Anti-asthma Drugs Formoterol and Budesonide (Symbicort) Induce Orofacial Clefts, Gastroschisis and Heart Septum Defects in an In Vivo Model.

BACKGROUND: We had a case in which three consecutive pregnancies resulted in birth of three children with an orofacial cleft. Their mother suffered from bronchial asthma and was treated using symbicort (corticosteroid budesonide plus bronchodilator formoterol) during her pregnancies. A hypothesis was assessed: these anti-asthmatics can induce an orofacial cleft in experimental model. MATERIALS AND METHODS: A single administration of one of five increasing doses (including therapeutically used ones) of Symbicort, budesonide or formoterol was injected into the amnion of a chick embryo on day 4 or 5 of incubation. The teratogenic/lethal effects of the anti-asthmatics were assessed on a total of 600 embryos. RESULTS: For budesonide, the teratogenic/lethal effect started at a dose 0.003 µg per embryo, for formoterol at 0.3 µg and for Symbicort 0.03 µg. Orofacial clefts and gastroschisis after exposure were found for all three anti-asthmatics. Heart septum defects occurred after exposure to formoterol. CONCLUSION: The present results support those clinical/epidemiological studies pointing out that anti-asthmatics have the potential to induce orofacial clefts, gastroschisis and heart malformations during prenatal development in human.

Correction: Targeting of the Plzf Gene in the Rat by Transcription Activator-Like Effector Nuclease Results in Caudal Regression Syndrome in Spontaneously Hypertensive Rats.

[This corrects the article DOI: 10.1371/journal.pone.0164206.].

Dentoalveolar Arch Dimensions in UCLP Boys After Neonatal Cheiloplasty or After Lip Surgery at the Age of 3 or 6 Months.

OBJECTIVE: To compare the influence of 3 different time protocols of cleft lip and palate operations on the growth of the dentoalveolar arch in patients with unilateral cleft lip and palate (UCLP). MATERIALS AND METHODS: We evaluated 64 plaster casts of 8-year-old boys with UCLP operated on according to 3 different time protocols: lip repair at the age of 6 months and palate repair at 4 years, lip repair at 3 months and palate repair at 9 months, and neonatal lip repair and palate repair at 9 months. The control group contained 13 plaster casts of 8-year-old boys. The dentoalveolar arch width was measured between deciduous canines and between the second deciduous molars; the length was measured between incisive papilla and the line connecting both tuber maxillae. RESULTS: All measured distances were statistically significantly smaller in boys with UCLP than in the control group. Intercanine width was not statistically significantly different between the patients operated on according to the different time protocols. In comparison to the lip repair at 6 months and palate repair at 4 years, the intermolar width was statistically significantly smaller in the group with neonatal lip repair; the alveolar arch length was statistically significantly shorter in both groups with lip repair performed neonatally or at 3 months. CONCLUSIONS: The length of the dentoalveolar arch is shorter after surgical repair of cleft lip neonatally or at the age of 3 months. Cleft palate repair at 9 months can contribute to a reduction in the width of the dentoalveolar arch.

Long-term significant seasonal differences in the numbers of new-borns with an orofacial cleft in the Czech Republic - a retrospective study.

BACKGROUND: Choosing the optimal season for conception is a part of family planning since it can positively influence the pregnancy outcome. Changes in the monthly number of infants born with a birth defect can signal prenatal damage - death or malformation - related to a harmful seasonal factor. The aim of our paper was to search for possible seasonal differences in the numbers of new-borns with an orofacial cleft and thus for a period of conception that can increase the risk of orofacial cleft development. METHODS: Mean monthly numbers of live births in the Bohemia region of the Czech Republic during the years 1964-2000 were compared within a group of 5619 new-borns with various types of orofacial clefts and the control group derived from natality data on 3,080,891 new-borns. RESULTS: The control group exhibited regular seasonal variation in the monthly numbers of new-borns: significantly more babies born during March-May and fewer babies born during October-December. Similar natural seasonal variation was also found in the group of babies with an orofacial cleft. However, after subdividing the cleft group according to gender and cleft type, in comparison to controls, significant differences appeared in the number of new-born girls with cleft lip during January-March and in the number of boys born with cleft palate in April - May. CONCLUSIONS: We found significant differences from controls in the number of new-born girls with CL and boys with CP, whose dates of birth correspond to conception from April to August and to the estimated prenatal critical period for cleft formation from May to October. The latter period includes the warm season, when various injurious physical, chemical and biological factors may act on a pregnant woman. This finding should be considered in pregnancy planning. Future studies are necessary to investigate the putative injurious factors during the warm season that can influence pregnancy outcome.

Early development of the human dentition revisited.

In this review, classical data on the early steps in human odontogenesis are summarized and updated with specific insights into the development of the upper and lower embryonic jaws to help in understanding some oral pathologies. The initial step of human odontogenesis is classically characterized by two parallel horseshoe-shaped epithelial laminae. These originate from the oral epithelium and an ingrowth into the jaw mesenchyme: the internal dental lamina gives rise to deciduous tooth primordia, while the external vestibular lamina represents the developmental base of the oral vestibule. However, a more complex situation was revealed by recent studies combining analyses of the dental and adjacent oral epithelia on histological sections and computer-aided three-dimensional (3D) reconstructions during the 2nd month of human embryonic development. The dental epithelium forms a mound, where swellings appear later, corresponding to the individual primordia of deciduous teeth. External to the developing deciduous dentition, the 3D reconstructions do not show any continuous vestibular lamina but instead a complex of discontinuous epithelial bulges and ridges. The patterns of these epithelial structures and their relationship to the dental epithelium differ not only between the upper and lower jaws but also between the lip and cheek segments in each jaw. Knowledge of early odontogenesis may help in understanding some oral pathologies. For example, the human lateral incisor has a dual origin: it arises in the area of fusion between the medial nasal and maxillary facial processes and involves material from these two regions. Such a dual origin at the site of fusion of facial processes represents a predisposition to developmental vulnerability for the upper lateral incisor, resulting in its frequent anomalies (absence, hypoplasia, duplication), especially in patients with a cleft lip and/or jaw. Other pathologies, such as a minute supernumerary tooth, desmoplastic ameloblastoma or extraosseous odontogenic cysts are located external to the upper or lower dentition, and might be derived from structures that transiently appear during early development of the oral vestibule in humans.

Targeting of the Plzf Gene in the Rat by Transcription Activator-Like Effector Nuclease Results in Caudal Regression Syndrome in Spontaneously Hypertensive Rats.

Recently, it has been found that spontaneous mutation Lx (polydactyly-luxate syndrome) in the rat is determined by deletion of a conserved intronic sequence of the Plzf (Promyelocytic leukemia zinc finger protein) gene. In addition, Plzf is a prominent candidate gene for quantitative trait loci (QTLs) associated with cardiac hypertrophy and fibrosis in the spontaneously hypertensive rat (SHR). In the current study, we tested the effects of Plzf gene targeting in the SHR using TALENs (transcription activator-like effector nucleases). SHR ova were microinjected with constructs pTAL438/439 coding for a sequence-specific endonuclease that binds to target sequence in the first coding exon of the Plzf gene. Out of 43 animals born after microinjection, we detected a single male founder. Sequence analysis revealed a deletion of G that resulted in frame shift mutation starting in codon 31 and causing a premature stop codon at position of amino acid 58. The Plzftm1Ipcv allele is semi-lethal since approximately 95% of newborn homozygous animals died perinatally. All homozygous animals exhibited manifestations of a caudal regression syndrome including tail anomalies and serious size reduction and deformities of long bones, and oligo- or polydactyly on the hindlimbs. The heterozygous animals only exhibited the tail anomalies. Impaired development of the urinary tract was also revealed: one homozygous and one heterozygous rat exhibited a vesico-ureteric reflux with enormous dilatation of ureters and renal pelvis. In the homozygote, this was combined with a hypoplastic kidney. These results provide evidence for the important role of Plzf gene during development of the caudal part of a body-column vertebrae, hindlimbs and urinary system in the rat.

3-D shape analysis of palatal surface in patients with unilateral complete cleft lip and palate.

Facial development of patients with unilateral complete cleft lip and palate (UCLP) is associated with many problems including deformity of the palate. The aim of this study was to evaluate palatal morphology and variability in patients with UCLP compared with Czech norms using methods of geometric morphometrics. The study was based on virtual dental cast analysis of 29 UCLP patients and 29 control individuals at the age of 15 years. The variability of palatal shape in UCLP patients was greater than that in nonclefted palates. Only 24% of clefted palates fell within the variability of controls. The palatal form of UCLP patients (range from 11.8 to 17.2 years) was not correlated with age. Compared with control palates, palates of UCLP patients were narrower, more anteriorly than posteriorly. Apart from the praemaxilla region, they were also shallower, and the difference increased posteriorly. The UCLP palate was characterised by the asymmetry of its vault. The maximum height of the palatal vault was anterior on the clefted side, whereas it was posterior on the nonclefted side. The slope of the UCLP palate was more inclined compared with the control group. The praemaxilla was therefore situated more inferiorly.

Developmental disorders of the dentition: an update.

Dental anomalies are common congenital malformations that can occur either as isolated findings or as part of a syndrome. This review focuses on genetic causes of abnormal tooth development and the implications of these abnormalities for clinical care. As an introduction, we describe general insights into the genetics of tooth development obtained from mouse and zebrafish models. This is followed by a discussion of isolated as well as syndromic tooth agenesis, including Van der Woude syndrome (VWS), ectodermal dysplasias (EDs), oral-facial-digital (OFD) syndrome type I, Rieger syndrome, holoprosencephaly, and tooth anomalies associated with cleft lip and palate. Next, we review delayed formation and eruption of teeth, as well as abnormalities in tooth size, shape, and form. Finally, isolated and syndromic causes of supernumerary teeth are considered, including cleidocranial dysplasia and Gardner syndrome.

Sequential Shh expression in the development of the mouse upper functional incisor.

The mouse incisor is a frequently used model in studies of the molecular control of organ development. The appropriate interpretation of data on normogenesis is essential for understanding the data obtained in mutant mice. For this reason, we performed a very detailed investigation of the development of the upper incisor in wild-type mice from embryonic day (ED) 11.5 till 14.5. A combination of histology, whole mount in situ hybridization, computer-aided three-dimensional reconstructions, and fluorescent microscopy, has been used. Several sonic hedgehog (Shh) expression domains have been detected in the upper incisor region during early prenatal development. At ED11.5-13.5, there was a single Shh positive domain present in the anterior part of left or right upper jaw arches, corresponding to the epithelial thickening. More posteriorly, a new Shh expression domain appeared in the incisor bud in the developmentally more advanced ED13.5 embryos. At ED14.5, only this posterior Shh expression in the incisor germ remained detectable. This study brings new insights into the early development of the upper incisor in mice and completes the data on normal mouse incisor development. The temporal-spatial pattern of Shh expression reflects the development of two tooth generations, being detectable in two successive, antero-posteriorly located areas in the prospective incisor region in the upper jaw. The first, anterior and superficial Shh expression domain reflects the rudimentary tooth development suppressed during evolution. Only the subsequent, posterior and deeper Shh expression region, appearing at ED13.5, correlates with the prospective upper functional incisor in wild-type mice.

The dynamics of supernumerary tooth development are differentially regulated by Sprouty genes.

In mice, a toothless diastema separates the single incisor from the three molars in each dental quadrant. In the prospective diastema of the embryo, small rudimentary buds are found that are presumed to be rudiments of suppressed teeth. A supernumerary tooth occurs in the diastema of adult mice carrying mutations in either Spry2 or Spry4. In the case of Spry2 mutants, the origin of the supernumerary tooth involves the revitalization of a rudimentary tooth bud (called R2), whereas its origin in the Spry4 mutants is not known. In addition to R2, another rudimentary primordium (called MS) arises more anteriorly in the prospective diastema. We investigated the participation of both rudiments (MS and R2) in supernumerary tooth development in Spry2 and Spry4 mutants by comparing morphogenesis, proliferation, apoptosis, size and Shh expression in the dental epithelium of MS and R2 rudiments. Increased proliferation and decreased apoptosis were found in MS and R2 at embryonic day (ED) 12.5 and 13.5 in Spry2(-/-) embryos. Apoptosis was also decreased in both rudiments in Spry4(-/-) embryos, but the proliferation was lower (similar to WT mice), and supernumerary tooth development was accelerated, exhibiting a cap stage by ED13.5. Compared to Spry2(-/-) mice, a high number of Spry4(-/-) supernumerary tooth primordia degenerated after ED13.5, resulting in a low percentage of supernumerary teeth in adults. We propose that Sprouty genes were implicated during evolution in reduction of the cheek teeth in Muridae, and their deletion can reveal ancestral stages of murine dental evolution.

Variability in palatal shape and size in patients with bilateral complete cleft lip and palate assessed using dense surface model construction and 3D geometric morphometrics.

Bilateral complete cleft lip and palate (BCLP) is the most severe of the common orofacial clefts and is associated with the greatest deformity during development. The aim of this study was to use geometric morphometrics to evaluate palatal shape and size variability in patients with BCLP in comparison to nonclefted Czech boys. The variability of palatal size and shape in BCLP patients was greater when compared with the nonclefted population. Though palate variability in BCLP was wide, nonclefted palatal shapes were generally different (a high, wide and vaulted palate) and fell almost outside the range of BCLP variability. The palatal size and shape of BCLP patients (range from 12.1 to 16.5 years) was not correlated with age. A comparison of the mean shapes of the clefted and nonclefted groups showed that the BCLP palate is flatter and narrower. The most notable size difference was found in the area between the maxilla and premaxilla. This phenomenon is associated with the persisting separation of the premaxilla from the rest of the palate. The shape of the palatal configuration of the premaxilla and adjacent area was concave in the nonclefted group and convex in BCLP patients.

Shh expression in a rudimentary tooth offers new insights into development of the mouse incisor.

For teeth as for any organ, knowledge of normal development is essential for the proper interpretation of developmental anomalies in mutant mice. It is generally accepted that tooth formation is initiated with a single signaling center that, in the incisor region, is exclusively related to the development of the functional adult incisor. Here, using a unique combination of computer-aided three-dimensional reconstructions and whole mount in situ hybridization of mandibles from finely staged wild-type mouse embryos, we demonstrate that several Sonic hedgehog (Shh) expression domains sequentially appear in the lower incisor region during early development. In contrast to the single Shh expression domain that is widely assumed to be present in each lower incisor area at ED12.5-13.5, we identified two spatially distinct regions of Shh expression that appear in an anterior-posterior sequence during this period. The initial anterior, more superficially located Shh expression region represented the rudimentary (so-called deciduous) incisor, whereas only the later posterior deeper situated region corresponded to the prospective functional incisor. In the more advanced embryos, only this posterior Shh expression in the incisor bud was detectable as a precursor of the enamel knot. This study offers a new interpretation of published molecular data on the mouse incisor from initiation through ED13.5. We suggest that, as with Shh expression, other molecular data that have been ascribed to the progressive development of the mouse functional incisor at early stages, in fact, correspond to a rudimentary incisor whose development is aborted.

Patterning by heritage in mouse molar row development.

It is known from paleontology studies that two premolars have been lost during mouse evolution. During mouse mandible development, two bud-like structures transiently form that may represent rudimentary precursors of the lost premolars. However, the interpretation of these structures and their significance for mouse molar development are highly controversial because of a lack of molecular data. Here, we searched for typical tooth signaling centers in these two bud-like structures, and followed their fate using molecular markers, 3D reconstructions, and lineage tracing in vitro. Transient signaling centers were indeed found to be located at the tips of both the anterior and posterior rudimentary buds. These centers expressed a similar set of molecular markers as the "primary enamel knot" (pEK), the signaling center of the first molar (M1). These two transient signaling centers were sequentially patterned before and anterior to the M1 pEK. We also determined the dynamics of the M1 pEK, which, slightly later during development, spread up to the field formerly occupied by the posterior transient signaling center. It can be concluded that two rudimentary tooth buds initiate the sequential development of the mouse molars and these have previously been mistaken for early stages of M1 development. Although neither rudiment progresses to form an adult tooth, the posterior one merges with the adjacent M1, which may explain the anterior enlargement of the M1 during mouse family evolution. This study highlights how rudiments of lost structures can stay integrated and participate in morphogenesis of functional organs and help in understanding their evolution, as Darwin suspected long ago.

Prenatal development of Crocodylus niloticus niloticus Laurenti, 1768.

Prenatal development in crocodilians represents a very interesting model for comparative studies. As the speed of prenatal development of crocodilians varies depending on incubation conditions, the staging of embryos and fetuses is a very important prerequisite for data correlation. To establish a background for future developmental studies on Crocodylus niloticus, we characterized its prenatal development in a collection comprising 169 animals during embryonic/incubation days 9-70. The characteristics included external morphology, head morphometry, and wet body weight determined before fixation. We documented the external morphology of prenatal Nile crocodiles in a large collection of photographs and described landmarks during the morphogenesis of the head, face and limbs. In the development of the facial processes (medial nasal, lateral nasal, maxillary), three phases could be distinguished: union, separation, reunion. At the free jaw margin, a regular series of prominences was present. The outer aspect of a prominence gave rise to a labial scale, the inner aspect to a tooth. In contrast to mammals (humans and mice), the hindlimbs of C. niloticus developed faster than the forelimbs. We also determined changes in basic measures of the head and of the wet body weight. Both morphological and morphometric characteristics showed an apparent inter-individual variability among animals of the same age. This variability decreased among animals of a similar body weight (irrespective of their age). Body weight can be considered as the most representative and complex parameter for crocodile staging reflecting the overall growth of a whole embryo/fetus.

Revitalization of a diastemal tooth primordium in Spry2 null mice results from increased proliferation and decreased apoptosis.

An understanding of the factors that promote or inhibit tooth development is essential for designing biological tooth replacements. The embryonic mouse dentition provides an ideal system for studying such factors because it consists of two types of tooth primordia. One type of primordium will go on to form a functional tooth, whereas the other initiates development but arrests at or before the bud stage. This developmental arrest contributes to the formation of the toothless mouse diastema. It is accompanied by the apoptosis of the rudimentary diastemal buds, which presumably results from the insufficient activity of anti-apoptotic signals such as fibroblast growth factors (FGFs). We have previously shown that the arrest of a rudimentary tooth bud can be rescued by inactivating Spry2, an antagonist of FGF signaling. Here, we studied the role of the epithelial cell death and proliferation in this process by comparing the development of a rudimentary diastemal tooth bud (R(2)) and the first molar in the mandibles of Spry2(-/-) and wild-type (WT) embryos using histological sections, image analysis and 3D reconstructions. In the WT R(2) at embryonic day 13.5, significantly increased apoptosis and decreased proliferation were found compared with the first molar. In contrast, increased levels of FGF signaling in Spry2(-/-) embryos led to significantly decreased apoptosis and increased proliferation in the R(2) bud. Consequently, the R(2) was involved in the formation of a supernumerary tooth primordium. Studies of the revitalization of rudimentary tooth primordia in mutant mice can help to lay the foundation for tooth regeneration by enhancing our knowledge of mechanisms that regulate tooth formation.

Temporal analysis of ectopic enamel production in incisors from sprouty mutant mice.

The mouse incisor has two unusual features: it grows continuously and it is covered by enamel exclusively on the labial side. The continuous growth is driven in part by epithelial stem cells in the cervical loop region that can both self-renew and give rise to ameloblasts. We have previously reported that ectopic enamel is found on the lingual side of the incisor in mice with loss-of-function of sprouty (spry) genes. Spry2(+/-); Spry4(-/-) mice, in which three sprouty alleles have been inactivated, have ectopic enamel as a result of upregulation of epithelial-mesenchymal FGF signaling in the lingual part of the cervical loop. Interestingly, lingual enamel is also present in the early postnatal period in Spry4(-/-) mice, in which only two sprouty alleles have been inactivated, but ectopic enamel is not found in adults of this genotype. To explore the mechanisms underlying the disappearance of lingual enamel in Spry4(-/-) adults, we studied the fate of the lingual enamel in Spry4(-/-) mice by comparing the morphology and growth of their lower incisors with wild type and Spry2(+/-); Spry4(-/-) mice at several timepoints between the perinatal period and adulthood. Ameloblasts and enamel were detected on the lingual side in postnatal Spry2(+/-); Spry4(+/-) incisors. By contrast, new ectopic ameloblasts ceased to differentiate after postnatal day 3 in Spry4(-/-) incisors, which was followed by a progressive loss of lingual enamel. Both the posterior extent of lingual enamel and the time of its last deposition were variable early postnatally in Spry4(-/-) incisors, but in all Spry4(-/-) adult incisors the lingual enamel was ultimately lost through continuous growth and abrasion of the incisor.

An FGF signaling loop sustains the generation of differentiated progeny from stem cells in mouse incisors.

Rodent incisors grow throughout adult life, but are prevented from becoming excessively long by constant abrasion, which is facilitated by the absence of enamel on one side of the incisor. Here we report that loss-of-function of sprouty genes, which encode antagonists of receptor tyrosine kinase signaling, leads to bilateral enamel deposition, thus impeding incisor abrasion and resulting in unchecked tooth elongation. We demonstrate that sprouty genes function to ensure that enamel-producing ameloblasts are generated on only one side of the tooth by inhibiting the formation of ectopic ameloblasts from self-renewing stem cells, and that they do so by preventing the establishment of an epithelial-mesenchymal FGF signaling loop. Interestingly, although inactivation of Spry4 alone initiates ectopic ameloblast formation in the embryo, the dosage of another sprouty gene must also be reduced to sustain it after birth. These data reveal that the generation of differentiated progeny from a particular stem cell population can be differently regulated in the embryo and adult.

Chernobyl: relationship between the number of missing newborn boys and the level of radiation in the Czech regions.

BACKGROUND: The number of newborn boys was higher than that of girls in the Czech Republic each month from 1950 to 2005. The only exception was November 1986, when the number of newborn boys was significantly reduced. This has been explained by a selective negative impact of the Chernobyl accident in April 1986 on male fetuses during the third month of their prenatal development. OBJECTIVES: The first and most radioactive cloud passed over the Czech Republic during 30 April-1 May 1986. Concurrent rainfall multiplied the radioactivity by up to > 10,000-fold in specific regions. We verified a hypothesis that the decrease in the male birth fraction in November 1986 correlated with the level of radiation in eight Czech regions after the Chernobyl disaster. RESULTS: We found a relationship between the level of radiation and the decrease in the number of newborn boys. The number of newborn boys was decreased in the six eastern regions where the radiation was strongly increased due to rain that accompanied the radioactive cloud. In contrast, the number of newborn boys was not reduced in the two western regions where the radioactivity was markedly lower. CONCLUSIONS: A negative impact of radiation on the prenatal population was manifested as a selective loss of newborn boys in November 1986. This loss correlated with level of radioactivity. The 131I probably played the most important role because of its up-take during primary saturation of fetal thyroid by iodine, which accompanies the onset of the gland function in 3-month-old fetuses.

Disruption of the palatal rugae pattern in Tabby (eda) mutant mice.

The eda mouse gene is linked with anomalies of ectodermal derivatives, such as hair, glands, and teeth. The palatal rugae (oral mucosa foldings on the hard palate) are also ectodermal derivatives. Therefore, we searched for and compared palatal rugae anomalies of Tabby mice bearing a mutation in the eda gene with their wild-type counterparts. We compared the number and shape of palatal rugae in 179 mutant and 102 wild-type mice from four different stocks of Tabby mice. Palatal rugae anomalies were documented at a low frequency in wild-type mice of different backgrounds, which may reflect a lack of robustness of palatal rugae development. However, the proportion of anomalies observed in the C57BL/6J background makes us recommend avoiding its use in further palate studies. We showed statistically that the phenotypic variability seen in wild-type animals is further increased in Tabby mutants. The anomalies mainly included various forms of reduction, with rugae IV-VI being more frequently affected. Those rugae were shortened, dotted or absent (mainly ruga V). By analogy to the role played by eda in other ectodermal derivatives, we propose that it might play a role in defining the pattern of the palatal rugae.

Early development of the lower deciduous dentition and oral vestibule in human embryos.

The aim of this work was to investigate the early development of the deciduous dentition and oral vestibule in the human embryonic lower jaw. Histological sections and three-dimensional reconstructions from prenatal weeks 6-9 were used. A continuous anlage for the oral vestibule did not exist in the mandible. In contrast to the upper jaw, where we previously observed that the dental and vestibular epithelia developed separately, two dento-vestibular bulges differentiated in the incisor region of the mandible. The lingual parts of each bulge were found to give rise to the respective central and lateral incisors, whereas the labial parts differentiated into the vestibular epithelium. In the canine and molar areas, the dental and vestibular epithelia originated separately. Later, the segments of the vestibular epithelium fused into the labial vestibular ridge, giving rise to the lower oral vestibule in the lip region. In the cheek region, the oral vestibule was found to originate in the mucosal inflection between the developing jaw and the cheek. A similar heterogeneous developmental base for the oral vestibule was also observed in the upper jaw. There is thus no general scheme for the early development of the dental and vestibular epithelia that applies to both the upper and lower jaws, and to both their anterior and posterior regions.