Weaver Syndrome-Associated EZH2 Protein Variants Show Impaired Histone Methyltransferase Function In Vitro.

Weaver syndrome (WS) is a rare congenital disorder characterized by generalized overgrowth, macrocephaly, specific facial features, accelerated bone age, intellectual disability, and susceptibility to cancers. De novo mutations in the enhancer of zeste homolog 2 (EZH2) have been shown to cause WS. EZH2 is a histone methyltransferase that acts as the catalytic agent of the polycomb-repressive complex 2 (PRC2) to maintain gene repression via methylation of lysine 27 on histone H3 (H3K27). Functional studies investigating histone methyltransferase activity of mutant EZH2 from various cancers have been reported, whereas WS-associated mutations remain poorly characterized. To investigate the role of EZH2 in WS, we performed functional studies using artificially assembled PRC2 complexes containing mutagenized human EZH2 that reflected the codon changes predicted from patients with WS. We found that WS-associated amino acid alterations reduce the histone methyltransferase function of EZH2 in this in vitro assay. Our results support the hypothesis that WS is caused by constitutional mutations in EZH2 that alter the histone methyltransferase function of PRC2. However, histone methyltransferase activities of different EZH2 variants do not appear to correlate directly with the phenotypic variability between WS patients and individuals with a common c.553G>C (p.Asp185His) polymorphism in EZH2.

The utility of quantitative methylation assays at imprinted genes for the diagnosis of fetal and placental disorders.

An imbalance of imprinted gene expression within 11p15.5 is observed in Beckwith-Wiedemann syndrome (BWS), as well as in a variety of placental abnormalities including complete hydatidiform mole (CHM), placental mesenchymal dysplasia (PMD) and triploidy. To facilitate the diagnosis of epigenetic errors and chromosomal imbalance of 11p15.5, we validated a pyrosequencing assay to measure methylation at KvDMR1 using blood samples from 13 BWS cases, 8 of which showed reduced methylation as compared to control blood. An imbalance between maternal and paternal genomes as is found in triploidy, CHM or PMD was also associated with altered KvDMR1 methylation. A reciprocal pattern of methylation was obtained in the triploid cases by assaying the proximal 11p15.5 ICR associated with H19. To distinguish chromosome 11 specific alterations from whole genome imbalance, other imprinted differentially methylated regions (DMRs) can be utilized. Thus, pyrosequencing assays for DMRs associated with SGCE, SNRPN, and MEST were also compared for their utility in diagnosing parental imbalance in placental samples. While each of these assays could successfully distinguish parental origin of triploidy, SGCE showed the clearest separation between groups. The combined use of a chromosome 11p15.5 assay (e.g. KvDMR1 or H19-ICR) and non-chromosome 11 assay (e.g. SGCE) provides a potentially valuable diagnostic tool in the rapid screening of methylation errors in placental disorders. These results also show the maintenance of imprinting status at these loci in the human placenta, even in the presence of abnormal pathology.

Sonographic 'molar tooth' sign in the diagnosis of Joubert syndrome.

The characteristic imaging finding common to Joubert syndrome and related disorders is the 'molar tooth' sign. The prenatal diagnosis of Joubert syndrome using both ultrasound and fetal magnetic resonance imaging (MRI) in families with an affected child has been reported previously. We report two cases in which the molar tooth sign was identified by sonography at 26 + 4 weeks and at 20 + 6 weeks, respectively, prior to fetal MRI or genetic testing. In both cases the finding was subsequently confirmed on fetal MRI. As definitive prenatal genetic testing may not be conclusive in Joubert syndrome, the ability to identify the molar tooth sign sonographically before 24 weeks provides a valuable adjunct to prenatal diagnosis.

Early sonographic evaluation for fetal growth delay and congenital malformations in pregnancies complicated by insulin-requiring diabetes. National Institute of Child Health and Human Development Diabetes in Early Pregnancy Study.

OBJECTIVE: It has been reported that early fetal growth retardation may be a useful marker for congenital malformations in diabetic pregnancies. To test this hypothesis, diabetic and nondiabetic women were sonographically evaluated during the first trimester. RESEARCH DESIGN AND METHODS: Fetal crown-rump lengths were measured sonographically at least once during the first 15 wk of pregnancy in 329 nondiabetic and 312 diabetic women. Of these, 289 nondiabetic and 269 diabetic women had sonograms before 10 wk of gestation and 283 nondiabetic and 269 diabetic women had sonograms between 10 and 15 wk of gestation. Early fetal growth delay was defined as a sonographic gestational age of greater than or equal to 6 days less than menstrual gestational age. RESULTS: The mean crown-rump lengths at 8 wk were 17.9 +/- 4.6 mm in the diabetic and 18.7 +/- 4.9 mm in the nondiabetic groups (P = 0.13). At 12 wk, the mean fetal crown-rump length was 58.5 +/- 8.8 mm for diabetic subjects and 60.6 +/- 8.7 mm for nondiabetic subjects (P = 0.04). Between 5 and 9 wk, 28 of 289 (9.7%) fetuses of nondiabetic subjects, 34 of 259 (13.1%) normal fetuses of diabetic subjects, and 2 of 10 (20%) malformed fetuses of diabetic subjects demonstrated growth delay (P = 0.31, normal vs. malformed diabetic). Between 10 and 15 wk of gestation, 28 of 283 (9.9%) fetuses of nondiabetic subjects, 32 of 256 (12.5%) normal fetuses of diabetic subjects, and 4 of 13 (30.8%) malformed fetuses of diabetic subjects demonstrated growth delay (P = 0.06, normal vs. malformed diabetic). Early fetal growth delay did not predict a reduced birth weight at term. CONCLUSIONS: Among insulin-dependent diabetic subjects who were moderately well controlled at conception, statistically significant but mild early fetal growth delay was present but did not appear to be useful clinically in predicting congenital malformations. Recommendations that growth delay demonstrated on early ultrasound be used as a predictor of congenital malformation require careful reexamination.

Intrinsic auricular muscles and auricular form.

Plical folding of the cartilagenous ear plate is largely determined by the insertions of four extrinsic and seven intrinsic auricular muscles. Dissection of ten external auricles from anencephalic fetuses of six or more months' gestation was done. The transversus and obliquus auricular muscles found in association with the antihelical fold were evaluated. Four auricles were of normal form and had the transversus and obliquus auricular muscles with typical insertions in the cartilage. Six of the ten auricles had a simplified form, with no antihelical folding in five and minimal development in one. Fibers from neither the transversus nor the obliquus auricular muscles were identified in these six ears. The results of these dissections strongly suggest that antihelical folding is determined by the presence of functioning transversus and obliquus auriculae muscles. Similarly it is proposed that normal folding of the auricular cartilage is dependent on the presence of functioning auricular muscles. In evaluation of the child with absence of normal ear form, these findings suggest that the unusual shape is due to abnormal placement and/or function of the intrinsic and extrinsic auricular muscles. These changes may be a subtle sign of neuromuscular weakness, abnormal muscle insertion or absence of muscle.

New multiple congenital anomalies syndrome in a stillborn infant of consanguinous parents and a prediabetic pregnancy.

We report on a term stillborn female infant with multiple congenital anomalies (MCA) which have not previously been reported as occurring together. The malformations include a first and second branchial arch sequence, ectopia cordis with congenital heart defect, caudal "regression" sequence with absent sacrum and hypoplastic right femur, ectrodactyly, left radial abnormality, islet cell hyperplasia, and skin lesions. The pregnancy was complicated by abdominal cramping with exercise, heavy vaginal bleeding, maternal obesity, and a normal screening glucose tolerance test at 6 months gestation. The infant was born to 20-year-old G3P1SAB2 Mexican-American parents who are first cousins. There was strong maternal family history of adult-onset diabetes. The malformations have some findings in common with those seen in infants of diabetic mothers. Structural defects similar to, but not inclusive of, those in our infant have been reported in 2 sibs born to a prediabetic mother with a first cousin marriage as well as in focal dermal hypoplasia. Parental consanguinity is suggestive of an autosomal recessive disorder. Alternatively, it may represent a combined multifactorial effect making the conceptus more sensitive to metabolic teratogens and thus placing it at increased risk for disruption of normal development.

Mixed sclerosing bone dysplasia, small stature, seizure disorder, and mental retardation: a syndrome?

We present a 40-year-old man with mental retardation, short stature, minor anomalies, and seizures, who was found to have osteopoikilosis with melorheostosis (mixed sclerosing bone dysplasia, MSBD). Cytogenetic findings of a low level trisomy 8 mosaicism were not confirmed by fluorescence in situ hybridization (FISH) of fibroblast cells. To our knowledge, the association of MSBD and mental retardation has not been previously reported.

Additional case of craniofacial and digital anomalies as reported by Harrod et al.

In 1977 Harrod et al. [BD:OAS XIII (3B): 111-115] reported 2 brothers with an unusual syndrome of mental retardation, unusual facial appearance, large protruding ears, arachnodactyly, hypogenitalism, failure to thrive, and minor anomalies. We report on a 46-year-old man with striking resemblance to the children described by Harrod who also has secondary megacolon and varicose veins, suggesting a connective tissue disorder.

Sutural exostoses, rib hyperostoses, craniosynostosis, mental retardation with focal fat deposition: Proteus syndrome?

We report on a 3-year-old boy with cartilaginous exostoses of the cranial sutures, rib hyperostosis, macrocephaly, metopic craniostenosis, epibulbar dermoid, hyperpigmented macules on the neck, focal fat deposition, and mild mental retardation with marked speech delay. Several of these manifestations were reported previously as an "unknown" by Thanos et al. [1977], with additional clinical information and a diagnosis of Proteus syndrome [Cohen, 1993].

Limb body wall complex: I. Pathogenesis.

Twenty-five fetuses with limb body wall complex (LBW complex) were evaluated. The diagnosis was based on two out of three of the following: exencephaly/encephalocele with facial clefts; thoraco- and/or abdominoschisis; and limb defect. Ninety-five percent (24/25) of the fetuses had associated internal structural defects. In 72% (18/25) the internal defects have been recognized as being secondary to vascular disruption. Concordance was not found between the side and location of the body wall defect versus the limb, internal, and cranial defects. In 85% there was evidence for persistence of the extraembryonic coelom by examination of the placenta. In this same group (85%) there was persistence of the ectodermal-amnion margin, with the amnion being continuous with the skin of the body wall defect. In 40% (10/25) there were tags and amniotic adhesions at other sites. There was no difference in the types or incidence of internal defects between those with and those without amniotic bands. The abnormalities in this collection and experimental animal models support vascular disruption during 4-6 weeks' gestation as an etiology for LBW complex. There is disruption and loss of existing tissues, persistence of embryonic structures, and secondary malformations. Persistence of the extraembryonic coelom may lead to the typical amniotic tags, ring constrictions, and adhesions seen in some specimens.

Noonan phenotype associated with neurofibromatosis.

We report on four patients with neurofibromatosis and manifestations of Noonan syndrome including short stature, ptosis, "midface hypoplasia," apparently short webbed neck, learning disabilities, and weakness. No family history of neurofibromatosis was present in any case. Average paternal and maternal age at birth was 37 and 28 years, respectively, suggestive of a new mutation. The presence of a distinct phenotype and hypotonia in these patients with neurofibromatosis is suggestive of a new separate disorder.

Health care concerns and guidelines for adults with Down syndrome.

Down syndrome (DS) is the most common cause of mental retardation in North America, yet little information is available on the natural history of DS in adults. We report on significant medical problems of adults with DS (DS adults) residing in a British Columbia provincial residential center, Woodlands, over the 12-year period from 1981 through 1992. Prospective, yearly health care reviews on 38 DS adults are summarized according to age. Group 1 consists of 18 middle-aged DS adults less than 50 years old, and group 2 comprises 20 elderly DS adults 50 years and older. Significant health problems in all DS adults include untreated congenital heart anomalies (15. 8%), acquired cardiac disease (15.8%), pulmonary hypertension (7.8%), recurrent respiratory infections/aspiration leading to chronic pulmonary interstitial changes (30%), complications from presenile dementia/Alzheimer-type disease (42%), adult-onset epilepsy (36.8%), osteoarthritic degeneration of the spine (31.6%), osteoporosis with resultant fractures of the long bones (55%) or vertebral bodies (30%), and untreated atlantooccipital instability (7.9%). Acquired sensory deficits are significant problems including loss of vision due to early onset of adult cataracts (50%), recurrent keratitis (21%) or keratoconus (15.8%), and significant hearing loss (25%). Behavioral problems (50%), loss of cognitive abilities, and onset of symptoms of Alzheimer disease (group 1: 5.5%; group 2: 75%) pose ongoing challenges for care. In conclusion, the quality of life for adults with DS can be improved by routine, systematic health care screening to identify treatable diseases that may be missed because of poor communication or confusion due to Alzheimer disease.

Trisomy 18 and a constitutional maternal translocation (2;18).

Parental chromosomes are usually not analyzed in cases of trisomy 18 because the extra 18 is assumed to have arisen through a meiotic nondisjunctional event. We report on a case of a trisomy 18 and a maternal translocation (2;18)(q34;q12).

In utero thrombosis and neonatal gangrene in an infant of a diabetic mother.

We report an infant of a diabetic mother (IDM) with in utero brachial artery thrombosis and neonatal gangrene to illustrate that there may be an increased risk for arterial as well as venous thrombosis in IDMs. The diagnosis of brachial artery thrombosis was made by using Doppler sonography flow studies and was confirmed with autopsy. The postnatal period was complicated by aortic and mesenteric artery thrombosis, with subsequent necrotizing enterocolitis, renal infarction, and death. Gangrene of a limb presenting at birth is rare, with 32 individuals reported in the literature, including this patient. Twenty-two percent (7/32) of the infants with peripartum limb gangrene were IDMs. This implies a marked increase in arterial thrombosis in IDMs over the general population. Changes in coagulation factors have been reported in newborn IDMs with poor control of maternal diabetes. Increased clotting and decreased fibrinolysis found in diabetics may lead to arterial thrombosis in IDMs in utero and postnatally. Use of anticoagulants in at-risk infants should be considered to prevent further thrombosis postnatally. Additionally, IDMs may be at increased risk for thrombotic complications from umbilical artery catheter. In utero thrombosis of the brachial artery may be one mechanism which leads to limb reduction defects in IDMs.

Deformations in infants of diabetic and control pregnancies.

Severe and mild deformations in newborn infants of insulin dependent diabetic mothers (IDDMs) and control mothers were evaluated with respect to the types of anomalies and previously hypothesized constraint factors. Factors evaluated were gestational length, birth weight, corrected birth weight for gestation (weight ratio), maternal height and parity, and severe deformations. Newborn infants from 81 control and 133 insulin dependent diabetic pregnancies were recruited periconceptually as part of a larger study of diabetes in early pregnancy. Examinations were done at 48 to 72 hours of life by one examiner blinded to maternal status using a checklist of major and minor deformations and malformations. Mild deformations were found to be common and were present in 84% of newborn infants. Severe deformations occurred in three (1.4%) IDMs, with two of three newborn infants having major malformations involving the CNS and/or musculoskeletal system which affected fetal movement. There was no significant difference between IDMs and control newborn infants with respect to the number with deformations; however, fetal macrosomia was not present in study participants. Using the entire cohort, a significantly greater number of deformations were present in newborn infants with a gestation > 36 weeks (P < 0.001), birth weight > 3,000 g (P < 0.001), and weight ratio > or = 1.2 (P = 0.05). There was no significant association with primiparous mothers or women with a height < 165 cm and the presence of deformations. For gestational age and birth weight, mild deformations were apparent only after 33 weeks gestation (P << 0.001) and/or birth weights of 2.0 kg or more (P << 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Trisomy 4 in a fetus with cyclopia and other anomalies.

We report on an 18-week fetus with cyclopia, alobar holoprosencephaly, complex congenital heart defect, anal atresia, oligosyndactyly, cystic hygroma, and skeletal abnormalities with trisomy 4. Structural anomalies were detected on routine ultrasound of the pregnancy of a 17-year-old G3 P1 TAB1 woman with sickle cell trait. Trisomy 4 conceptuses usually miscarry in the first trimester. We are aware of no other reports of a fetus with trisomy 4 and cyclopia. Causal association of chromosome abnormalities and holoprosencephaly sequence may be more apparent in embryos and early fetuses than term fetuses because of poor viability of affected conceptuses.

Partial duplication of 3q (q25.1-->q26.1) without the Brachmann-de Lange phenotype.

Partial duplications of chromosome 3 have previously been reported to have phenotypic characteristics similar to Brachmann-de Lange syndrome (BDLS). We present the case of a 13-year-old girl with an apparent duplication in the 3q25.1-->q26.1 region but none of the manifestations commonly seen in BDLS. The chromosome 3 duplication was confirmed with a FISH painting probe of the involved region. These results suggest that the region critical for Brachmann-de Lange syndrome is not within the duplicated region of 3q25.1-->q26.1.

Craniosynostosis associated with partial duplication of 15q and deletion of 2q.

We report on an infant with multiple congenital anomalies including complex craniosynostosis associated with an unbalanced karyotype, 46,XY,-2,+der(2),t(2;15)(q37;q26)pat. The previous report of a child with cloverleaf skull and partial duplication of 15q25----qter and the Man-on-Mouse Homology map suggests that a critical segment for synostosis of sutures may be in this region.

Limb-body wall complex: II. Limb and spine defects.

Limb defects from 25 fetuses with limb-body wall (LBW) complex were evaluated to determine the mechanism of limb damage. The limb defects could be divided into 3 pathogenetic groups: (1) secondary to disruption of embryonic vessels and surrounding tissue (84%), (2) secondary to amniotic bands or adhesions (16%), and (3) deformation versus hemorrhage (44% with club feet), with some fetuses having more than one pathogenetic mechanism causing limb defects. The hypothesis that the majority of limb defects resulted from disruption of embryonic vessels was supported by the following findings: 96% of the LBW complex fetuses had limb defects; the lower limbs were at greater risk of damage than the upper limbs (28% rt arm, 52% lt arm, 60% rt leg, 72% lt leg); there was a distal to proximal progression of limb damage in 92% of the fetuses; statistical analysis of comparing the location of the most severe limb defect and the body wall defect did not find concordance between the side (p = 1.0) and the region (p = 0.18) of the body wall defect; and limb defects found in the human specimens were similar to those produced in experimental animals following disruption of embryonic vessels at a corresponding gestation. In the specimens with amniotic band related limb defects (16%), the most likely pathogenesis is mechanical rupture through the amnion in the presence of a persistent extraembryonic coelom or from adhesion of the amnion to necrotic embryonic tissue after the initial disruptive event. Club feet were present in 44% and may be due either to disruption of embryonic vessels or to deformation. Further studies are needed to resolve this question.

Constriction bands and limb reduction defects in two newborns with fetal ultrasound evidence for vascular disruption.

Most structural anomalies attributed to vascular disruption have been inferred, though not proven, to be the result of disruptive events in utero. We report on 2 pregnancies with ultrasound evidence of disruptive events resulting in terminal limb "reduction" defects with constriction bands and other anomalies. In the first patient a fetal ultrasound study at 12 weeks post-LMP demonstrated a monochorionic (MC) twin pregnancy with a nonviable co-twin and no evidence of amniotic bands. At birth, there was a left cleft lip and palate, and terminal limb "reduction" defects with ring constrictions of the left hand and both feet. In the second patient, a routine fetal ultrasound study at 18 weeks post-LMP identified a subhepatic cyst which subsequently resolved. Fetal ultrasound examination and neonatal computed tomography (CT) scan of the liver were consistent with a hepatic infarct due to emboli from the umbilical vein. At birth, patient 2 had acrosyndactyly of the left hand with ring constrictions of the digits and reduction of the left big toe. There was no evidence of abnormal amnion. Postnatal development has been normal in both cases. We present ultrasound evidence supporting the hypothesis that vascular disruption from death of a co-twin or from in utero embolic infarcts can cause: 1) terminal limb "reduction" defects and possibly cleft lip and palate; and 2) ring constrictions similar to those of "amniotic band disruption sequence" in the absence of an abnormal amnion. Serial pregnancy ultrasound studies are recommended for evaluation of the development of fetal structural anomalies following ultrasound evidence of a disruptive event in utero.