenerative capacity, due to the presence of satellite cells, adult muscle stem cells. We used fate-mapping techniques in avian and mouse models to show that trunk (Pax3(+)) and cranial (MesP1(+)) skeletal muscle and satellite cells derive from separate genetic lineages. Similar lineage heterogeneity is seen within the head musculature and satellite cells, due to their shared, heterogenic embryonic origins. Lineage tracing experiments with Isl1Cre mice demonstrated the robust contribution of Isl1(+) cells to distinct jaw muscle-derived satellite cells. Transplantation of myofiber-associated, Isl1-derived satellite cells into damaged limb muscle contributed to muscle regeneration. In vitro experiments demonstrated the cardiogenic nature of cranial- but not trunk-derived satellite cells. Finally, overexpression of Isl1 in the branchiomeric muscles of chick embryos inhibited skeletal muscle differentiation in vitro and in vivo, suggesting that this gene plays a role in the specification of cardiovascular and skeletal muscle stem cell progenitors.

The contribution of Islet1-expressing splanchnic mesoderm cells to distinct branchiomeric muscles reveals significant heterogeneity in head muscle development.

During embryogenesis, paraxial mesoderm cells contribute skeletal muscle progenitors, whereas cardiac progenitors originate in the lateral splanchnic mesoderm (SpM). Here we focus on a subset of the SpM that contributes to the anterior or secondary heart field (AHF/SHF), and lies adjacent to the cranial paraxial mesoderm (CPM), the precursors for the head musculature. Molecular analyses in chick embryos delineated the boundaries between the CPM, undifferentiated SpM progenitors of the AHF/SHF, and differentiating cardiac cells. We then revealed the regionalization of branchial arch mesoderm: CPM cells contribute to the proximal region of the myogenic core, which gives rise to the mandibular adductor muscle. SpM cells contribute to the myogenic cells in the distal region of the branchial arch that later form the intermandibular muscle. Gene expression analyses of these branchiomeric muscles in chick uncovered a distinct molecular signature for both CPM- and SpM-derived muscles. Islet1 (Isl1) is expressed in the SpM/AHF and branchial arch in both chick and mouse embryos. Lineage studies using Isl1-Cre mice revealed the significant contribution of Isl1(+) cells to ventral/distal branchiomeric (stylohyoid, mylohyoid and digastric) and laryngeal muscles. By contrast, the Isl1 lineage contributes to mastication muscles (masseter, pterygoid and temporalis) to a lesser extent, with virtually no contribution to intrinsic and extrinsic tongue muscles or extraocular muscles. In addition, in vivo activation of the Wnt/beta-catenin pathway in chick embryos resulted in marked inhibition of Isl1, whereas inhibition of this pathway increased Isl1 expression. Our findings demonstrate, for the first time, the contribution of Isl1(+) SpM cells to a subset of branchiomeric skeletal muscles.

Mesoderm progenitor cells of common origin contribute to the head musculature and the cardiac outflow tract.

During early embryogenesis, heart and skeletal muscle progenitor cells are thought to derive from distinct regions of the mesoderm (i.e. the lateral plate mesoderm and paraxial mesoderm, respectively). In the present study, we have employed both in vitro and in vivo experimental systems in the avian embryo to explore how mesoderm progenitors in the head differentiate into both heart and skeletal muscles. Using fate-mapping studies, gene expression analyses, and manipulation of signaling pathways in the chick embryo, we demonstrate that cells from the cranial paraxial mesoderm contribute to both myocardial and endocardial cell populations within the cardiac outflow tract. We further show that Bmp signaling affects the specification of mesoderm cells in the head: application of Bmp4, both in vitro and in vivo, induces cardiac differentiation in the cranial paraxial mesoderm and blocks the differentiation of skeletal muscle precursors in these cells. Our results demonstrate that cells within the cranial paraxial mesoderm play a vital role in cardiogenesis, as a new source of cardiac progenitors that populate the cardiac outflow tract in vivo. A deeper understanding of mesodermal lineage specification in the vertebrate head is expected to provide insights into the normal, as well as pathological, aspects of heart and craniofacial development.