Symmetry and fluctuation of cell movements in neural crest-derived facial mesenchyme

In the face, symmetry is established when bilateral streams of neural crest cells leave the neural tube at the same time, follow identical migration routes and then give rise to the facial prominences. However developmental instability exists, particularly surrounding the steps of lip fusion. The causes of instability are unknown but inability to cope with developmental fluctuations are a likely cause of congenital malformations such as non-syndromic orofacial clefts. Here, we tracked cell movements over time in the frontonasal mass which forms the facial midline and participates in lip fusion using live-cell imaging. Our mathematical examination of cell velocity vectors uncovered temporal fluctuations in several parameters including order/disorder, symmetry/asymmetry and divergence/convergence. We found that treatment with a RhoGTPase inhibitor completely disrupted the temporal fluctuations in all measures and blocked morphogenesis. Thus we discovered that genetic control of symmetry extends to mesenchymal cell movements and that these movements are of the type that could be perturbed in in asymmetrical malformations such as non-syndromic cleft lip.

Development of the squamate naso-palatal complex: detailed 3D analysis of the vomeronasal organ and nasal cavity in the brown anole Anolis sagrei (Squamata: Iguania)

Background Despite the diverse morphology of the adult squamate naso-palatal complex – consisting of the nasal cavity, vomeronasal organ (VNO), choanal groove, lacrimal duct and superficial palate – little is known about the embryology of these structures. Moreover, there are no comprehensive studies concerning development of the nasal cavity and VNO in relation to the superficial palate. In this investigation, we used X-ray microtomography and histological sections to describe embryonic development of the naso-palatal complex of iguanian lizard, the brown anole (Anolis sagrei). The purpose of the study was to describe the mechanism of formation of adult morphology in this species, which combines the peculiar anole features with typical iguanian conditions. Considering the uncertain phylogenetic position of the Iguania within Squamata, embryological data and future comparative studies may shed new light on the evolution of this large squamate clade. Results Development of the naso-palatal complex was divided into three phases: early, middle and late. In the early developmental phase, the vomeronasal pit originates from medial outpocketing of the nasal pit, when the facial prominences are weakly developed. In the middle developmental phase, the following events can be noted: the formation of the frontonasal mass, separation of the vestibulum, appearance of the lacrimal duct, and formation of the choanal groove, which leads to separation of the VNO from the nasal cavity. In late development, the nasal cavity and the VNO attain their adult morphology. The lacrimal duct establishes an extensive connection with the choanal groove, which eventually becomes largely separated from the oral cavity. Conclusions Unlike in other tetrapods, the primordium of the lacrimal duct in the brown anole develops largely beyond the nasolacrimal groove. In contrast to previous studies on squamates, the maxillary prominence is found to participate in the initial fusion with the frontonasal mass. Moreover, formation of the choanal groove occurs due to the fusion of the vomerine cushion to the subconchal fold, rather than to the choanal fold. The loss or significant reduction of the lateral nasal concha is secondary. Some features of anole adult morphology, such as the closure of the choanal groove, may constitute adaptations to vomeronasal chemoreception.

Development of the squamate naso-palatal complex: detailed 3D analysis of the vomeronasal organ and nasal cavity in the brown anole Anolis sagrei (Squamata: Iguania)

Background Despite the diverse morphology of the adult squamate naso-palatal complex – consisting of the nasal cavity, vomeronasal organ (VNO), choanal groove, lacrimal duct and superficial palate – little is known about the embryology of these structures. Moreover, there are no comprehensive studies concerning development of the nasal cavity and VNO in relation to the superficial palate. In this investigation, we used X-ray microtomography and histological sections to describe embryonic development of the naso-palatal complex of iguanian lizard, the brown anole ( Anolis sagrei ). The purpose of the study was to describe the mechanism of formation of adult morphology in this species, which combines the peculiar anole features with typical iguanian conditions. Considering the uncertain phylogenetic position of the Iguania within Squamata, embryological data and future comparative studies may shed new light on the evolution of this large squamate clade. Results Development of the naso-palatal complex was divided into three phases: early, middle and late. In the early developmental phase, the vomeronasal pit originates from medial outpocketing of the nasal pit, when the facial prominences are weakly developed. In the middle developmental phase, the following events can be noted: the formation of the frontonasal mass, separation of the vestibulum, appearance of the lacrimal duct, and formation of the choanal groove, which leads to separation of the VNO from the nasal cavity. In late development, the nasal cavity and the VNO attain their adult morphology. The lacrimal duct establishes an extensive connection with the choanal groove, which eventually becomes largely separated from the oral cavity. Conclusions Unlike in other tetrapods, the primordium of the lacrimal duct in the brown anole develops largely beyond the nasolacrimal groove. In contrast to previous studies on squamates, the maxillary prominence is found to participate in the initial fusion with the frontonasal mass. Moreover, formation of the choanal groove occurs due to the fusion of the vomerine cushion to the subconchal fold, rather than to the choanal fold. The loss or significant reduction of the lateral nasal concha is secondary. Some features of anole adult morphology, such as the closure of the choanal groove, may constitute adaptations to vomeronasal chemoreception.

Development of the squamate naso-palatal complex: detailed 3D analysis of the vomeronasal organ and nasal cavity in the brown anole Anolis sagrei (Squamata: Iguania)

Background Despite the diverse morphology of the adult squamate naso-palatal complex – consisting of the nasal cavity, vomeronasal organ (VNO), choanal groove, lacrimal duct and soft palate –little is known about the embryology of these structures. Moreover, there are no comprehensive studies concerning development of the nasal cavity and VNO in relation to the soft palate. In this investigation, we used X-ray microtomography and light microscopy to describe embryonic development of the naso-palatal complex of iguanian lizard, the brown anole ( Anolis sagrei ). The purpose of the study was to describe the mechanism of formation of adult morphology in this species, which combines the peculiar anole features with iguanian conditions. Considering the uncertain phylogenetic position of the Iguania within Squamata, embryological data and future comparative studies may shed new light on the evolution of this large squamate clade. Results Development of the naso-palatal complex was divided into three phases: early, middle and late. In the early developmental phase, the vomeronasal pit originates from medial outpocketing of the nasal pit, when the facial prominences are weakly developed. In the middle developmental phase, the following events can be noted: the formation of the frontonasal mass, separation of the vestibulum, appearance of the lacrimal duct, and formation of the choanal groove, which leads to separation of the VNO from the nasal cavity. In late development, the nasal cavity and the VNO attain their adult-like morphology. The lacrimal duct establishes an extensive connection with the choanal groove, which eventually becomes largely separated from the oral cavity. Conclusions Unlike in other tetrapods, the primordium of the lacrimal duct in the brown anole develops largely beyond the nasolacrimal groove. In contrast to previous studies on squamates, the maxillary prominence is found to participate in the initial fusion with the frontonasal mass. Moreover, formation of the choanal groove occurs due to the fusion of the vomerine cushion to the subconchal fold, rather than to the choanal fold. The loss or significant reduction of the lateral nasal concha is secondary. Some features of anole adult-like morphology, such as the closure of the choanal groove, may constitute adaptations to vomeronasal chemoreception.

Endogenous bone morphogenetic proteins regulate outgrowth and epithelial survival during avian lip fusion

Our expression studies of bone morphogenetic proteins (BMPs) and Noggin (a BMP antagonist) in the embryonic chicken face suggested that BMP signals were important for closure of the upper lip or primary palate. We noted that Noggin expression was restricted to the frontonasal mass epithelium but was reduced at the corners of the frontonasal mass (globular processes) just prior to fusion with the adjacent maxillary prominences. We therefore performed gain- and loss-of-function experiments to determine the role of BMPs in lip formation. Noggin treatment led to reduced proliferation and outgrowth of the frontonasal mass and maxillary prominences and ultimately to the deletion of the maxillary and palatine bones. The temporary block in BMP signalling in the mesenchyme also promoted epithelial survival. Noggin treatment also upregulated expression of endogenous BMPs, therefore we investigated whether increasing BMP levels would lead to the same phenotype. A BMP2 bead was implanted into the globular process and a similar phenotype to that produced by Noggin resulted. However, instead of a decrease in proliferation, defects were caused by increased programmed cell death, first in the epithelium and then in the mesenchyme. Programmed cell death was induced primarily in the lateral frontonasal mass with very little cell death medial to the bead. The asymmetric cell death pattern was correlated with a rapid induction of Noggin in the same embryos, with transcripts complementary to the regions with increased cell death. We have demonstrated a requirement for endogenous BMP in the proliferation of facial mesenchyme and that mesenchymal signals promote either survival or thinning of the epithelium. We furthermore demonstrated in vivo that BMP homeostasis is regulated by increasing expression of ligand or antagonist and that such mechanisms may help to protect the embryo from changes in growth factor levels during development or after exposure to teratogens.