tretinoin and Craniofacial-Abnormalities

Fetal Alcohol Spectrum Disorder (FASD) is a set of neurodevelopmental malformations caused by maternal consumption of alcohol during pregnancy. FASD sentinel facial features are unique to the disorder, and microcephaly is common in severe forms of FASD. Retinoic acid deficiency has been shown to cause craniofacial malformations and microcephaly in animal models reminiscent of those caused by prenatal alcohol exposure. Alcohol exposure affects the migration and survival of cranial neural crest cells, which are required for proper frontonasal prominence and pharyngeal arch development. Defects in craniofacial development are further amplified by the many downstream pathways that are transcriptionally controlled retinoic acid target genes, including Shh signaling. Recent evidence shows that alcohol exposure itself is sufficient to induce retinoic acid deficiency in the embryo. These data suggest that retinoic acid deficiency is an important underlying etiology of FASD. In disorders like Vitamin A Deficiency, FASD, DiGeorge (22q11.2 Deletion Syndrome), CHARGE, Smith-Magenis, Matthew-Wood, and Congenital Zika Syndromes, evidence is accumulating to link reduced retinoic acid signaling with developmental defects like craniofacial malformations and microcephaly. Research focus on characterizing the effects of retinoic acid deficiency during early development and on understanding the downstream signaling pathways involved in aberrant head, and craniofacial development will reveal underlying etiologies of these disorders.

Topics: Animals; Craniofacial Abnormalities; Fetal Alcohol Spectrum Disorders; Humans; Microcephaly; Neural Crest; Tretinoin

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.

Topics: Animals; Craniofacial Abnormalities; Embryonic Induction; Eye Abnormalities; Humans; Neural Crest; Signal Transduction; Tretinoin

Prenatal exposure to ethanol results in fetal alcohol spectrum disorder (FASD), a syndrome characterised by a broad range of clinical manifestations including craniofacial dysmorphologies and neurological defects. The characterisation of the mechanisms by which ethanol exerts its teratogenic effects is difficult due to the pleiotropic nature of its actions. Different experimental model systems have been employed to investigate the aetiology of FASD. Here, I will review studies using these different model organisms that have helped to elucidate how ethanol causes the craniofacial abnormalities characteristic of FASD. In these studies, ethanol was found to impair the prechordal plate-an important embryonic signalling centre-during gastrulation and to negatively affect the induction, migration and survival of the neural crest, a cell population that generates the cartilage and most of the bones of the skull. At the cellular level, ethanol appears to inhibit Sonic hedgehog signalling, alter levels of retionoic acid activity, trigger a Ca(2+)-CamKII-dependent pathway that antagonises WNT signalling, affect cytoskeletal dynamics and increase oxidative stress. Embryos of the domestic chick Gallus gallus domesticus have played a central role in developing a working model for the effects of ethanol on craniofacial development because they are easily accessible and because key steps in craniofacial development are particularly well established in the avian embryo. I will finish this review by highlighting some potential future avenues of fetal alcohol research.

Topics: Abnormalities, Drug-Induced; Animals; Calcium Signaling; Chick Embryo; Craniofacial Abnormalities; Disease Models, Animal; Embryo, Mammalian; Embryo, Nonmammalian; Endoderm; Ethanol; Face; Fetal Alcohol Spectrum Disorders; Gastrula; Genetic Predisposition to Disease; Hedgehog Proteins; Holoprosencephaly; Humans; Maxillofacial Development; Neural Crest; Signal Transduction; Skull; Species Specificity; Tretinoin; Wnt Signaling Pathway

The shaping of the vertebrate head results from highly dynamic integrated processes involving the growth and exchange of signals between the ectoderm, the endoderm, the mesoderm and Cephalic Neural Crest Cells (CNCCs). During embryonic development, these tissues change their shape and relative position rapidly and come transiently in contact with each other. Molecular signals exchanged in restricted regions of tissue interaction are crucial in providing positional identity to the mesenchymes which will form the different skeletal and muscular components of the head. Slight spatio-temporal modifications of these signalling maps can result in profound changes in craniofacial development and might have contributed to the evolution of facial diversity. Abnormal signalling patterns could also be at the origin of congenital craniofacial malformations. This review brings into perspective recent work on spatial and temporal aspects of facial morphogenesis with particular focus on the molecular mechanisms of jaw specification.

Topics: Animals; Body Patterning; Craniofacial Abnormalities; Ectoderm; Endoderm; Endothelin-1; Gene Expression Regulation, Developmental; Humans; Jaw; Mesoderm; Models, Biological; Neural Crest; Signal Transduction; Time Factors; Tretinoin

Fetal alcohol syndrome (FAS) is a collection of signs and symptoms seen in children exposed to alcohol in the prenatal period. It is characterized mainly by a distinct pattern of craniofacial malformations, physical and mental retardation. However, with the increased incidence of FAS, there is a great variation in the clinical features of FAS.. Narrative review.. This review describes data from clinical and experimental studies, and in vitro models. Experimental studies have shown that alcohol has a direct toxic effect on the ectodermal and mesodermal cells of the developing embryo, particularly in the cells destined to give rise to dentofacial structures (i.e. cranial neural crest cells). Other effects, such as, abnormal pattern of cranial and mandibular growth and altered odontogenesis are described in detail. The exact mechanism by which alcohol induces its teratogenic effects remains still unknown. The possible mechanisms are outlined here, with an emphasis on the developing face and tooth. Possible future research directions and treatment strategies are also discussed.. Early identification of children affected by prenatal alcohol exposure leads to interventions, services, and improved outcomes. FAS can be prevented with the elimination of alcohol consumption during pregnancy. We need to provide education, target high-risk groups, and make this issue a high priority in terms of public health.

Topics: Animals; Cell Membrane; Craniofacial Abnormalities; Facies; Female; Fetal Alcohol Spectrum Disorders; Free Radicals; Gene Expression Regulation, Developmental; Growth Substances; Humans; Neural Crest; Pregnancy; Tooth Abnormalities; Tretinoin

In this paper I have tried to bring together work that highlights the role of homeobox genes in generating craniofacial form. I review both normal and disrupted embryogenesis and ask whether mis-expression of the homeobox genes outside their normal domains could be contributing to congenital facial abnormalities arising from either genetic or teratogenic actions. Experimentally generated transgenic mice carrying loss- or gain-of-function mutations in homeobox genes, in combination with their normal expression patterns, have allowed us to compile and test models of embryonic specification based around a Hox/homeobox code. These models form the basis on which the functional questions are considered. There are four major sections covering different experimental approaches designed to ectopically induce homeobox genes in the head. Transgenic mice, where heterologous promoters drive a given Hox gene in the head, have shown that the more posteriorly expressed Hox genes tend to have a significant effect only on the skull bones of mesodermal origin whereas those normally expressed more anteriorly, in the hindbrain and branchial arches, can affect more anterior branchial arch and neural crest-derived structures. Manipulation experiments which can induce homeobox genes in small, localised regions of the facial precursors show clear and dramatic effects of this expression on facial development. Null mutations in predicted repressors of Hox gene expression, however, do not appear to give rise to substantial craniofacial abnormalities. Retinoic acid, on the other hand, is well known for its teratogenic actions and its ability to induce Hox gene expression. Evidence is now accumulating that at least some of its teratogenic actions may be mediated by its regulation of the Hox and other homeobox genes in the head.

Topics: Abnormalities, Drug-Induced; Animals; Brain; Chick Embryo; Craniofacial Abnormalities; Face; Facial Bones; Gene Expression Regulation, Developmental; Genes, Homeobox; Head; Homeodomain Proteins; Mice; Mice, Knockout; Mice, Transgenic; Morphogenesis; Neural Crest; Organ Specificity; Promoter Regions, Genetic; Recombinant Fusion Proteins; Tretinoin

AP-2 is a recent significant addition to the list of transcription factors that have been demonstrated by targeted gene disruption to be essential for normal development. Two recent reports of AP-2 null mutant mice indicate that AP-2 holds a key position in the network of genes and proteins controlling developmental pattern and morphogenesis, and that it is particularly important for development of the cranial region and for midline fusions.

Topics: Animals; Craniofacial Abnormalities; DNA-Binding Proteins; Eye Abnormalities; Limb Deformities, Congenital; Mice; Mice, Knockout; Mutation; Neural Crest; Phenotype; Transcription Factor AP-2; Transcription Factors; Tretinoin

Environmental teratogens such as smoking are known risk factors for developmental disorders such as cleft palate. While smoking rates have declined, a new type of smoking, called vaping is on the rise. Vaping is the use of e-cigarettes to vaporize and inhale an e-liquid containing nicotine and food-like flavors. There is the potential that, like smoking, vaping could also pose a danger to the developing human. Rather than waiting for epidemiological and mammalian studies, we have turned to an aquatic developmental model, Xenopus laevis, to more quickly assess whether e-liquids contain teratogens that could lead to craniofacial malformations. Xenopus, like zebrafish, has the benefit of being a well-established developmental model and has also been effective in predicting whether a chemical could be a teratogen. We have determined that embryonic exposure to dessert flavored e-liquids can cause craniofacial abnormalities, including an orofacial cleft in Xenopus. To better understand the underlying mechanisms contributing to these defects, transcriptomic analysis of the facial tissues of embryos exposed to a representative dessert flavored e-liquid vapor extract was performed. Analysis of differentially expressed genes in these embryos revealed several genes associated with retinoic acid metabolism or the signaling pathway. Consistently, retinoic acid receptor inhibition phenocopied the craniofacial defects as those embryos exposed to the vapor extract of the e-liquid. Such malformations also correlated with a group of common differentially expressed genes, two of which are associated with midface birth defects in humans. Further, e-liquid exposure sensitized embryos to forming craniofacial malformations when they already had depressed retinoic acid signaling. Moreover, 13-cis-retinoic acid treatment could significantly reduce the e-liquid induced malformation in the midface. Such results suggest the possibility of an interaction between retinoic acid signaling and e-liquid exposure. One of the most popular and concentrated flavoring chemicals in dessert flavored e-liquids is vanillin. Xenopus embryos exposed to this chemical closely resembled embryos exposed to dessert-like e-liquids and a retinoic acid receptor antagonist. In summary, we determined that e-liquid chemicals, in particular vanillin, can cause craniofacial defects potentially by dysregulating retinoic acid signaling. This work warrants the evaluation of vanillin and other such flavoring additiv

Topics: Animals; Benzaldehydes; Craniofacial Abnormalities; Embryo, Nonmammalian; Flavoring Agents; Signal Transduction; Tobacco Products; Tretinoin; Xenopus laevis

Embryonic craniofacial development depends on the coordinated outgrowth and fusion of multiple facial primordia, which are populated with cranial neural crest cells and covered by the facial ectoderm. Any disturbance in these developmental events, their progenitor tissues, or signaling pathways can result in craniofacial deformities such as orofacial clefts, which are among the most common birth defects in humans. In the present study, we show that

Topics: Animals; Cleft Lip; Cleft Palate; Craniofacial Abnormalities; Embryonic Development; Hedgehog Proteins; Mice; Neural Crest; Tretinoin

Adverse outcome pathway (AOP) is a conceptual framework that links a molecular initiating event (MIE) via intermediate key events (KEs) with adverse effects (adverse outcomes, AO) relevant for risk assessment, through defined KE relationships (KERs). The aim of the present work is to describe a linear AOP, supported by experimental data, for skeletal craniofacial defects as the AO. This AO was selected in view of its relative high incidence in humans and the suspected relation to chemical exposure. We focused on inhibition of CYP26, a retinoic acid (RA) metabolizing enzyme, as MIE, based on robust previously published data. Conazoles were selected as representative stressors. Intermediate KEs are RA disbalance, aberrant HOX gene expression, disrupted specification, migration, and differentiation of neural crest cells, and branchial arch dysmorphology. We described the biological basis of the postulated events and conducted weight of evidence (WoE) assessments. The biological plausibility and the overall empirical evidence were assessed as high and moderate, respectively, the latter taking into consideration the moderate evidence for concordance of dose-response and temporal relationships. Finally, the essentiality assessment of the KEs, considered as high, supported the robustness of the presented AOP. This AOP, which appears of relevance to humans, thus contributes to mechanistic underpinning of selected test methods, thereby supporting their application in integrated new approach test methodologies and strategies and application in a regulatory context.

Topics: Adverse Outcome Pathways; Animals; Azoles; Craniofacial Abnormalities; Cytochrome P450 Family 26; Female; Gene Expression Regulation; Humans; Male; Mice; Neural Crest; Risk Assessment; Tretinoin

Multiple syndromes share congenital heart and craniofacial muscle defects, indicating there is an intimate relationship between the adjacent cardiac and pharyngeal muscle (PM) progenitor fields. However, mechanisms that direct antagonistic lineage decisions of the cardiac and PM progenitors within the anterior mesoderm of vertebrates are not understood. Here, we identify that retinoic acid (RA) signaling directly promotes the expression of the transcription factor Nr2f1a within the anterior lateral plate mesoderm. Using zebrafish nr2f1a and nr2f2 mutants, we find that Nr2f1a and Nr2f2 have redundant requirements restricting ventricular cardiomyocyte (CM) number and promoting development of the posterior PMs. Cre-mediated genetic lineage tracing in nr2f1a; nr2f2 double mutants reveals that tcf21+ progenitor cells, which can give rise to ventricular CMs and PM, more frequently become ventricular CMs potentially at the expense of posterior PMs in nr2f1a; nr2f2 mutants. Our studies reveal insights into the molecular etiology that may underlie developmental syndromes that share heart, neck and facial defects as well as the phenotypic variability of congenital heart defects associated with NR2F mutations in humans.

Topics: Animals; Animals, Genetically Modified; Body Patterning; Cell Lineage; COUP Transcription Factor II; Craniofacial Abnormalities; DNA-Binding Proteins; Embryonic Stem Cells; Heart Defects, Congenital; Heart Ventricles; Humans; Mesoderm; Models, Animal; Mutation; Myocytes, Cardiac; Pharyngeal Muscles; Promoter Regions, Genetic; Signal Transduction; Transcription Factors; Tretinoin; Zebrafish; Zebrafish Proteins

Retinoic acid (RA) is required for embryonic formation of the anterior segment of the eye and craniofacial structures. The present study further investigated the role of RA in maintaining the function of these neural crest-derived structures in adult zebrafish.. Morphology and histology were analyzed by using live imaging, methylacrylate sections, and TUNEL assay. Functional analysis of vision and aqueous humor outflow were assayed with real-time imaging.. Both decreased and increased RA signaling altered craniofacial and ocular structures in adult zebrafish. Exogenous treatment with all-trans RA for 5 days resulted in a prognathic jaw, while inhibition of endogenous RA synthesis through treatment with 4-diethylaminobenzaldehyde (DEAB) decreased head height. In adult eyes, RA activity was localized to the retinal pigment epithelium, photoreceptors, outer plexiform layer, inner plexiform layer, iris stroma, and ventral canalicular network. Exogenous RA increased apoptosis in the iris stroma and canalicular network in the ventral iridocorneal angle, resulting in the loss of these structures and decreased aqueous outflow. DEAB, which decreased RA activity throughout the eye, induced widespread apoptosis, resulting in corneal edema, cataracts, retinal atrophy, and loss of iridocorneal angle structures. DEAB-treated fish were blind with no optokinetic response and no aqueous outflow from the anterior chamber.. Tight control of RA levels is required for normal structure and function of the adult anterior segment. These studies demonstrated that RA plays an important role in maintaining ocular and craniofacial structures in adult zebrafish.

Topics: Animals; Anterior Eye Segment; Antineoplastic Agents; Apoptosis; Aqueous Humor; Craniofacial Abnormalities; Female; Gene Expression Regulation, Developmental; In Situ Nick-End Labeling; Male; Neural Crest; Nystagmus, Optokinetic; Real-Time Polymerase Chain Reaction; Tretinoin; Vision, Ocular; Zebrafish

Development of the face and mouth is orchestrated by a large number of transcription factors, signaling pathways and epigenetic regulators. While we know many of these regulators, our understanding of how they interact with each other and implement changes in gene expression during orofacial development is still in its infancy. Therefore, this study focuses on uncovering potential cooperation between transcriptional regulators and one important signaling pathway, retinoic acid, during development of the midface.. Transcriptome analyses was performed on facial tissues deficient for retinoic acid receptor function at two time points in development; early (35 hpf) just after the neural crest migrates and facial tissues are specified and later (60 hpf) when the mouth has formed and facial structures begin to differentiate. Functional and network analyses revealed that retinoic acid signaling could cooperate with novel epigenetic factors and calcium-NFAT signaling during early orofacial development. At the later stage, retinoic acid may work with WNT and BMP and regulate homeobox containing transcription factors. Finally, there is an overlap in genes dysregulated in Xenopus embryos with median clefts with human genes associated with similar orofacial defects.. This study uncovers novel signaling pathways required for orofacial development as well as pathways that could interact with retinoic acid signaling during the formation of the face. We show that frog faces are an important tool for studying orofacial development and birth defects.

Topics: Animals; Computational Biology; Craniofacial Abnormalities; Gene Expression Profiling; Gene Expression Regulation, Developmental; Gene Regulatory Networks; Humans; Organ Specificity; Organogenesis; Phenotype; Receptors, Retinoic Acid; Signal Transduction; Transcriptome; Tretinoin; Xenopus

Syndromes that display craniofacial anomalies comprise a major class of birth defects. Both genetic and environmental factors, including prenatal retinoic acid (RA) exposure, have been associated with these syndromes. While next generation sequencing has allowed the discovery of new genes implicated in these syndromes, some are still poorly characterized such as Oculo-Auriculo-Vertebral Spectrum (OAVS). Due to the lack of clear diagnosis for patients, developing new strategies to identify novel genes involved in these syndromes is warranted. Thus, our study aimed to explore the link between genetic and environmental factors. Owing to a similar phenotype of OAVS reported after gestational RA exposures in humans and animals, we explored RA targets in a craniofacial developmental context to reveal new candidate genes for these related disorders. Using a proteomics approach, we detected 553 dysregulated proteins in the head region of mouse embryos following their exposure to prenatal RA treatment. This novel proteomic approach implicates changes in proteins that are critical for cell survival/apoptosis and cellular metabolism which could ultimately lead to the observed phenotype. We also identified potential molecular links between three major environmental factors known to contribute to craniofacial defects including maternal diabetes, prenatal hypoxia and RA exposure. Understanding these links could help reveal common key pathogenic mechanisms leading to craniofacial disorders. Using both in vitro and in vivo approaches, this work identified two new RA targets, Gnai3 and Eftud2, proteins known to be involved in craniofacial disorders, highlighting the power of this proteomic approach to uncover new genes whose dysregulation leads to craniofacial defects.

Topics: Craniofacial Abnormalities; Female; Humans; Pregnancy; Prenatal Exposure Delayed Effects; Tretinoin

To characterize the prenatal and postnatal craniofacial bone development in mouse model of all-trans retinoic acid (ATRA) exposure at different ages by a quantitative and morphological analysis of skull morphology.. Pregnant mice were exposed to ATRA at embryonic day 10 (E10) and 13 (E13) by oral gavage. Skulls of mice embryos at E19.5 and adult mice at postnatal day 35 (P35) were collected for high-resolution microcomputed tomography (microCT) imaging scanning and section HE staining. Reconstruction and measurement of mouse skulls were performed for prenatal and postnatal analysis of the control and ATRA-exposed mice.. Craniofacial malformations in mouse models caused by ATRA exposure were age dependent. ATRA exposure at E10 induced cleft palate in 81.8% of the fetuses, whereas the palatine bone of E13-exposed mice was intact. Inhibitions of maxilla and mandible development with craniofacial asymmetry induced were observed at E19.5 and P35. Compared with control and E13-exposed mice, the palatine bones of E10-exposed mice were not elevated and were smaller in dimension. Some E10-exposed mice exhibited other craniofacial abnormalities, including premature fusion of mandibular symphysis with a missing mandibular incisor and a smaller mandible. Severe deviated snouts and amorphous craniofacial suture were detected in E13-exposed mice at P35.. These morphological variations in E10- and E13-exposed mice suggested that ATRA was teratogenic in craniofacial bone development in mice and the effect was age dependent.

Topics: Age Factors; Animals; Animals, Newborn; Craniofacial Abnormalities; Disease Models, Animal; Female; Mice; Pregnancy; Radiographic Image Interpretation, Computer-Assisted; Tretinoin; X-Ray Microtomography

We assessed feeding-related developmental anomalies in the LgDel mouse model of chromosome 22q11 deletion syndrome (22q11DS), a common developmental disorder that frequently includes perinatal dysphagia--debilitating feeding, swallowing and nutrition difficulties from birth onward--within its phenotypic spectrum. LgDel pups gain significantly less weight during the first postnatal weeks, and have several signs of respiratory infections due to food aspiration. Most 22q11 genes are expressed in anlagen of craniofacial and brainstem regions critical for feeding and swallowing, and diminished expression in LgDel embryos apparently compromises development of these regions. Palate and jaw anomalies indicate divergent oro-facial morphogenesis. Altered expression and patterning of hindbrain transcriptional regulators, especially those related to retinoic acid (RA) signaling, prefigures these disruptions. Subsequently, gene expression, axon growth and sensory ganglion formation in the trigeminal (V), glossopharyngeal (IX) or vagus (X) cranial nerves (CNs) that innervate targets essential for feeding, swallowing and digestion are disrupted. Posterior CN IX and X ganglia anomalies primarily reflect diminished dosage of the 22q11DS candidate gene Tbx1. Genetic modification of RA signaling in LgDel embryos rescues the anterior CN V phenotype and returns expression levels or pattern of RA-sensitive genes to those in wild-type embryos. Thus, diminished 22q11 gene dosage, including but not limited to Tbx1, disrupts oro-facial and CN development by modifying RA-modulated anterior-posterior hindbrain differentiation. These disruptions likely contribute to dysphagia in infants and young children with 22q11DS.

Topics: Animals; Animals, Newborn; Body Patterning; Chromosome Deletion; Cranial Nerves; Craniofacial Abnormalities; Deglutition; Deglutition Disorders; DiGeorge Syndrome; Disease Models, Animal; Embryo, Mammalian; Feeding Behavior; Female; Gene Dosage; Gene Expression Regulation, Developmental; Male; Mice; Phenotype; Rhombencephalon; Signal Transduction; T-Box Domain Proteins; Tretinoin

To characterize the abnormal metabolic profile of all-trans-retinoic acid (ATRA)-induced craniofacial development in mouse embryos using proton magnetic resonance spectroscopy (1H-MRS).. Timed-pregnant mice were treated by oral gavage on the morning of embryonic gestation day 11 (E11) with all-trans-retinoic acid (ATRA). Dosing solutions were adjusted by maternal body weight to provide 30, 70, or 100 mg/kg RA. The control group was given an equivalent volume of the carrier alone. Using an Agilent 7.0 T MR system and a combination of surface coil coils, a 3 mm×3 mm×3 mm 1H-MRS voxel was selected along the embryonic craniofacial tissue. 1H-MRS was performed with a single-voxel method using PRESS sequence and analyzed using LCModel software. Hematoxylin and eosin was used to detect and confirm cleft palate.. 1H-MRS revealed elevated choline levels in embryonic craniofacial tissue in the RA70 and RA100 groups compared to controls (P<0.05). Increased choline levels were also found in the RA70 and RA100 groups compared with the RA30 group (P<0.01). High intra-myocellular lipids at 1.30 ppm (IMCL13) in the RA100 group compared to the RA30 group were found (P<0.01). There were no significant changes in taurine, intra-myocellular lipids at 2.10 ppm (IMCL21), and extra-myocellular lipids at 2.30 ppm (EMCL23). Cleft palate formation was observed in all fetuses carried by mice administered 70 and 100 mg/kg RA.. This novel study suggests that the elevated choline and lipid levels found by 1H-MRS may represent early biomarkers of craniofacial defects. Further studies will determine performance of this test and pathogenetic mechanisms of craniofacial malformation.

Topics: Abnormalities, Drug-Induced; Animals; Antineoplastic Agents; Choline; Cleft Palate; Craniofacial Abnormalities; Creatine; Dose-Response Relationship, Drug; Embryo, Mammalian; Female; Lipids; Male; Mice; Pregnancy; Proton Magnetic Resonance Spectroscopy; Taurine; Tretinoin

Rare, recurrent genomic imbalances facilitate the association of genotype with abnormalities at the "whole body" level. However, at the cellular level, the functional consequences of recurrent genomic abnormalities and how they can be linked to the phenotype are much less investigated.. We report an example of a functional analysis of two genes from a new, overlapping microdeletion of 2p13.2 region (from 72,140,702-72,924,626). The subjects shared intellectual disability (ID), language delay, hyperactivity, facial asymmetry, ear malformations, and vertebral and/or craniofacial abnormalities. The overlapping region included two genes, EXOC6B and CYP26B1, which are involved in exocytosis/Notch signaling and retinoic acid (RA) metabolism, respectively, and are of critical importance for early morphogenesis, symmetry as well as craniofacial, skeleton and brain development. The abnormal function of EXOC6B was documented in patient lymphoblasts by its reduced expression and with perturbed expression of Notch signaling pathway genes HES1 and RBPJ, previously noted to be the consequence of EXOC6B dysfunction in animal and cell line models. Similarly, the function of CYP26B1 was affected by the deletion since the retinoic acid induced expression of this gene in patient lymphoblasts was significantly lower compared to controls (8% of controls).. Haploinsufficiency of CYP26B1 and EXOC6B genes involved in retinoic acid and exocyst/Notch signaling pathways, respectively, has not been reported previously in humans. The developmental anomalies and phenotypic features of our subjects are in keeping with the dysfunction of these genes, considering their known role. Documenting their dysfunction at the cellular level in patient cells enhanced our understanding of biological processes which contribute to the clinical phenotype.

Topics: Abnormalities, Multiple; Adolescent; Bone Diseases, Developmental; Cell Line; Child; Chromosome Deletion; Chromosomes, Human, Pair 2; Craniofacial Abnormalities; Cytochrome P-450 Enzyme System; Developmental Disabilities; Exocytosis; Genotype; GTP-Binding Proteins; Haploinsufficiency; Humans; Male; Oligonucleotide Array Sequence Analysis; Phenotype; Retinoic Acid 4-Hydroxylase; Tretinoin

To determine changes on craniofacial growth morphometrically in newborn mice with cleft palate induced by retinoic acid. DESIGN, SETTING, PARTICIPANTS, INTERVENTIONS: Gestation day 10 or 12 pregnant female C57BL/6N mice were given a single dose of all-trans retinoic acid (atRA) by gastric intubations via oral gavage. Sixty newborn mice with cleft palate (CP), 52 without CP from the experimental group, and 30 without CP from the control group were collected, and lateral cephalograms were taken of all of the mice.. Cephalometric analysis of the craniofacial skeleton was performed by means of a personal computer.. Inhibition of craniofacial growth was found in the experimental groups but not in the control groups. In the maxillary bone and mandible, the amount of growth was significantly reduced.. These results suggest that craniofacial growth is inhibited in newborn mice with cleft palate induced by retinoic acid.

Topics: Animals; Animals, Newborn; Cephalometry; Cleft Palate; Craniofacial Abnormalities; Embryo, Mammalian; Female; Fetus; Gestational Age; Mice; Mice, Inbred C57BL; Pregnancy; Radiography; Tretinoin

The purpose of the present study was to identify the potential effect of prenatal vitamin B12 administration on retinoic acid (RA)-induced early craniofacial abnormalities in mice and to investigate the possible mechanisms by which vitamin B12 reduces malformations.. In our study, whole embryo culture was used to explore the effect of vitamin B12 on mouse embryos during the critical period of organogenesis. All embryos were exposed to 0.4 µM RA and different concentrations of vitamin B12 and scored for their growth in the branchial region at the end of a 48-hour culture period. The endothelin-1 (ET-1)/dHAND protein expression levels in the first branchial arch were investigated using an immunohistochemical method.. In the whole embryo culture, 100 and 10 µM vitamin B12 dose-dependently prevented branchial region malformations and decreased craniofacial defects by 90.5% and 77.3%, respectively. ET-1 and dHAND protein levels were significantly increased in vitamin B12-supplemented embryos compared to the RA-exposed group in embryonic branchial region.. These results suggest that vitamin B12 may prevent RA-induced craniofacial abnormalities via prevention of an RA-induced decrease of ET-1 and dHAND protein levels in the branchial region during the organogenic period. This study may shed new light on preventing craniofacial abnormalities.

Topics: Animals; Basic Helix-Loop-Helix Transcription Factors; Branchial Region; Craniofacial Abnormalities; Dose-Response Relationship, Drug; Embryo Culture Techniques; Embryonic Development; Endothelin-1; Facial Bones; Female; Male; Mice; Mice, Inbred ICR; Microcephaly; Neural Tube Defects; Tretinoin; Vitamin B 12; Vitamin B Complex

Retinoic acid is a widely used drug in the treatment of cystic acne. It has teratogenic effects that depend on the gestational period in which it is used. We report a seven months old female whose mother was exposed to retinoic acid in both pre-gestational and gestational periods. She had a retardation of psychomotor development and a brain MRI showed frontal atrophy and a malformation of the posterior fossa. We discuss the mechanisms of the teratogenic effects of retinoic acid.

Topics: Abnormalities, Drug-Induced; Abnormalities, Multiple; Acne Vulgaris; Atrophy; Cranial Fossa, Posterior; Craniofacial Abnormalities; Female; Frontal Lobe; Humans; Infant; Isotretinoin; Keratolytic Agents; Maternal Exposure; Pregnancy; Prenatal Exposure Delayed Effects; Psychomotor Disorders; Teratogens; Tretinoin

The cellular uptake of vitamin A from its RBP4-bound circulating form (holo-RBP4) is a homeostatic process that evidently depends on the multidomain membrane protein STRA6. In humans, mutations in STRA6 are associated with Matthew-Wood syndrome, manifested by multisystem developmental malformations. Here we addressed the metabolic basis of this inherited disease. STRA6-dependent transfer of retinol from RBP4 into cultured NIH 3T3 fibroblasts was enhanced by lecithin:retinol acyltransferase (LRAT). The retinol transfer was bidirectional, strongly suggesting that STRA6 acts as a retinol channel/transporter. Loss-of-function analysis in zebrafish embryos revealed that Stra6 deficiency caused vitamin A deprivation of the developing eyes. We provide evidence that, in the absence of Stra6, holo-Rbp4 provokes nonspecific vitamin A excess in several embryonic tissues, impairing retinoic acid receptor signaling and gene regulation. These fatal consequences of Stra6 deficiency, including craniofacial and cardiac defects and microphthalmia, were largely alleviated by reducing embryonic Rbp4 levels by morpholino oligonucleotide or pharmacological treatments.

Topics: Abnormalities, Multiple; Acyltransferases; Animals; Cardiovascular Abnormalities; Craniofacial Abnormalities; Disease Models, Animal; Eye; Gene Deletion; Gene Expression Regulation, Developmental; Homeostasis; Humans; Membrane Proteins; Membrane Transport Proteins; Mice; Morpholines; NIH 3T3 Cells; Oligonucleotides, Antisense; Retinol-Binding Proteins, Plasma; Syndrome; Time Factors; Transduction, Genetic; Tretinoin; Vitamin A; Zebrafish; Zebrafish Proteins

Intake of retinoic acid (RA) or of its precursor, vitamin A, during early pregnancy is associated with increased incidence of craniofacial lesions. The origin of these teratogenic effects remains enigmatic as in cranial neural crest cells (CNCCs), which largely contribute to craniofacial structures, the RA-transduction pathway is not active. Recent results suggest that RA could act on the endoderm of the first pharyngeal arch (1stPA), through a RARbeta-dependent mechanism.. Here we show that RA provokes dramatically different craniofacial malformations when administered at slightly different developmental times within a narrow temporal interval corresponding to the colonization of the 1(st) PA by CNCCs. We provide evidence showing that RA acts on the signalling epithelium of the 1(st) PA, gradually reducing the expression of endothelin-1 and Fgf8. These two molecular signals are instrumental in activating Dlx genes in incoming CNCCs, thereby triggering the morphogenetic programs, which specify different jaw elements.. The anatomical series induced by RA-treatments at different developmental times parallels, at least in some instances, the supposed origin of modern jaws (e.g., the fate of the incus). Our results might provide a conceptual framework for the rise of jaw morphotypes characteristic of gnathostomes.

Topics: Animals; Craniofacial Abnormalities; Early Growth Response Protein 2; Embryo, Mammalian; Endothelin-1; Female; Gene Expression Regulation, Developmental; Homeodomain Proteins; In Situ Hybridization; Jaw; Keratolytic Agents; Mice; Mice, Transgenic; Molecular Dynamics Simulation; Neural Crest; Pregnancy; Receptors, Retinoic Acid; Signal Transduction; Tretinoin

To analyze the expression and role of three proteins (HSP110, caspase-3 and caspase-9) during craniofacial development.. Seven pregnant C57Bl/6J mice received, by force-feeding at gestation day 9 (E9), 80 mg/kg of all-trans retinoic acid mixed to sesame oil. Seven pregnant NMRI mice received two grays irradiation at the same gestation day. Control mice of both strains (seven mice for each strain) were not submitted to any treatment. Embryos were obtained at various stages after exposition (3, 6, 12 and 24 h), fixed, dehydrated and embedded. Coronal sections (5 microm) were made. Slide staining occurred alternatively using anti-Hsp110, anti-caspase-3 and anti-caspase-9 immunohistochemistry.. Expression of HSP110, caspase-3 and caspase-9 was found in cells of well-known locations of programmed cell death. After retinoic acid exposure, expressions were increased especially in neural crest cells of mandibular and hyoid arches. Quantification of positive cells shows that caspase-9 and Hsp110 were expressed before caspase-3. After irradiation, the expression of the three proteins quickly increased with a maximum 3 h after irradiation. For all three models of apoptosis (physiological, retinoic-induced and irradiation-induced) HSP110 positive cells were more numerous than caspase-3 positive cells. Caspase-3 positive cells were more numerous than caspase-9 positive cells especially in mesectodermal irradiation-induced apoptotic cells.. The findings show a potential function of HSP110 in apoptosis during embryo development. Caspase-3-expressing cells are more numerous than cells expressing caspase-9, especially irradiation-induced apoptotic neural crest cells. This suggests that other caspases, still to be identified, may activate caspase-3 in this model.

Topics: Abnormalities, Drug-Induced; Abnormalities, Radiation-Induced; Animals; Apoptosis; Caspase 3; Caspase 9; Caspases; Craniofacial Abnormalities; Embryonic Development; Female; HSP110 Heat-Shock Proteins; Immunohistochemistry; Maxillofacial Development; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Neural Crest; Pregnancy; Tretinoin

Prevention of retinoic acid-induced craniofacial abnormalities by folinic acid, and endothelin-1 (ET-1)/dHAND protein and mRNA expression were investigated in mouse embryos using the whole embryo culture, streptavidin-biotin peroxidase complex method, and whole-mount in situ hybridization. In the whole embryo culture, 1.0 and 0.1 mm-folinic acid dose dependently prevented branchial region malformations and decreased defects by 93 % and 77 %, respectively. Folinic acid at concentrations of 1.0 and 0.1 mm significantly increased ET-1 and dHAND protein expression levels compared to retinoic acid-exposed values in embryonic branchial areas. Folinic acid also increased ET-1 and dHAND mRNA levels in the same region. The present results suggest that folinic acid may prevent retinoic acid-induced craniofacial abnormalities via increasing ET-1 and dHAND levels in the branchial region during the organogenic period.

Topics: Animals; Basic Helix-Loop-Helix Transcription Factors; Brain; Branchial Region; Craniofacial Abnormalities; Embryo Culture Techniques; Embryo, Mammalian; Endothelin-1; Female; Gene Expression Regulation; Immunohistochemistry; In Situ Hybridization; Leucovorin; Male; Mice; RNA, Messenger; Teratogens; Tretinoin; Vitamin B Complex

(1) In 1998 several cases of malformations similar to those induced by oral retinoids were reported in children exposed in utero to topical retinoids (adapalene and tretinoin). The results of two somewhat flawed epidemiological studies were reassuring. (2) New cases of birth defects were subsequently reported in children exposed in utero to topical tretinoin. (3) Epidemiological data are still scant and unconvincing: they neither confirm this risk nor rule it out completely. (4) It is best to avoid using topical retinoids altogether in early pregnancy. Women of child-bearing age must be fully informed of the risks and the importance of effective contraception. This also applies to patients with moderate forms of psoriasis, for which topical tazaroten is indicated.

Topics: Abnormalities, Drug-Induced; Acitretin; Administration, Topical; Craniofacial Abnormalities; Female; France; Humans; Infant, Newborn; Maternal Exposure; Naphthalenes; Pregnancy; Prenatal Exposure Delayed Effects; Retinoids; Tretinoin

Triadimefon and triadimenol, fungicides used in agriculture, are suspected of producing craniofacial malformations. As the results of FETAX analysis showed that Triadimefon was highly teratogenic, we studied the action of these triazoles on the development of the branchial apparatus in Xenopus, using early molecular markers and analysis of the cartilaginous-muscular elements of tadpoles. Teratogenic effects were observed, after exposure at the neurula stage, at the level of cartilages and muscles of the 1st and 2nd branchial arches. By in situ hybridization, we observed that in exposed specimens the territories of the branchial arches are perturbed. From these results, we considered the craniofacial malformations related to the influence of triazoles on the differentiation of branchial arches. Comparing the anomalies caused by triazoles and by retinoic acid (RA) led us to the conclusion that triazoles can affect endogenous RA content, as has been shown for mammals.

Topics: Animals; Branchial Region; Craniofacial Abnormalities; Fungicides, Industrial; Histological Techniques; Immunohistochemistry; In Situ Hybridization; Tretinoin; Triazoles; Xenopus laevis

Development of the frontonasal mass (FNM), branchial arches, heart, and limbs depends on neural crest-mediated epithelial-mesenchymal (E-M) interactions. Teratogenesis by retinoic acid (RA) or blockade of serotonergic (5-HT) signaling by the pan-5-HT(2) receptor antagonist, ritanserin, perturbs development of these embryonic structures. In both cases, resulting phenotypes include forebrain and olfactory placode anomalies, malformations of the face, eye and lens, as well as posterior neural tube and cardiac defects. Similar sites of malformations, together with the presence of RA response elements in the 5-HT(2B) receptor promoter, have led to the suggestion that a negative regulatory relationship may exist between RA and 5-HT(2)-mediated 5-HT signaling at sites of E-M interaction (Choi et al. 1997); however, another possibility is that RA and 5-HT act independently as opposing signals to regulate development of common embryonic targets. Together with recent evidence for opposite effects on chondrogenic differentiation in hindlimb micromass cultures (Bhasin et al. 2003a), results of the present study raise the possibility that these pathways may act as opposing signals for common targets in the mouse embryo. The RA receptors, co-factors and metabolic enzymes, and 5-HT(2B) receptors were found to be are coordinately expressed at sites of E-M interaction, including the FNM, in the embryonic day (E)10.5 mouse. Cell proliferation experiments using [(3)H]thymidine incorporation demonstrated that RA or activation of 5-HT(2B) receptors caused opposite effects in FNM explants, namely stimulation or inhibition of cell proliferation, respectively, 5-HT(2B) receptor activation did not appreciably alter patterning in FNM explants. While RA has been shown to regulate lateral patterning in the FNM (LaMantia et al. 2000), 5-HT(2B) receptor activation did not alter patterning in FNM explants. Quantification of 5-HT(2B) receptor transcripts by real-time PCR provided no evidence of negative regulation of 5-HT(2B) receptor expression by RA in FNM explants, although preliminary studies using in situ hybridization had suggested that this was a possibility in both explants and RA teratogenized embryos. Future studies using quantitative PCR may still show this to be the case in teratogenized embryos. Together with the finding of coordinate expression of 5-HT(2B )receptors and RA signaling molecules, results of the present study suggest that RA, and 5-HT mediated by 5-HT(2B )rece

Topics: Animals; Brain; Cell Proliferation; Craniofacial Abnormalities; Embryonic Development; Eye; Face; Gene Expression Regulation, Developmental; Heart; Immunohistochemistry; Keratolytic Agents; Mice; Mice, Inbred ICR; Receptor, Serotonin, 5-HT2B; Signal Transduction; Tretinoin

Endogenous retinoids are important for patterning many aspects of the embryo including the branchial arches and frontonasal region of the embryonic face. The nasal placodes express retinaldehyde dehydrogenase-3 (RALDH3) and thus retinoids from the placode are a potential patterning influence on the developing face. We have carried out experiments that have used Citral, a RALDH antagonist, to address the function of retinoid signaling from the nasal pit in a whole embryo model. When Citral-soaked beads were implanted into the nasal pit of stage 20 chicken embryos, the result was a specific loss of derivatives from the lateral nasal prominences. Providing exogenous retinoic acid residue development of the beak demonstrating that most Citral-induced defects were produced by the specific blocking of RA synthesis. The mechanism of Citral effects was a specific increase in programmed cell death on the lateral (lateral nasal prominence) but not the medial side (frontonasal mass) of the nasal pit. Gene expression studies were focused on the Bone Morphogenetic Protein (BMP) pathway, which has a well-established role in programmed cell death. Unexpectedly, blocking RA synthesis decreased rather than increased Msx1, Msx2, and Bmp4 expression. We also examined cell survival genes, the most relevant of which was Fgf8, which is expressed around the nasal pit and in the frontonasal mass. We found that Fgf8 was not initially expressed along the lateral side of the nasal pit at the start of our experiments, whereas it was expressed on the medial side. Citral prevented upregulation of Fgf8 along the lateral edge and this may have contributed to the specific increase in programmed cell death in the lateral nasal prominence. Consistent with this idea, exogenous FGF8 was able to prevent cell death, rescue most of the morphological defects and was able to prevent a decrease in retinoic acid receptorbeta (Rarbeta) expression caused by Citral. Together, our results demonstrate that endogenous retinoids act upstream of FGF8 and the balance of these two factors is critical for regulating programmed cell death and morphogenesis in the face. In addition, our data suggest a novel role for endogenous retinoids from the nasal pit in controlling the precise downregulation of FGF in the center of the frontonasal mass observed during normal vertebrate development.

Topics: Acyclic Monoterpenes; Animals; Body Patterning; Bone and Bones; Bone Morphogenetic Protein 4; Bone Morphogenetic Proteins; Cell Death; Chick Embryo; Craniofacial Abnormalities; Embryo, Mammalian; Embryo, Nonmammalian; Embryonic Structures; Face; Fibroblast Growth Factor 8; Fibroblast Growth Factors; Gene Expression Regulation, Developmental; Homeodomain Proteins; In Situ Hybridization; In Situ Nick-End Labeling; Monoterpenes; Morphogenesis; MSX1 Transcription Factor; Nose; Receptors, Retinoic Acid; Signal Transduction; Skeleton; Transcription Factors; Tretinoin

Fusion and hypoplasia of the first two branchial arches, a defect typically observed in retinoic acid (RA) embryopathy, is generated in cultured mouse embryos upon treatment with BMS453, a synthetic compound that exhibits retinoic acid receptor beta (RARbeta) agonistic properties in transfected cells. By contrast, no branchial arch defects are observed following treatment with synthetic retinoids that exhibit RARalpha or RARgamma agonistic properties. The BMS453-induced branchial arch defects are mediated through RAR activation, as they are similar to those generated by a selective pan-RAR agonist, are prevented by a selective pan-RAR antagonist and cannot be mimicked by exposure to a pan-RXR agonist alone. They are enhanced in the presence of a pan-RXR agonist, and cannot be generated in Rarb-null embryos. Furthermore, they are accompanied, in the morphologically altered region, by ectopic expression of Rarb and of several other direct RA target genes. Therefore, craniofacial abnormalities characteristic of the RA embryopathy are mediated through ectopic activation of RARbeta/RXR heterodimers, in which the ligand-dependent activity of RXR is subordinated to that of RARbeta. Endodermal cells lining the first two branchial arches respond to treatment with the RARbeta agonist, in contrast to neural crest cells and ectoderm, which suggests that a faulty endodermal regionalization is directly responsible for RA-induced branchial arch dysmorphologies. Additionally, we provide the first in vivo evidence that the synthetic RARbeta agonist BMS453 exhibits an antagonistic activity on the two other RAR isotypes.

Topics: Animals; Branchial Region; Craniofacial Abnormalities; Dimerization; Embryo, Mammalian; Endoderm; Gestational Age; In Situ Hybridization; Mice; Mice, Transgenic; Morphogenesis; Pharynx; Protein Isoforms; Receptors, Retinoic Acid; Retinoic Acid Receptor gamma; Retinoid X Receptors; Retinoids; Signal Transduction; Transcription Factors; Transgenes; Tretinoin

Fibroblast growth factor homologous factors (FHFs) have been implicated in limb and nervous system development. In this paper we describe the expression of the cFHF-4 gene during chicken craniofacial development. cFHF-4 is expressed in the mesenchyme of the frontonasal process, and in the mesenchyme and ectoderm of the mandibular processes. The expression of cFHF-4 and other genes implicated in facial patterning have been analyzed in talpid(2) embryos or in the presence of exogenous retinoic acid. Talpid(2) mutants show abnormal patterns of gene expression, including up-regulation of cFHF-4 in the developing face, which correlate with defects in cartilage formation. By contrast, expression of cFHF-4 in the developing face is strongly downregulated by teratogenic doses of all-trans retinoic acid in a dose-dependent manner. Low levels of retinoic acid that produce distal upper beak truncations do not affect cShh, c-Patched-1, or c-Bmp-2 expression in the face, but downregulate cFHF-4 in the frontonasal process.

Topics: Animals; Chick Embryo; Craniofacial Abnormalities; Facial Bones; Fibroblast Growth Factors; Gene Expression Regulation, Developmental; In Situ Hybridization; Mutation; Signal Transduction; Skull; Tretinoin

The RAR and RXR families of retinoid nuclear receptors each comprise three isotypes (alpha, beta and gamma). In vitro, RARs bind to their cognate DNA response elements as heterodimers with RXRs. Null mutations of all six isotypes have been generated. The defects displayed by RAR alpha, beta and gamma single null mutant mice are confined to a small subset of the tissues normally expressing these receptors. This discrepancy reflects the existence of a functional redundancy, since RAR double null mutants exhibit congenital malformations in almost every organ system. In particular, most of the structures derived from the mesectoderm are severely affected. Analysis of mutant mice lacking both RARs and RXRs indicates that RXR alpha:RAR gamma heterodimers are instrumental in the patterning of craniofacial skeletal elements, whereas RXR alpha:RAR alpha heterodimers may be preferentially involved in the generation of neural crest cell-derived arterial smooth muscle cells. Both RXR alpha:RAR beta and RXR alpha:RAR gamma heterodimers appear to function during the development of the ocular mesenchyme. Moreover, atavistic reptilian cranial structures are generated in RAR mutants, suggesting that the RA signal has been implicated in the modification of developmental programs in the mesectoderm during evolution.

Topics: Abnormalities, Multiple; Animals; Biological Evolution; Craniofacial Abnormalities; Dimerization; Ectoderm; Eye; Facial Bones; Mesoderm; Mice; Mice, Mutant Strains; Muscle, Smooth, Vascular; Neural Crest; Odontogenesis; Receptors, Retinoic Acid; Retinoid X Receptors; Skull; Thymus Gland; Transcription Factors; Tretinoin

Topics: Acne Vulgaris; Administration, Cutaneous; Central Nervous System; Craniofacial Abnormalities; Female; Fetus; Humans; Keratolytic Agents; Pregnancy; Teratogens; Tretinoin

Embryonic levels of retinoic acid (RA) and the response of cells to RA are critical to the normal development of vertebrates. To understand the effects of RA signaling in Fundulus heteroclitus, we exposed embryos to a range of RA concentrations of 2 h during gastrulation. Embryos exposed to low concentrations of RA (10(-10)-10(-7) M) develop normally, whereas those exposed to higher concentrations (5 x 10(-7)-10(-4) M) develop characteristic dose-dependent defects. We describe, in detail, four stages of development that represent morphological effects of RA on (1) cell death and defects in the brain, heart, and eye, (2) relative size and differentiation, (3) duplications of pectoral fins, and (4) deletions in craniofacial cartilage elements. Analysis of cartilaginous skeletal elements demonstrates distinct patterns of deletions in the neurocranium and pharyngeal skeleton in response to increasing concentrations of RA. In F. heteroclitus, RA treatment during gastrulation results in five highly consistent phenotypes, which we have incorporated into an index of embryonic RA defects. This index should be valuable in the genetic analysis of RA pathways and in evaluating chemicals that interfere with embryonic RA signaling.

Topics: Animals; Body Patterning; Cartilage; Craniofacial Abnormalities; Female; Gastrula; Killifishes; Male; Phenotype; Signal Transduction; Tretinoin

Craniofacial deformities caused by retinoic acid (RA) treatment are well known, but its mechanisms remain to be elucidated. In order to study RA-induced deformities during craniofacial morphogenesis, several different stages and regions of quail embryo in ovo were examined. It was found that the RA injection to the subgerminal space in early-stage (H.H. stage 8) affects specifically craniofacial morphogenesis. Superficially, the acrania and the exencephalia are the most remarkable deformities. Bone and cartilage staining has revealed the hypoplasia of maxilla in high frequencies. Histochemical analyses have demonstrated abnormal development and growth of the central nervous system (CNS). The expression of the Nau gene, a marker of post-mitotic neuroblasts, was also examined by in situ hybridization. Strong expression of the Nau gene in normal embryo was observed in the mantle zone in CNS in the E5 embryo. In contrast, Nau negative nervous tissues were often observed in the RA-treated E5 quail embryo, which indicates abnormal proliferation and differentiation of the neuroblasts in the CNS.

Topics: Animals; Craniofacial Abnormalities; Facial Bones; Morphogenesis; Quail; Skull; Tretinoin

Exogenous retinoic acid (RA) administered during mouse embryogenesis can alter the pattern of the axial skeleton during two developmental periods: an early window (7 to 8.5 days post-coitum; dpc) and a late window (9.5 to 11.5 dpc). Treatment during the early window results in vertebral homeotic transformations (predominantly posteriorizations) concomitant with rostral shifts in Hox gene expression, while treatment at the later window results in similar transformations without detectable alterations in Hox gene expression patterns. Mice null for retinoic acid receptor gamma (RAR gamma) exhibit axial defects, including homeosis of several vertebrae, therefore establishing a role for this receptor in normal axial specification RAR gamma null mutants are also completely resistant to RA-induced spina bifida, which occurs in wildtype embryos treated at 8.5-9.0 dpc, suggesting that this receptor specifically transduces at least a subset of the teratogenic effects of retinoids. To further investigate the role of RAR gamma in RA-induced defects during the early and late windows of retinoid-sensitive vertebral patterning, RAR gamma heterozygotes were intercrossed, pregnant females treated with vehicle or RA at 7.3, 10.5 or 11.5 dpc and full-term fetuses assessed for skeletal defects. Relative to wildtype littermates, RAR gamma null mutants treated at 7.3 dpc were markedly resistant to RA-induced embryolethality, craniofacial malformations, and neural tube defects. Furthermore, while RAR gamma null mutants were modestly resistant to certain vertebral malformations elicited by RA treatment at 7.3, they exhibited more pronounced resistance following treatment at 10.5 and 11.5 dpc. Moreover, several of the vertebral defects inherent to the RAR gamma null phenotype were abolished by RA treatment specifically at 10.5 dpc, suggesting that RAR alpha and/or RAR beta isoforms may substitute for certain RAR gamma functions, and that RAR gamma may elicit its normal effects on vertebral morphogenesis at this developmental stage.

Topics: Animals; Body Patterning; Craniofacial Abnormalities; Embryonic and Fetal Development; Female; Keratolytic Agents; Mice; Pregnancy; Receptors, Retinoic Acid; Retinoic Acid Receptor gamma; Tretinoin

The growth and differentiation factor transforming growth factor-beta2 (TGFbeta2) is thought to play important roles in multiple developmental processes. Targeted disruption of the TGFbeta2 gene was undertaken to determine its essential role in vivo. TGFbeta2-null mice exhibit perinatal mortality and a wide range of developmental defects for a single gene disruption. These include cardiac, lung, craniofacial, limb, spinal column, eye, inner ear and urogenital defects. The developmental processes most commonly involved in the affected tissues include epithelial-mesenchymal interactions, cell growth, extracellular matrix production and tissue remodeling. In addition, many affected tissues have neural crest-derived components and simulate neural crest deficiencies. There is no phenotypic overlap with TGFbeta1- and TGFbeta3-null mice indicating numerous non-compensated functions between the TGFbeta isoforms.

Topics: Abnormalities, Multiple; Animals; Bone and Bones; Cleft Palate; Craniofacial Abnormalities; Cyanosis; Ear, Inner; Embryonic Induction; Epithelium; Eye Abnormalities; Genes, Homeobox; Heart Defects, Congenital; Mesoderm; Mice; Mice, Inbred C57BL; Mice, Knockout; Phenotype; Transforming Growth Factor beta; Tretinoin; Urogenital Abnormalities

The face is one of the most intricately patterned structures in human and yet little is known of the mechanisms by which the tissues are instructed to grow, fuse, and differentiate. We undertook a study to determine if the craniofacial primordia used the same molecular cues that mediate growth and patterning in other embryonic tissues such as the neural tube and the limb. Here we provide evidence for the presence of organizer-like tissues in the craniofacial primordia. These candidate organizers express the polarizing signal sonic hedghog (shh) and its putative receptor, patched, as well as fibroblast growth factor 8 and bone morphogeneic protein 2. Shh-expressing epithelial grafts functioned as organizing tissues in a limb bud assay system, where they evoked duplications of the digit pattern. High doses of retinoic acid, which are known to truncate the growth of the frontonasal and maxillary processes and thus produce bilateral clefting of the lip and palate, inhibited the expression of shh and patched but not fgf8, in the craniofacial primordia, and abolished polarizing activity of these tissues. From these studies we conclude that the embryonic face contains signaling centers in the epithelium that participate in craniofacial growth and patterning. In addition, we discuss a novel mechanism whereby retinoids can exert a teratogenic effect on craniofacial morphogenesis independent of its effects on Hox gene expression or neural crest cell migration.

Topics: Animals; Chick Embryo; Cleft Lip; Cleft Palate; Craniofacial Abnormalities; Ectoderm; Embryonic Induction; Endoderm; Epithelium; Face; Gene Expression Regulation, Developmental; Hedgehog Proteins; Humans; Limb Buds; Maxilla; Morphogenesis; Protein Biosynthesis; Proteins; Skull; Teratogens; Trans-Activators; Transcription, Genetic; Tretinoin

The gene Fgf-3 is expressed in rhombomeres 5 and 6 of the hindbrain and has been functionally implicated in otic development. We describe new sites of expression of this gene in mouse embryos in the forebrain, the midbrain-hindbrain junction region, rhombomere boundaries, a cranial surface ectodermal domain that includes the otic placode, and in the most recently formed somite. In the early hindbrain, high levels of Fgf-3 transcripts are present in rhombomere 4. The surface ectodermal domain at first (day 8 1/2) extends laterally from rhombomeres 4 and 5 (prorhombomere B), in which neuroepithelial levels of expression are highest, to the second pharyngeal arch ventrally; at day 9, when the region of highest level of neuroepithelial Fgf-3 expression is in rhombomeres 5 and 6, the dorsal origin of the surface ectodermal domain is also at this level, extending obliquely to the otic placode and the second arch. The initially high level of Fgf-3 transcripts in the otic placode is downregulated as the placode invaginates to form the otic pit. Fgf-3 is a good marker for the epithelium of pharyngeal arches 2 and 3, and our in situ hybridization results confirm the dual identity of the apparently fused first and second arches in some retinoic acid-exposed embryos, and the fusion of the first arch with the maxillary region in others. Correlation between Fgf-3 expression and morphological pattern in craniofacial tissues of normal and retinoic acid-exposed embryos indicates that prorhombomere B, the second arch and the otic ectoderm represent a cranial segment whose structural integrity is maintained when hindbrain morphology and pharyngeal arch morphology are altered. Comparison of normal Fgf-3 expression domains with those of Fgf-4 and with the phenotype of Fgf-3-deficient mutant embryos suggests that there is some functional redundancy between Fgf-3 and Fgf-4 in otic induction and second arch development.

Topics: Abnormalities, Drug-Induced; Animals; Branchial Region; Craniofacial Abnormalities; Ear; Embryonic and Fetal Development; Female; Fibroblast Growth Factor 3; Fibroblast Growth Factors; Gene Expression Regulation, Developmental; Genetic Markers; In Situ Hybridization; Mice; Mice, Inbred C57BL; Pregnancy; Proto-Oncogene Proteins; Reference Values; Rhombencephalon; Tretinoin