Teratogenic action and embryo lethality of AY 9944R. Prevention by a hypercholesterolemia-provoking diet.

Embryomortality and teratogenesis provoked by inhibitors of cholesterol synthesis are well demonstrated. Teratogenic action is particularly reflected by holoprosencephalies, but also by uro-genital abnormalities. A hypercholesterolemia-provoking diet has been shown to be completely effective for preventing holoprosencephaly, but only partially so for preventing the uro-genital malformations and fetal mortality. It is thus possible that the two types of abnormalities are governed by different mechanisms. In addition, the diet itself, whose hypercholesterolemic effect is considerable, has certain disadvantages. It seems to have a certain effect on fetal mortality and could be responsible for several uro-genital malformations. This deserves further study.     

The spectrum of median craniofacial dysplasia

Given the multiple permutations in craniofacial malformations, classification of median craniofacial dysplasia or midline Tessier no. 0 to 14 clefts has been difficult and disjointed. In this review, the authors present a summary of normal embryology, prior terminology, and their proposed new classification system. Median craniofacial dysplasia has tissue agenesis and holoprosencephaly at one end (the hypoplasias), frontonasal hyperplasia and excessive tissue (the hyperplasias) at the other end, and abnormal splitting or clefting and normal tissue volume (dysraphia) occupying the middle portion of the spectrum. These three distinct subclassifications have different forms of anomalies within their groups.     

Holoprosencephaly: clinical, anatomic, and molecular dimensions

Holoprosencephaly is addressed under the following headings: alobar, semilobar, and lobar holoprosencephaly; arrhinencephaly; agenesis of the corpus callosum; pituitary abnormalities; hindbrain abnormalities; syntelencephaly; aprosencephaly/atelencephaly; neural tube defects; facial anomalies; median cleft lip; minor facial anomalies; single maxillary central incisor; holoprosencephaly-like phenotype; epidemiology; genetic causes of holoprosencephaly; teratogenic causes of holoprosencephaly; SHH mutations; ZIC2 mutations; SIX3 mutations; TGIF mutations; PTCH mutations; GLI2 mutations; FAST1 mutations; TDGF1 mutations; and DHCR7 mutations.     

The Ubiquitin E3 Ligase NOSIP Modulates Protein Phosphatase 2A Activity in Craniofacial Development

Holoprosencephaly is a common developmental disorder in humans characterised by incomplete brain hemisphere separation and midface anomalies. The etiology of holoprosencephaly is heterogeneous with environmental and genetic causes, but for a majority of holoprosencephaly cases the genes associated with the pathogenesis could not be identified so far. Here we report the generation of knockout mice for the ubiquitin E3 ligase NOSIP. The loss of NOSIP in mice causes holoprosencephaly and facial anomalies including cleft lip/palate, cyclopia and facial midline clefting. By a mass spectrometry based protein interaction screen we identified NOSIP as a novel interaction partner of protein phosphatase PP2A. NOSIP mediates the monoubiquitination of the PP2A catalytic subunit and the loss of NOSIP results in an increase in PP2A activity in craniofacial tissue in NOSIP knockout mice. We conclude, that NOSIP is a critical modulator of brain and craniofacial development in mice and a candidate gene for holoprosencephaly in humans.     

The pathology of trisomy 13 syndrome

Summary Anatomical and histopathological findings in 12 cases of trisomy 13 syndrome (nine with classic full trisomy and three with trisomy 13 and an unbalanced Robertsonian 13/13 translocation) are reported. Emphasis is on the brain defects, cardiovascular anomalies, and histological organ dysplasia. Eight patients showed abnormal development of the forebrain and midline facial structures (holoprosencephaly). Cardiovascular malformations were invariably present, the leading malformation being an infundibular ventricular septal defect often in combination with dextroposition of the aorta and abnormalities of the semilunar valves. Histological abnormalities giving evidence of organ dysplasia were observed in the central nervous system, eyes, pancreas, kidneys, and ovaries. Mild cystic renal dysplasia was a constant feature. Foci of persistent nodular renal blastema were found in six cases. The pancreatic dysplasia appears to be pathognomonic for trisomy 13. These observations illustrate the importance of pathological studies in the recognition of chromosome abnormalities and, more specifically, of trisomy 13 syndrome. Based on autopsy data, trisomy 13 can be diagnosed — or ruled out — with certainty, even in the absence of karyotyping.     

Limb reduction defects in humans associated with prenatal isotretinoin exposure.

Retinoic acid has long been used to induce limb reductions defects in experimental animal studies. No limb malformations, however, have been reported among malformed retinoic acid-exposed human fetuses from case reports or epidemiologic studies. We report a child and a fetus with limb reduction malformations following maternal use of isotretinoin (13-cis-retinoic acid) during the first trimester of pregnancy. The child had a unilaterally absent clavicle and nearly absent scapula, with a short humerus and short, synostotic forearm bones. He also had ventriculomegaly and developmental delay, minor dysmorphic facial features, and a short sternum with a sterno-umbilical raphe. The fetus had a unilaterally absent thumb with normal proximal bony structures. Other findings included hydrocephalus, craniofacial anomalies, thymic agenesis, supracristal ventricular septal defect, single umbilical artery, anal and vaginal atresia, and urethral agenesis with dysplastic, multicystic kidneys. Although the limb malformations were quite dissimilar, a number of anomalies that are frequently found among isotretinoin-exposed fetuses/infants were present in both cases. This increases the probability that retinoic acid caused these limb defects, but a causal association cannot be conclusively drawn on the basis of these two retrospective case reports.     

Teratogenic effect of AY 9944 in rats: importance of the day of administration and maternal plasma cholesterol level

An inhibitor of cholesterol synthesis, AY 9944 (trans-1,4-bis(2-chlorobenzylaminomethyl) cyclohexane dihydrochloride) is teratogenic. A single dose of AY 9944 (50 mg/kg or 75 mg/kg) given to Wistar pregnant rats on the second, fourth, sixth, seventh, or eighth day of gestation induced malformations such as holoprosencephaly. They were often limited to isolated pituitary agenesis. The highest percentage of holoprosencephalic fetuses was found when AY 9944 was given on the fourth day of gestation. Whatever the dose and the day of administration, the lower the maternal plasma cholesterol level, the more frequent were holoprosencephalic fetuses. Therefore, it is suggested that the decrease in maternal plasma cholesterol level is at least one of the factors provoking holoprosencephaly.     

Potentialities of magnetic resonance imaging in the complex of prenatal radiation diagnosis of fetal malformations

The purpose of the study was to investigate the potentialities of magnetic resonance imaging (MRI) in the complex of prenatal radiation diagnosis of fetal malformations. Twenty-eight female patients with suspected fetal malformations were examined. Ultrasound study was supplemented by MRI according to a specially developed protocol. Various fetal CNS malformations were diagnosed. These included the Arnold-Chiari syndrome, the Dandy-Walker syndrome, occlusive hydrocephaly, lobular holoprosencephaly, porencephaly, diaphragmatic hernias, anomalies of the abdomen and retroperitoneal space, as well as anomalies of facial structures, including median clefts, and dacryocystocele. The use of MRI in the complex prenatal radiation diagnosis makes it possible to visualize fetal malformation more clearly, contributes to the more adequate prediction of the outcome of pregnancy and to the choice of a management policy for a female patient.     

Mutations in Hedgehog Acyltransferase (Hhat) Perturb Hedgehog Signaling, Resulting in Severe Acrania-Holoprosencephaly-Agnathia Craniofacial Defects

Holoprosencephaly (HPE) is a failure of the forebrain to bifurcate and is the most common structural malformation of the embryonic brain. Mutations in SHH underlie most familial (17%) cases of HPE; and, consistent with this, Shh is expressed in midline embryonic cells and tissues and their derivatives that are affected in HPE. It has long been recognized that a graded series of facial anomalies occurs within the clinical spectrum of HPE, as HPE is often found in patients together with other malformations such as acrania, anencephaly, and agnathia. However, it is not known if these phenotypes arise through a common etiology and pathogenesis. Here we demonstrate for the first time using mouse models that Hedgehog acyltransferase (Hhat) loss-of-function leads to holoprosencephaly together with acrania and agnathia, which mimics the severe condition observed in humans. Hhat is required for post-translational palmitoylation of Hedgehog (Hh) proteins; and, in the absence of Hhat, Hh secretion from producing cells is diminished. We show through downregulation of the Hh receptor Ptch1 that loss of Hhat perturbs long-range Hh signaling, which in turn disrupts Fgf, Bmp and Erk signaling. Collectively, this leads to abnormal patterning and extensive apoptosis within the craniofacial primordial, together with defects in cartilage and bone differentiation. Therefore our work shows that Hhat loss-of-function underscrores HPE; but more importantly it provides a mechanism for the co-occurrence of acrania, holoprosencephaly, and agnathia. Future genetic studies should include HHAT as a potential candidate in the etiology and pathogenesis of HPE and its associated disorders.     

A trisomy 13 case with Robertsonian translocation presenting with atypical findings

A trisomy 13 case with Robertsonian translocation presenting with atypical findings: Trisomy 13 is an autosomal trisomy caused by the presence of an extra copy of chromosome 13. Anomalies associated with this syndrome are severe mental retardation, coloboma, hypotonia, skeletal anomalies, midline anomalies, facial defects, holoprosencephaly, cardiac defects, omphalocele and polydactyly. Here we report a case of trisomy 13 with Robertsonian translocation, 160 day old, presenting with atypical findings like posterior fusion defect of the vertebra, hyperplasia of the right lobe of the liver, dilatation at pelvicalyxial system, scoliosis and complex heart disease including cardiomyopathy.     

Selected clinical research involving the central nervous system

This paper updates three clinical research projects involving the central nervous system. Discussions of conditions with encephalocele include several associations: encephalocele/craniostenosis, transsphenoidal encephalocele/hypothalamic-pituitary dysfunction, encephalocele/oculo-auriculo-vertebral spectrum, and encephalocele/frontonasal dysplasia. The relationship between oculo-auriculo-vetebral spectrum with encephalocele and frontonasal dysplasia with epibulbar dermoids and ear tags is also discussed and an explanation for encephalocele formation in the Apert syndrome is provided. Studies of the central nervous system in Apert syndrome indicate that distortion ventriculomegaly is common, but progressive hydrocephalus occurs infrequently. A recurrent pattern of abnormalities was discerned consisting of megalencephaly, gyral abnormalities, and defects of the corpus callosum and limbic structures. Five neuropathologic studies lend further support to this pattern of CNS anomalies in the Apert syndrome. In a study of holoprosencephaly, eight principles governing associated facial dysmorphism were derived. Each diagnostic category was shown to have its own frequency and range of holoprosencephalic faces. Some categories, such as del(13q), have narrow ranges; others, such as trisomy 13 syndrome, have broad ranges. However, no broad diagnostic range is known to include agnathia-holoprosencephaly and other severe forms of facial dysmorphism without agnathia. Absent maxillary incisors and a single maxillary central incisor are extremely common in holoprosencephaly with severe facial dysmorphism and may occur on occasion as a striking microform of holoprosencephaly, most commonly in the autosomal dominant form.     

Holoprosencephaly and midline facial anomalies: redefining classification and management.

Holoprosencephaly encompasses a series of midline defects of the brain and face. Most cases are associated with severe malformations of the brain which are incompatible with life. At the other end of the spectrum, however, are patients with midline facial defects and normal or near-normal brain development. Although some are mentally retarded, others have the potential for achieving near-normal mentality and a full life expectancy. The latter patients do not fit clearly into the previously defined classification system. Proposed is a new classification focusing on those patients with normal or lobar brain morphology but with a wide range of facial anomalies. The classification aids in planning treatment. Coupled with CT scan findings of the brain and a period of observation, patients unlikely to thrive can be distinguished from those who will benefit from surgical intervention. Repair of the false median cleft lip and palate may suffice in patients with moderate mental retardation. Patients exhibiting normal or near-normal mentality with hypotelorbitism and nasomaxillary hypoplasia can be treated with a simultaneous midface advancement, facial bipartition expansion, and nasal reconstruction.     

Heterozygous mutations in SIX3 and SHH are associated with schizencephaly and further expand the clinical spectrum of holoprosencephaly

Schizencephaly (SCH) is a clinically and etiologically heterogeneous cerebral malformation presenting as unilateral or bilateral hemispheric cleft with direct connection between the inner and outer liquor spaces. The SCH cleft is usually lined by gray matter, which appears polymicrogyric implying an associated impairment of neuronal migration. The majority of SCH patients are sporadic, but familial SCH has been described. An initial report of heterozygous mutations in the homeobox gene EMX2 could not be confirmed in 52 patients investigated in this study in agreement with two independent SCH patient cohorts published previously. SCH frequently occurs with additional cerebral malformations like hypoplasia or aplasia of the septum pellucidum or optic nerve, suggesting the involvement of genes important for the establishment of midline forebrain structures. We therefore considered holoprosencephaly (HPE)-associated genes as potential SCH candidates and report for the first time heterozygous mutations in SIX3 and SHH in a total of three unrelated patients and one fetus with SCH; one of them without obvious associated malformations of midline forebrain structures. Three of these mutations have previously been reported in independent patients with HPE. SIX3 acts directly upstream of SHH, and the SHH pathway is a key regulator of ventral forebrain patterning. Our data indicate that in a subset of patients SCH may develop as one aspect of a more complex malformation of the ventral forebrain, directly result from mutations in the SHH pathway and hence be considered as yet another feature of the broad phenotypic spectrum of holoprosencephaly.     

Perspectives on holoprosencephaly: Part I. Epidemiology, genetics, and syndromology

This paper provides an updated, comprehensive, critical review of the epidemiology, genetics, and syndromic aspects of holoprosencephaly and is divided into four parts. In the first part, epidemiologic aspects are discussed under the following headings: prevalence, temporal trends, socioeconomic status, exposure to environmental teratogens, maternal and paternal ages, pregnancy histories, and birth weights. The second part analyzes the facial phenotypes because the genetic and syndromic aspects of holoprosencephaly cannot be understood without knowledge of facial variability and its meaning. Topics discussed include cyclopia, ethmocephaly, cebocephaly, median cleft lip, and less severe facial dysmorphism. The third section, on genetics, analyzes associated anomalies, chromosomal and non-chromosomal holoprosencephaly, family studies, twin studies, genetics of nonsyndromic holoprosencephaly, and recurrence risks. The final section on syndromology summarizes 48 conditions in which some degree of holoprosencephaly may be a feature.     

Distal 4p microdeletion in a case of Wolf-Hirschhorn syndrome with congenital diaphragmatic hernia

BACKGROUND: Wolf-Hirschhorn syndrome (WHS) is a well-known genetic condition characterized by typical facial anomalies, midline defects, skeletal anomalies, prenatal and postnatal growth retardation, hypotonia, mental retardation, and seizures. Affected patients with a microdeletion on distal 4p present a milder phenotype that lacks congenital malformations. WHS is rarely associated with congenital diaphragmatic hernia (CDH), and only 8 cases are reported in the literature. In almost all cases of CDH and WHS a large deletion of the short arm of chromosome 4 is present. CASE: A microdeletion of 2.6 Mb on distal 4p associated with CDH and multiple congenital malformations (i.e., cleft palate) is reported for the first time. CONCLUSIONS: Such a microdeletion should prompt a molecular study for WHS when in a fetus/newborn with CDH the association with cleft lip/palate and typical facial appearance (flat facial profile, hypertelorism) is found.     

Comparative ultrasound and magnetic resonance diagnosis of fetal CNS malformations

The study was undertaken to optimize the diagnosis of fetal CNS and facial malformations, by using a complex of ultrasound (US) and magnetic resonance imaging (MRI) studies. A hundred and forty-four fetuses with suspected CNS and facial malformations were examined. The US study conducted by a specially developed protocol was supplemented by MRI (48 fetuses) also made by a specially developed protocol. Various fetal CNS malformations, such as neural tube defects, congenital endbrain malformations, cystic lesions, tumors, ventricular complex anomalies, defects of the face and eyes, multiple defects, including CNS and facial anomalies, were detected. With MRI, the diagnosis was changed in 33% of cases. The application of a complex of US and MRI studies enhances the efficiency of diagnosis of congenital CNS and facial malformations in the fetus. MRI in the diagnosis of fetal CNS and facial malformations has a number of advantages and should be used if there is some difficulty in establishing a diagnosis when an US study is performed.     

Holoprosencephaly in a fetal macaque (Macaca nemestrina) following weekly exposure to ethanol

Previous studies in rodents have indicated that the facial changes of fetal alcohol syndrome (FAS) closely resemble those of a mild form of holoprosencephaly. In order to examine this relationship in non-human primates, we evaluated a 133-day gestation macaque (Macaca nemestrina) with holoprosencephaly, median cleft lip and palate, and encephalocele. The mother had been given ethanol once per week (1.8 g/kg body weight) from weeks 2 to 19 postconception. Diagnosis of holoprosencephaly was made following ultrasound evaluation for polyhydramnios and delivery of the female fetus by caesarean section. Another fetus of identical age was delivered by caesarean section for use as a control. Both fetuses were studied by anthropometric, gross, radiographic, and histologic techniques. In the fetus exposed to alcohol, no extracranial anomalies were identified and the karyotype was normal. The brain was micrencephalic, with absent olfactory bulbs, tracts, optic nerves and chiasma, fused frontal lobes, and a single, dilated lateral ventricle; a parietooccipital encephalocele consisted of thin, dysplastic cortex bordering the ventricle; the cerebellum was dysplastic and superiorly displaced. Within the craniofacial complex, anophthalmia was bilateral; premaxillary components were absent, palatal shelves separate, the maxillae closeset, and the ethmoid bone small and deformed. Most of these defects are similar to those encountered in humans with holoprosencephaly and support the hypothesis of shared etiologic and pathogenetic relations between the facial anomalies of fetal alcohol syndrome and holoprosencephaly.     

The role of sonic hedgehog in normal and abnormal craniofacial morphogenesis.

There is growing evidence that implicates a role for Sonic hedgehog (SHH) in morphogenesis of the craniofacial complex. Mutations in human and murine SHH cause midline patterning defects that are manifested in the head as holoprosencephaly and cyclopia. In addition, teratogens such as jervine, which inhibit the response of tissues to SHH, also produce cyclopia. Thus, the loss of SHH signaling during early stages of neural plate patterning has a profound influence of craniofacial morphogenesis. However, the severity of these defects precludes analyses of SHH function during later stages of craniofacial development. We have used an embryonic chick system to study the role of SHH during these later stages of craniofacial development. Using a combination of surgical and molecular experiments, we show here that SHH is essential for morphogenesis of the frontonasal and maxillary processes (FNP and MXPs), which give rise to the mid- and upper face. Transient loss of SHH signaling in the embryonic face inhibits growth of the primordia and results in defects analogous to hypotelorism and cleft lip/palate, characteristics of the mild forms of holoprosencephaly. In contrast, excess SHH leads to a mediolateral widening of the FNP and a widening between the eyes, a condition known as hypertelorism. In severe cases, this widening is accompanied by facial duplications. Collectively, these experiments demonstrate that SHH has multiple and profound effects on the entire spectrum of craniofacial development, and perturbations in SHH signaling are likely to underlie a number of human craniofacial anomalies.