Ellis Van Creveld2 is Required for Postnatal Craniofacial Bone Development

Ellis‐van Creveld (EvC) syndrome is a genetic disorder with mutations in either EVC or EVC2 gene. Previous case studies reported that EvC patients underwent orthodontic treatment, suggesting the presence of craniofacial bone phenotypes. To investigate whether a mutation in EVC2 gene causes a craniofacial bone phenotype, Evc2 knockout (KO) mice were generated and cephalometric analysis was performed. The heads of wild type (WT), heterozygous (Het) and homozygous Evc2 KO mice (1‐, 3‐, and 6‐week‐old) were prepared and cephalometric analysis based on the selected reference points on lateral X‐ray radiographs was performed. The linear and angular bone measurements were then calculated, compared between WT, Het and KO and statistically analyzed at each time point. Our data showed that length of craniofacial bones in KO was significantly lowered by ～20% to that of WT and Het, the growth of certain bones, including nasal bone, palatal length, and premaxilla was more affected in KO, and the reduction in these bone length was more significantly enhanced at later postnatal time points (3 and 6 weeks) than early time point (1 week). Furthermore, bone‐to‐bone relationship to cranial base and cranial vault in KO was remarkably changed, i.e. cranial vault and nasal bone were depressed and premaxilla and mandible were developed in a more ventral direction. Our study was the first to show the cause‐effect relationship between Evc2 deficiency and craniofacial defects in EvC syndrome, demonstrating that Evc2 is required for craniofacial bone development and its deficiency leads to specific facial bone growth defect. Anat Rec, 299:1110–1120, 2016. © 2016 Wiley Periodicals, Inc.     

First Natural Endocranial Cast of a Fossil Snake (Cretaceous of Patagonia,Argentina)

In this study, we describe a natural endocranial cast included in a partially preserved medium‐sized skull of the Upper Cretaceous South American snake Dinilysia patagonica. The endocast is composed of sedimentary filling of the cranial cavity in which the posterior brain, the vessels, the cranial nerves, and the inner ear surrounded by delicate semicircular canals, are represented. It is simple in form, with little differentiation between the three main areas (Forebrain, Midbrain, and Hindbrain), and without flexures. The nervous system is well preserved. The posterior brain surface is smooth, except for two small prominences that make up the cerebellum. A large inner ear is preserved on the right side; it consists of a voluminous central mass, the vestibule, which occupies most of the space defined by the three semicircular canals. In particular, the lateral semicircular canal is very close to the vestibule. This characteristic, in combination with the medium to large body size of Dinilysia, its large skull and dorsally exposed orbits, and vertebrae bearing a rather high neural spine on a depressed neural arch, suggests that this snake would have had a semifossorial lifestyle. Anat Rec, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 301:9–20, 2018. © 2017 Wiley Periodicals, Inc.     

Notch pathway regulation of neural crest cell development in vivo

Background : The function of Notch signaling in murine neural crest–derived cell lineages in vivo was examined. Results : Conditional gain (Wnt1Cre;Rosa^Notch) or loss (Wnt1Cre;RBP‐J^f/f) of Notch signaling in neural crest cells (NCCs) in vivo results in craniofacial, cardiac, and trunk abnormalities. Severe craniofacial malformations are apparent in Wnt1Cre;Rosa^Notch embryos, while less severe skull abnormalities are evident in Wnt1Cre;RBP‐J^f/f mice. Deficient cardiac neural crest migration, resulting in cardiac outflow tract malformations, occurs with increased or decreased Notch signaling in NCCs. Smooth muscle cell differentiation also is impaired in pharyngeal NCC derivatives in both Wnt1Cre;Rosa^Notch and Wnt1Cre;RBP‐J^f/f embryos. Neurogenesis is absent and gliogenesis is increased in the dorsal root ganglia of Wnt1Cre;Rosa^Notch embryos, while neurogenesis is increased and gliogenesis is decreased in Wnt1Cre;RBP‐J^f/f embryos. Conclusions : Together, these studies demonstrate essential cell‐autonomous roles for appropriate levels of Notch signaling during NCC migration, proliferation, and differentiation with critical implications in craniofacial, cardiac, and neurogenic development and disease. Developmental Dynamics 241:376–389, 2012. © 2011 Wiley Periodicals, Inc.     

Skull Anatomy and Ontogeny of Legless Lizard Pseudopus apodus (Pallas, 1775): Heterochronic Influences on Form

Pseudopus apodus (Pallas, 1775) is the largest extant legless species of the subfamily Anguinae (Anguimorpha, Anguidae) living mostly in the sub‐arid territories ranging from the Balkan area in Europe to Kazakhstan in Asia. The species of other two genera live in North America, South‐East Asia and North Africa (Ophisaurus) and Europe and South‐West Asia (Anguis). The interrelationships of Anguinae are unresolved; this is in part the consequence of the insufficient knowledge of the cranial, postcranial and integumentary anatomy of the individual anguine species. The aim of this article is to fulfill this gap in our knowledge of the anguine anatomy. Now, in the first part of the project, the individual bones of the exocranium and visceral endocranium of the anguine legless lizard P. apodus are described in detail. In the present study, P. apodus is revealed to have autoapomorphic features of the skull which clearly distinguish it from Anguis and Ophisaurus. In addition, the study of posthatchling ontogeny of exocranium of P. apodus revealed some features, such as a nasal process of premaxilla being slightly widened in about its mid‐length, that are also typical for adults of the Ophisaurus and Anguis species as well as extinct species of Pseudopus. This strongly indicates that peramorphic heterochronic process played role in the evolution of the P. apodus skull. Anat Rec, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 300:460–502, 2017. © 2016 Wiley Periodicals, Inc.     

Morphometric Variations in the Skin Layers of Frogs: An Exploration Into Their Relation With Ecological Parameters in Leptodactylus (Anura,Leptodactylidae), With an Emphasis on the Eberth‐Kastschenko Layer

Leptodactylus is a genus of frogs known to live in diverse habitats and to show both aquatic and terrestrial breeding habits. We studied 21 species of Leptodactylus to explore whether skin structure specialization relates to habitats and habit variation. Morphometric analyses of the skin thickness revealed that phylogeny has a strong influence on variations in the thickness of the epidermis, stratum spongiosum, Eberth‐Kastschenko layer, and stratum compactum, while habitat and habits display no significant correlation. The optimization of the phylogenetic hypothesis suggested that a pattern of intermediate values for skin layer thickness are plesiomorphic for this group. Anat Rec, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 300:1895–1909, 2017. © 2017 Wiley Periodicals, Inc.     

Pten regulates neural crest proliferation and differentiation during mouse craniofacial development

Background: The phosphatase and tensin homolog deleted on chromosome TEN (Pten) is implicated in a broad range of developmental events and diseases. However, its role in neural crest and craniofacial development has not been well illustrated. Results: Using genetically engineered mouse models, we showed that inactivating Pten specifically in neural crest cells causes malformation of craniofacial structures. Pten conditional knockout mice exhibit perinatal lethality with overgrowth of craniofacial structures. At the cellular level, Pten deficiency increases cell proliferation rate and enhances osteoblast differentiation. Our data further revealed that inactivating Pten elevates PI3K/Akt signaling activity in neural crest derivatives, and confirmed that attenuation of PI3K/Akt activity led to decreased neural crest cell proliferation and differentiation both in vitro and in vivo. Conclusions: Our study revealed that Pten is essential for craniofacial morphogenesis in mice. Inactivating Pten in neural crest cells increases proliferation rate and promotes their differentiation toward osteoblasts. Our data further indicate that Pten acts via modulating PI3K/Akt activity during these processes. Developmental Dynamics 247:304–314, 2018. © 2017 Wiley Periodicals, Inc.     

Cranial Ontogeny of the Early Triassic Basal Cynodont Galesaurus planiceps

Ontogenetic changes in the skull and mandible of thirty‐one specimens of Galesaurus planiceps, a basal non‐mammaliaform cynodont from the Early Triassic of South Africa, are documented. The qualitative survey indicated eight changes in the craniomandibular apparatus occurred during growth, dividing the sample into three ontogenetic stages: juvenile, subadult, and adult. Changes in the temporal region, zygomatic arch, occiput, and mandible occurred during the transition from the subadult to adult stage at a basal skull length of 90 mm. At least four morphological and allometric differences divided the adult specimens into two morphs, indicating the presence of sexual dimorphism in Galesaurus. Differences include extensive lateral flaring of the zygomatic arches in the “male” morph resulting in a more anterior orientation of the orbits, and a narrower snout in the “female”. This is the first record of sexual dimorphism in a basal cynodont, and the first time it is quantitatively documented in a non‐mammaliaform cynodont. An ontogenetic comparison between Galesaurus and the more derived basal cynodont Thrinaxodon revealed differences in the timing and extent of sagittal crest development. In Galesaurus, the posterior sagittal crest, located behind the parietal foramen, developed relatively later in ontogeny, and the anterior sagittal crest rarely formed suggesting the anterior fibres of the temporalis were less developed than in Thrinaxodon. In contrast, craniomandibular features related to the masseters became more developed during the ontogeny of Galesaurus. The development of the adductor musculature appears to be one of the main factors influencing skull growth in these basal non‐mammaliaform cynodonts. Anat Rec, 300:353–381, 2017. © 2016 Wiley Periodicals, Inc.     

Craniosynostosis in Twist heterozygous mice: A model for Saethre‐Chotzen syndrome

Saethre‐Chotzen syndrome is a common autosomal dominant form of craniosynostosis, the premature fusion of the sutures of the calvarial bones of the skull. Most Saethre‐Chotzen syndrome cases are caused by haploinsufficiency for the TWIST gene. Mice heterozygous for a null mutation of the Twist gene replicate certain features of Saethre‐Chotzen syndrome, but have not been reported to exhibit craniosynostosis. We demonstrate that Twist heterozygous mice exhibit fusions of the coronal suture and other cranial suture abnormalities, indicating that Twist heterozygous mice constitute a better animal model for Saethre‐Chotzen syndrome than was previously appreciated. Anat Rec 268:90–92, 2002. © 2002 Wiley‐Liss, Inc.     

The First Record of Incidences of Ankylosis in Seven Triplefin Fishes (Pisces: Tripterygiidae) from New Zealand

Different cases of vertebral ankylosis were examined in seven tripterygiid species obtained from waters around New Zealand. The skeletal deformities observed are located in the caudal region of the vertebral column. Those occurred in Forsterygion nigripenne, Matanui bathytaton and in one specimen of Ruanoho whero were severe cases. The mechanism of the formation of vertebral ankylosis and the causes behind such anomaly were discussed. Further studies are needed to relate specific pollutants with the observed types of deformities. Anat Rec, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 301:39–45, 2018. © 2017 Wiley Periodicals, Inc.     

Effects of Antitumor Drug Sorafenib on Chick Embryo Development

Sorafenib has been used as an oral anti‐cancer drug because of its ability to inhibit tumor growth. However, the pharmacological effect of sorafenib is still the lack of in vivo experimental evidence. Tumor and embryonic cells share some similar features, so we investigated the effects of sorafenib on the development of gastrulating chick embryos. We found that sorafenib exposure was markedly attributed to the number of embryonic cell in proliferation and apoptosis. We also detected sorafenib significantly interfered with epithelial‐mesenchymal transition (EMT). Furthermore, sorafenib treatment impaired the production and migration of neural crest cells. Anat Rec, 298:1271–1281, 2015. © 2015 Wiley Periodicals, Inc.     

Escapees from Rhombomeric Lineage Restriction: Extensive Migration Rostral to the r4/r5 Border of Hox‐a3 Expression

The rhombomeric compartments of the hindbrain are characterized by lineage restriction; cells born in one compartment generally remain there and do not migrate to neighboring rhombomeres. Two well‐known exceptions are the substantial migrations of the pontine nuclei and the mammalian facial nucleus. In this study we used Hoxa3‐Cre lineage to permanently mark cells that originate in rhombomeres caudal to r4. We found that cells born caudal to the r4/r5 border migrate forwards to a number of different locations in rhombomeres 1–4; the final locations include the interfascicular trigeminal nucleus, the principal trigeminal nucleus, the pontine nuclei, the reticulotegmental nucleus, the ventral nucleus of the lateral lemniscus, and the lateral and medial vestibular nuclei. We suggest that there are numerous exceptions to the principle of rhombomeric lineage restriction that have previously gone unnoticed. Anat Rec, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 300:1838–1846, 2017. © 2017 Wiley Periodicals, Inc.     

Pdgfra and Pdgfrb genetically interact during craniofacial development

Background: One of the most prevalent congenital birth defects is cleft palate. The palatal skeleton is derived from the cranial neural crest and platelet‐derived growth factors (Pdgf) are critical in palatogenesis. Of the two Pdgf receptors, pdgfra is required for neural crest migration and palatogenesis. However, the role pdgfrb plays in the neural crest, or whether pdgfra and pdgfrb interact during palatogenesis is unclear. Results: We find that pdgfrb is dispensable for craniofacial development in zebrafish. However, the palatal defect in pdgfra;pdgfrb double mutants is significantly more severe than in pdgfra single mutants. Data in mouse suggest this interaction is conserved and that neural crest requires both genes. In zebrafish, pdgfra and pdgfrb are both expressed by neural crest within the pharyngeal arches, and pharmacological analyses demonstrate Pdgf signaling is required at these times. While neither proliferation nor cell death appears affected, time‐lapsed confocal analysis of pdgfra;pdgfrb mutants shows a failure of proper neural crest condensation during palatogenesis. Conclusions: We provide data showing that pdgfra and pdgfrb interact during palatogenesis in both zebrafish and mouse. In zebrafish, this interaction affects proper condensation of maxillary neural crest cells, revealing a previously unknown interaction between Pdgfra and Pdgfrb during palate formation. Developmental Dynamics 245:641–652, 2016. © 2016 Wiley Periodicals, Inc.     

The Relationship Between Linear Osteological and Radiographic Measurements of the Human Calcaneus and Talus

Plain radiographs of the foot are a common form of examination in individuals when medical evaluation is indicated. Their availability makes them useful for studying human variation where large sample sizes are crucial. Calcaneal and talar morphology are critical in the understanding of human foot form and function, but few studies have examined the accuracy of linear measurements of these bones taken from radiographs. If linear measurements are to be used in quantitative analysis, their accuracy must first be demonstrated. For this study, 54 human feet from amputated limbs were imaged in standard radiographic views and skeletonized. Selected linear measurements were made on each skeletonized talus and calcaneus. Corresponding landmarks were identified on the radiographic images and the distances between these points determined. Using the measurements taken from the physical bone, the accuracy of the radiographic measurements was evaluated employing paired Student's t‐tests and correlation analysis. Most measurements were not significantly different (P ≥ 0.05). Overall, the sagittal plane measurements were correlated (r² > 0.61–0.88, all P < 0.001) with the radiographic measurements; transverse plane measurements less so (r² = 0.20–0.63, P < 0.001). These results demonstrate that measurements from radiographic images provide useful information concerning both the calcaneus and talus, indicating that radiographs of living individuals can be evaluated quantitatively and compared to osteology collections of modern humans. This information can be applied to the understanding of functional anatomy of the modern human foot, interpreting the hominin fossil pedal record, and establishing identification in forensic analysis. Anat Rec, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 301:21–33, 2018. © 2017 Wiley Periodicals, Inc.     

2D or Not 2D? Testing the Utility of 2D Vs. 3D Landmark Data in Geometric Morphometrics of the Sculpin Subfamily Oligocottinae (Pisces; Cottoidea)

We contrast 2D vs. 3D landmark‐based geometric morphometrics in the fish subfamily Oligocottinae by using 3D landmarks from CT‐generated models and comparing the morphospace of the 3D landmarks to one based on 2D landmarks from images. The 2D and 3D shape variables capture common patterns across taxa, such that the pairwise Procrustes distances among taxa correspond and the trends captured by principal component analysis are similar in the xy plane. We use the two sets of landmarks to test several ecomorphological hypotheses from the literature. Both 2D and 3D data reject the hypothesis that head shape correlates significantly with the depth at which a species is commonly found. However, in taxa where shape variation in the z‐axis is high, the 2D shape variables show sufficiently strong distortion to influence the outcome of the hypothesis tests regarding the relationship between mouth size and feeding ecology. Only the 3D data support previous studies which showed that large mouth sizes correlate positively with high percentages of elusive prey in the diet. When used to test for morphological divergence, 3D data show no evidence of divergence, while 2D data show that one clade of oligocottines has diverged from all others. This clade shows the greatest degree of z‐axis body depth within Oligocottinae, and we conclude that the inability of the 2D approach to capture this lateral body depth causes the incongruence between 2D and 3D analyses. Anat Rec, 301:806–818, 2018. © 2017 Wiley Periodicals, Inc.     

Size and shape changes during late fetal growth (137–157 gestational days) in the pigtailed macaque (Macaca nemestrina) craniofacial complex: An application using three‐dimensional coordinate data and finite element scaling analysis

Form changes within the fetal pigtailed macaque (Macaca nemestrina) craniofacial complex was documented using finite element scaling analysis (FESA) and three‐dimensional (3D) coordinate data for 35 craniofacial landmarks. Coordinate data were digitized from 3D reconstructions of computed tomography (CT) images and 2D axial slices. Twenty‐two fetal pigtailed macaques ranging in age from 137 to 157 gestational days were included (in this species, birth is estimated at 170 gestational days). The null hypothesis that the craniofacial complex grows with isometry during late fetal growth of the craniofacial complex was tested (P < 0.05), and the prediction that morphological change along an anteroposterior axis dominates late fetal growth was also investigated. The null hypothesis was rejected, indicating that allometric growth is present during late fetal growth. Growth along an anteroposterior axis is localized in the palate and mandible. The neurocranium grows along a superoinferior axis, while the neurofacial junction displays growth along both the anteroposterior and superoinferior axes. Mediolateral changes are localized between asterions, the external auditory meati, and maxillary and mandibular alveolar points. Finally, a 3D model of craniofacial growth for this species was created, localizing size and shape changes that occur during late fetal growth for each of the 35 craniofacial landmarks defined in this study. Anat Rec 267:307–320, 2002. © 2002 Wiley‐Liss, Inc.     

Natural development of dermal ectopic bone in the american alligator (Alligator mississippiensis) resembles heterotopic ossification disorders in humans

Heterotopic ossification (HO) occurs when soft tissues are inappropriately converted to bony tissue. Several human diseases result in HO with few reliable treatment options. Animal models that naturally produce dermal ectopic bone (i.e., osteoderms), such as crocodilians, have never been utilized as models for studying these disorders in humans. Here, a histological evaluation and staging criteria for osteoderm development is described for the first time in the American alligator (Alligator mississipiensis). Differential staining and immunohistochemistry of alligator scales depict a progressive change during development, where woven bone forms from the differentiated dermis. Bone formation proceeds via intramembranous ossification, which is initiated in part by endothelial cell precursors that undergo endothelial‐to‐mesenchymal transition and eventually acquire an osteoblast phenotype. As such, the development of osteoderms in the American alligator bears morphological and mechanistic similarities to HO in humans, presenting a potential model for future study of soft tissue mineralization pathologies and providing insight into the morphological and molecular development of osteoderms in other vertebrate lineages. Anat Rec, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 301:56–76, 2018. © 2017 Wiley Periodicals, Inc.