Bone labeling experiments and intraskeletal growth patterns in captive leopard geckos (Eublepharis macularius).

An understanding of the dynamics of bone growth is key to interpreting life-history parameters of vertebrates. In this study, we used fluorochrome labels in captive leopard geckos (Eublepharis macularius) to track bone growth and intraskeletal variability from embryonic to adult growth stages. Thirteen individuals were administered fluorochromes from pre-hatching to 4 years of age.

Resilience of the replacing dentition in adult reptiles.

The dentition is critical to animal survival and teeth are present in modern vertebrates including teleost fish, sharks, amphibians, mammals and reptiles. The developmental processes that give rise to teeth are not just preserved through evolution but also share high level of similarity with the embryogenesis of other ectodermal organs. In this review we go beyond the embryonic phase of tooth development to life-long tooth replacement.

Craniofacial studies in chicken embryos confirm the pathogenicity of human FZD2 variants associated with Robinow syndrome.

Robinow syndrome is a rare disease caused by variants of seven WNT pathway genes. Craniofacial features include widening of the nasal bridge and jaw hypoplasia. We used the chicken embryo to test whether two missense human FZD2 variants (1301G>T, p.

3D atlas of the human fetal chondrocranium in the middle trimester.

The chondrocranium provides the key initial support for the fetal brain, jaws and cranial sensory organs in all vertebrates. The patterns of shaping and growth of the chondrocranium set up species-specific development of the entire craniofacial complex. The 3D development of chondrocranium have been studied primarily in animal model organisms, such as mice or zebrafish.

Tissues and signals with true organizer properties in craniofacial development.

Transplantation experiments have shown that a true organizer provides instructive signals that induce and pattern ectopic structures in the responding tissue. Here, we review craniofacial experiments to identify tissues with organizer properties and signals with organizer properties. In particular, we evaluate whether transformation of identity took place in the mesenchyme.

Mechanistic studies in Drosophila and chicken give new insights into functions of DVL1 in dominant Robinow syndrome.

The study of rare genetic diseases provides valuable insights into human gene function. The autosomal dominant or autosomal recessive forms of Robinow syndrome are genetically heterogeneous, and the common theme is that all the mutations lie in genes in Wnt signaling pathways. Cases diagnosed with Robinow syndrome do survive to adulthood with distinct skeletal phenotypes, including limb shortening and craniofacial abnormalities.

Tooth Removal in the Leopard Gecko and the Formation of Replacement Teeth.

Many reptiles are able to continuously replace their teeth through life, an ability attributed to the existence of epithelial stem cells. Tooth replacement occurs in a spatially and temporally regulated manner, suggesting the involvement of diffusible factors, potentially over long distances. Here, we locally disrupted tooth replacement in the leopard gecko () and followed the recovery of the dentition.

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.

The Effects of Premature Tooth Extraction and Damage on Replacement Timing in the Green Iguana.

Reptiles with continuous tooth replacement, or polyphyodonty, replace their teeth in predictable, well-timed waves in alternating tooth positions around the mouth. This process is thought to occur irrespective of tooth wear or breakage. In this study, we aimed to determine if damage to teeth and premature tooth extraction affects tooth replacement timing long-term in juvenile green iguanas (Iguana iguana).

Getting out of an egg: Merging of tooth germs to create an egg tooth in the snake.

Background: The egg tooth is a vital structure allowing hatchlings to escape from the egg. In squamates (snakes and lizards), the egg tooth is a real tooth that develops within the oral cavity at the top of the upper jaw. Most squamates have a single large midline egg tooth at hatching, but a few families, such as Gekkonidae, have two egg teeth.

Analysis of facial skeletal asymmetry during foetal development using μCT imaging.

Objectives: Asymmetry has been noted in the human craniofacial region in several pathological conditional and growth abnormalities, often with a directional predilection. Physiological asymmetry has also been reported in normal adults and adolescents, with certain regions of the cranioskeleton, such as the mandible, displaying prevalent asymmetry. However, the timing at which such asymmetries arise has not been evaluated.

Robinow syndrome skeletal phenotypes caused by the WNT5AC83S variant are due to dominant interference with chondrogenesis.

Heterozygous missense mutations in several genes in the WNT5A signaling pathway cause autosomal dominant Robinow syndrome 1 (DRS1). Our objective was to clarify the functional impact of a missense mutation in WNT5A on the skeleton, one of the main affected tissues in RS. We delivered avian replication competent retroviruses (RCAS) containing human wild-type WNT5A (wtWNT5A), WNT5AC83S variant or GFP/AlkPO4 control genes to the chicken embryo limb.

Craniofacial development: discoveries made in the chicken embryo.

The aim of this review is to highlight some of the key contributions to our understanding of craniofacial research from work carried out with the chicken and other avian embryos. From the very first observations of neural crest cell migration to the fusion of the primary palate, the chicken has proven indispensable in facilitating craniofacial research. In this review we will look back to the premolecular studies where "cut and paste" grafting experiments mapped the fate of cranial neural crest cells, the role of different tissue layers in patterning the face, and more recently the contribution of neural crest cells to jaw size and identity.

Coordination of bilateral tooth replacement in the juvenile gecko is continuous with in ovo patterning.

We performed a test of how function impacts a genetically programmed process that continues into postnatal life. Using the dentition of the polyphyodont gecko as our model, tooth shedding was recorded longitudinally across the jaw. We compared two time periods: one in which teeth were patterned symmetrically in ovo and a later period when teeth were initiated post-hatching.

Design of lipid nanoparticles for in vitro and in vivo delivery of plasmid DNA.

Lipid nanoparticles (LNPs) containing distearoylphosphatidlycholine (DSPC), and ionizable amino-lipids such as dilinoleylmethyl-4-dimethylaminobutyrate (DLin-MC3-DMA) are potent siRNA delivery vehicles in vivo. Here we explore the utility of similar LNP systems as transfection reagents for plasmid DNA (pDNA). It is shown that replacement of DSPC by unsaturated PCs and DLin-MC3-DMA by the related lipid DLin-KC2-DMA resulted in highly potent transfection reagents for HeLa cells in vitro.

MORN5 Expression during Craniofacial Development and Its Interaction with the BMP and TGFβ Pathways.

MORN5 (MORN repeat containing 5) is encoded by a locus positioned on chromosome 17 in the chicken genome. The MORN motif is found in multiple copies in several proteins including junctophilins or phosphatidylinositol phosphate kinase family and the MORN proteins themselves are found across the animal and plant kingdoms. MORN5 protein has a characteristic punctate pattern in the cytoplasm in immunofluorescence imaging.

Pannexin 3 is required for late stage bone growth but not for initiation of ossification in avian embryos.

Background: Pannexin 3 (PANX3) is a channel-forming protein capable of stimulating osteogenesis in vitro. Here, we studied the in vivo roles of PANX3 in the chicken embryo using the RCAS retroviral system to over-express and knockdown expression during endochondral bone formation.

Results: In the limbs, PANX3 RNA was first detected in the cartilage condensations and became restricted to the prehypertrophic cartilage of the epiphyses, diaphysis, and perichondrium.

BMP signaling regulates the fate of chondro-osteoprogenitor cells in facial mesenchyme in a stage-specific manner.

Background: Lineage tracing has shown that most of the facial skeleton is derived from cranial neural crest cells. However, the local signals that influence postmigratory, neural crest-derived mesenchyme also play a major role in patterning the skeleton. Here, we study the role of BMP signaling in regulating the fate of chondro-osteoprogenitor cells in the face.

Identification and functional analysis of novel facial patterning genes in the duplicated beak chicken embryo.

Cranial neural crest cells form the majority of the facial skeleton. However exactly when the pattering information and hence jaw identity is established is not clear. We know that premigratory neural crest cells contain a limited amount of information about the lower jaw but the upper jaw and facial midline are specified later by local tissue interactions.

Analysis of human soft palate morphogenesis supports regional regulation of palatal fusion.

It is essential to complete palate closure at the correct time during fetal development, otherwise a serious malformation, cleft palate, will ensue. The steps in palate formation in humans take place between the 7th and 12th week and consist of outgrowth of palatal shelves from the paired maxillary prominences, reorientation of the shelves from vertical to horizontal, apposition of the medial surfaces, formation of a bilayered seam, degradation of the seam and bridging of mesenchyme. However, in the soft palate, the mechanism of closure is unclear.

Recent insights into the morphological diversity in the amniote primary and secondary palates.

The assembly of the upper jaw is a pivotal moment in the embryonic development of amniotes. The upper jaw forms from the fusion of the maxillary, medial nasal, and lateral nasal prominences, resulting in an intact upper lip/beak and nasal cavities; together called the primary palate. This process of fusion requires a balance of proper facial prominence shape and positioning to avoid craniofacial clefting, whilst still accommodating the vast phenotypic diversity of adult amniotes.

Diversity in primary palate ontogeny of amniotes revealed with 3D imaging.

The amniote primary palate encompasses the upper lip and the nasal cavities. During embryonic development, the primary palate forms from the fusion of the maxillary, medial nasal and lateral nasal prominences. In mammals, as the primary palate fuses, the nasal and oral cavities become completely separated.

Avian facial morphogenesis is regulated by c-Jun N-terminal kinase/planar cell polarity (JNK/PCP) wingless-related (WNT) signaling.

Wingless-related proteins (WNTs) regulate extension of the central axis of the vertebrate embryo (convergent extension) as well as morphogenesis of organs such as limbs and kidneys. Here, we asked whether WNT signaling directs facial morphogenesis using a targeted approach in chicken embryos. WNT11 is thought to mainly act via β-catenin-independent pathways, and little is known about its role in craniofacial development.