Overlapping origins of pharyngeal arch crest cells on the postotic hind-brain

The developing hind-brain of vertebrates consists of segmental units called rhombomeres. Although crest cells emigrate from the hind-brain, they are subsequently subdivided into several cell populations that are attached to restricted regions of the hind-brain. At the preotic level, only even-numbered rhombomeres are accompanied by crest cells, while the odd-numbered ones are not. At the postotic level, such the birhombomeric repetition becomes obscure. In order to map the origins and distributions of postotic crest cells, focal injections of Dil were made into various axial levels of the postotic neural tube. Cephalic crest cells at the postotic level first form a single cell population deposited by cells along the dorsolateral pathway. They are called the circumpharyngeal crest cells (CP cells) and are secondarily subdivided into each pharyngeal arch ectomesenchyme. The neural tube extending from r5 to the somite 3/4 boundary gave rise to CP cells. The neuraxial origins of each pharyngeal ectomesenchyme extended for more than three somite lengths, most of which overlapped with the other. Unlike in the preotic region, there is no segmental registration between neuraxial levels and pharyngeal arches. Caudal portions of the CP cell population show a characteristic distribution pattern that circumscribes the postotic pharyngeal arches caudally. Heterotopic transplantation of the Dil-labeled neural crest into the somite 3 level resulted in a distribution of labeled cells similar to that of CP cells, suggesting that the pattern of distribution depends upon dynamic modification of the body wall associated with pharyngeal arch formation.     

Evolutionary developmental biology and vertebrate head segmentation: A perspective from developmental constraint

Summary The question of vertebrate head segmentation has become one of the central issues in Evolutionary Developmental Biology. Beginning as a theory based in comparative anatomy, a segmental theory of the head has been adopted and further developed by comparative embryologists. With the use of molecular and cellular biology, and in particular analyses of the Hox gene complex, the question has been addressed at new levels, but it remains unresolved. In this review, vertebrate head segmentation is reevaluated, by introducing findings from experimental embryology and evolutionary biology. Developmental biology has shown that pattern is generated through hierarchically organized and causally linked series of events. The question of head segmentation can be viewed as a question of generative constraint, that is whether segmentation in the head is imposed by underlying segmental patterns, as it is in the trunk. In this respect, amphioxus appears to be segmented along the entire anteroposterior axis, with myotomes and peripheral nerves repeating with the same rhythm (somitomerism). Similarly, in the vertebrate trunk, the segmental patterns shared by myotomes, peripheral nerves and vertebrae are derived from the somites. However, in the head of vertebrates there is no such mesodermal pattern, although neuromerism and branchiomerism do indicate the presence of constraints derived from rhombomeres and pharyngeal pouches, respectively. These data fit better the concept of dual metamerism of the vertebrate body proposed by Romer (1972), than the traditional head cavity-based segmental model by Goodrich (1930).     

Disruption of Smad4 in neural crest cells leads to mid-gestation death with pharyngeal arch, craniofacial and cardiac defects

TGFβ/BMP signaling pathways are essential for normal development of neural crest cells (NCCs). Smad4 encodes the only common Smad protein in mammals, which is a critical nuclear mediator of TGFβ/BMP signaling. In this work, we sought to investigate the roles of Smad4 for development of NCCs. To overcome the early embryonic lethality of Smad4 null mice, we specifically disrupted Smad4 in NCCs using a Cre/loxP system. The mutant mice died at mid-gestation with defects in facial primordia, pharyngeal arches, outflow tract and cardiac ventricles. Further examination revealed that mutant embryos displayed severe molecular defects starting from E9.5. Expression of multiple genes, including Msx1, 2, Ap-2α, Pax3, and Sox9, which play critical roles for NCC development, was downregulated by NCC disruption of Smad4. Moreover, increased cell death was observed in pharyngeal arches from E10.5. However, the cell proliferation rate in these areas was not substantially altered. Taken together, these findings provide compelling genetic evidence that Smad4-mediated activities of TGFβ/BMP signals are essential for appropriate NCC development.     

Hox genes, neural crest cells and branchial arch patterning

Proper craniofacial development requires the orchestrated integration of multiple specialized tissue interactions. Recent analyses suggest that craniofacial development is not dependent upon neural crest pre-programming as previously thought but is regulated by a more complex integration of cell and tissue interactions. In the absence of neural crest cells it is still possible to obtain normal arch patterning indicating that neural crest is not responsible for patterning all of arch development. The mesoderm, endoderm and surface ectoderm tissues play a role in the patterning of the branchial arches, and there is now strong evidence that Hoxa2 acts as a selector gene for the pathways that govern second arch structures.     

What's retinoic acid got to do with it? Retinoic acid regulation of the neural crest in craniofacial and ocular development

Retinoic acid (RA), the active derivative of vitamin A (retinol), is an essential morphogen signaling molecule and major regulator of embryonic development. The dysregulation of RA levels during embryogenesis has been associated with numerous congenital anomalies, including craniofacial, auditory, and ocular defects. These anomalies result from disruptions in the cranial neural crest, a vertebrate‐specific transient population of stem cells that contribute to the formation of diverse cell lineages and embryonic structures during development. In this review, we summarize our current knowledge of the RA‐mediated regulation of cranial neural crest induction at the edge of the neural tube and the migration of these cells into the craniofacial region. Further, we discuss the role of RA in the regulation of cranial neural crest cells found within the frontonasal process, periocular mesenchyme, and pharyngeal arches, which eventually form the bones and connective tissues of the head and neck and contribute to structures in the anterior segment of the eye. We then review our understanding of the mechanisms underlying congenital craniofacial and ocular diseases caused by either the genetic or toxic disruption of RA signaling. Finally, we discuss the role of RA in maintaining neural crest‐derived structures in postembryonic tissues and the implications of these studies in creating new treatments for degenerative craniofacial and ocular diseases.     

Developmental Morphology of the Head Mesoderm and Reevaluation of Segmental Theories of the Vertebrate Head: Evidence from Embryos of an Agnathan Vertebrate, Lampetra japonica

Due to the peculiar morphology of its preotic head, lampreys have long been treated as an intermediate animal which links amphioxus and gnathostomes. To reevaluate the segmental theory of classical comparative embryology, mesodermal development was observed in embryos of a lamprey, Lampetra japonica, by scanning electron microscopy and immunohistochemistry. Signs of segmentation are visible in future postotic somites at an early neurula stage, whereas the rostral mesoderm is unsegmented and rostromedially confluent with the prechordal plate. The premandibular and mandibular mesoderm develop from the prechordal plate in a caudal to rostral direction and can be called the preaxial mesoderm as opposed to the caudally developing gastral mesoderm. With the exception of the premandibular mesoderm, the head mesodermal sheet is secondarily regionalized by the otocyst and pharyngeal pouches into the mandibular mesoderm, hyoid mesoderm, and somite 0. The head mesodermal components never develop into cephalic myotomes, but the latter develop only from postotic somites. These results show that the lamprey embryo shows a typical vertebrate phylotype and that the basic mesodermal configuration of vertebrates already existed prior to the split of agnatha-gnathostomata; lamprey does not represent an intermediate state between amphioxus and gnathostomes. Unlike interpretations of theories of head segmentation that the mesodermal segments are primarily equivalent along the axis, there is no evidence in vertebrate embryos for the presence of preotic myotomes. We conclude that mesomere-based theories of head metamerism are inappropriate and that the formulated vertebrate head should possess the distinction between primarily unsegmented head mesoderm which includes preaxial components at least in part and somites in the trunk which are shared in all the known vertebrate embryos as the vertebrate phylotype.     

Constitutive expression of preproendothelin in the cardiac neural crest selectively promotes expansion of the adventitia of the great vessels in vivo

Cardiac neural crest cells are essential for normal development of the great vessels and the heart, giving rise to a range of cell types, including both neuronal and non-neuronal adventitial cells and smooth muscle. Endothelin (ET) signaling plays an important role in the development of cardiac neural crest cell lineages, yet the underlying mechanisms that act to control their migration, differentiation, and proliferation remain largely unclear. We examined the expression patterns of the receptor, ET(A), and the ET-specific converting enzyme, ECE-1, in the pharyngeal arches and great vessels of the developing chick embryo. In situ hybridization analysis revealed that, while ET(A) is expressed in the pharyngeal arch mesenchyme, populated by cardiac neural crest cells, ECE-1 expression is localized to the outermost ectodermal cells of the arches and then to the innermost endothelial cells of the great vessels. This dynamic pattern of expression suggests that only a subpopulation of neural crest cells in these regions is responsive to ET signaling at particular developmental time points. To test this, retroviral gene delivery was used to constitutively express preproET-1, a precursor of mature ET-1 ligand, in the cardiac neural crest. This resulted in a selective expansion of the outermost, adventitial cell population in the great vessels. In contrast, neither differentiation nor proliferation of neural crest-derived smooth muscle cells was significantly affected. These results suggest that constitutive expression of exogenous preproET-1 in the cardiac neural crest results in expansion restricted to an adventitial cell population of the developing great vessels.     

Endothelin regulates neural crest deployment and fate to form great vessels through Dlx5/Dlx6-independent mechanisms

Endothelin-1 (Edn1), originally identified as a vasoconstrictor peptide, is involved in the development of cranial/cardiac neural crest-derived tissues and organs. In craniofacial development, Edn1 binds to Endothelin type-A receptor (Ednra) to induce homeobox genes Dlx5/Dlx6 and determines the mandibular identity in the first pharyngeal arch. However, it remains unsolved whether this pathway is also critical for pharyngeal arch artery development to form thoracic arteries. Here, we show that the Edn1/Ednra signaling is involved in pharyngeal artery development by controlling the fate of neural crest cells through a Dlx5/Dlx6-independent mechanism. Edn1 and Ednra knock-out mice demonstrate abnormalities in pharyngeal arch artery patterning, which include persistent first and second pharyngeal arteries, resulting in additional branches from common carotid arteries. Neural crest cell labeling with Wnt1-Cre transgene and immunostaining for smooth muscle cell markers revealed that neural crest cells abnormally differentiate into smooth muscle cells at the first and second pharyngeal arteries of Ednra knock-out embryos. By contrast, Dlx5/Dlx6 knockout little affect the development of pharyngeal arch arteries and coronary arteries, the latter of which is also contributed by neural crest cells through an Edn-dependent mechanism. These findings indicate that the Edn1/Ednra signaling regulates neural crest differentiation to ensure the proper patterning of pharyngeal arch arteries, which is independent of the regional identification of the pharyngeal arches along the dorsoventral axis mediated by Dlx5/Dlx6.