Prenatal correction of IGF2 to rescue the growth phenotypes in mouse models of Beckwith-Wiedemann and Silver-Russell syndromes.

Beckwith-Wiedemann syndrome (BWS) and Silver-Russell syndrome (SRS) are imprinting disorders manifesting as aberrant fetal growth and severe postnatal-growth-related complications. Based on the insulator model, one-third of BWS cases and two-thirds of SRS cases are consistent with misexpression of insulin-like growth factor 2 (IGF2), an important facilitator of fetal growth. We propose that the IGF2-dependent BWS and SRS cases can be identified by prenatal diagnosis and can be prevented by prenatal intervention targeting IGF2. We test this hypothesis using our mouse models of IGF2-dependent BWS and SRS. We find that genetically normalizing IGF2 levels in a double rescue experiment corrects the fetal overgrowth phenotype in the BWS model and the growth retardation in the SRS model. In addition, we pharmacologically rescue the BWS growth phenotype by reducing IGF2 signaling during late gestation. This animal study encourages clinical investigations to target IGF2 for prenatal diagnosis and prenatal prevention in human BWS and SRS.

Epigenetic imprinting during assisted reproductive technologies: The effect of temporal and cumulative fluctuations in methionine cycling on the DNA methylation state.

Assisted reproductive technology (ART) exposes gametes and embryos to an artificial environment that does not resemble the conditions of natural conception, and therefore might change epigenetic regulation of genes that are imprinted during development. In the present review, we discuss the relationship between susceptibility of specific genes to receive an altered epigenetic composition during ART processes, possibly via alterations in the biochemical folate and methionine cycle. We provide a comprehensive view of the current state of epigenetic patterning in ART-conceived healthy children and in Angelman syndrome (AS) and Beckwith-Wiedemann syndrome (BWS) patients. We illustrate that similar genes--that is, MEST, KCNQ1OT1, and IGF2--possess an altered DNA methylation profile in animal models, ART-conceived healthy children, and AS and BWS patients. The developmental stage at which these genes receive their epigenetic imprint appears to coincide with the specific moment that ART takes place. We highlight that ART procedures affect physiological levels of enzymes and substrates involved in the folate and methionine cycle thereby altering the DNA methylation state. Moreover, although the DNA methylation rate appears to be robust: (i) temporal imbalances coinciding with defined moments of epigenetic imprinting of specific genes affect the eventual DNA methylation state of those genes and (ii) cumulative ART effects on methionine and folate cycling can alter DNA methylation rates. These observations underscore the necessity to further investigate consequences of ART treatments on the epigenetic profile.

Characterization of global loss of imprinting in fetal overgrowth syndrome induced by assisted reproduction.

Embryos generated with the use of assisted reproductive technologies (ART) can develop overgrowth syndromes. In ruminants, the condition is referred to as large offspring syndrome (LOS) and exhibits variable phenotypic abnormalities including overgrowth, enlarged tongue, and abdominal wall defects. These characteristics recapitulate those observed in the human loss-of-imprinting (LOI) overgrowth syndrome Beckwith-Wiedemann (BWS). We have recently shown LOI at the KCNQ1 locus in LOS, the most common epimutation in BWS. Although the first case of ART-induced LOS was reported in 1995, studies have not yet determined the extent of LOI in this condition. Here, we determined allele-specific expression of imprinted genes previously identified in human and/or mouse in day ∼105 Bos taurus indicus × Bos taurus taurus F1 hybrid control and LOS fetuses using RNAseq. Our analysis allowed us to determine the monoallelic expression of 20 genes in tissues of control fetuses. LOS fetuses displayed variable LOI compared with controls. Biallelic expression of imprinted genes in LOS was associated with tissue-specific hypomethylation of the normally methylated parental allele. In addition, a positive correlation was observed between body weight and the number of biallelically expressed imprinted genes in LOS fetuses. Furthermore, not only was there loss of allele-specific expression of imprinted genes in LOS, but also differential transcript amounts of these genes between control and overgrown fetuses. In summary, we characterized previously unidentified imprinted genes in bovines and identified misregulation of imprinting at multiple loci in LOS. We concluded that LOS is a multilocus LOI syndrome, as is BWS.

Chromosome 11 segmental paternal isodisomy in amniocytes from two fetuses with omphalocoele: new highlights on phenotype-genotype correlations in Beckwith-Wiedemann syndrome.

BACKGROUND: The phenotypic variability in Beckwith-Wiedemann syndrome (BWS) reflects the genetic heterogeneity of the mechanism which by default leads to the deregulation of genes located at 11p15.5. Genotype-phenotype correlation studies have demonstrated an association between omphalocoele and CDKN1C/p57 mutations or hypermethylation. Paternal uniparental disomy 11 (pUPD11) has been described only in the mosaic condition with both uniparental and biparental cell lines, and no association with omphalocoele has been pointed out. METHODS: Two cases are presented here, in which a paternal segmental UPD11 was detected by molecular investigation of amniotic fluid cell cultures after the presence of apparently isolated omphalocoele was revealed in the fetuses by ultrasound scan. Further studies were performed on additional autoptic feto-placental tissues to characterise the distribution of the uniparental cell line and to unmask any biparental lineage in order to document in more detail the as yet unreported association between omphalocoele and pUPD11. RESULTS: Results on the UPD distribution profile showed that the abdominal organs have a predominant uniparental constitution. This condition could mimic the effect of CDKN1C/p57 inactivation, causing the omphalocoele. CONCLUSION: New genotype-phenotype correlations emerge from the investigated cases, suggesting that molecular analysis be extended to all cases with fetal omphalocoele in order to establish the incidence of pUPD11 in complete BWS and in monosymptomatic/mild forms.

Bilateral hemorrhagic adrenal cysts in an incomplete form of Beckwith-Wiedemann syndrome: MRI and prenatal US findings.

Benign hemorrhagic adrenal cysts are a cause of subdiaphragmatic mass in the fetus and neonate with Beckwith-Wiedemann syndrome. Characteristic imaging features on ultrasonography, color Doppler, and magnetic resonance imaging help differentiate adrenal hemorrhage from neonatal neuroblastoma and help avoid unnecessary surgery in these patients. Bilateral adrenal hemorrhage is self-limiting, and spontaneous resolution is the usual outcome. This report presents this rare condition with prenatal ultrasonographic and magnetic resonance imaging findings and reviews the differential diagnosis of neonatal adrenal masses.

Beckwith-Wiedemann syndrome.

Beckwith-Wiedemann syndrome (BWS) is a clinically heterogeneous overgrowth syndrome associated with an increased risk for embryonal tumor development. BWS provides an ideal model system to study epigenetic mechanisms. This condition is caused by a variety of genetic or epigenetic alterations within two domains of imprinted growth regulatory genes on human chromosome 11p15. Molecular studies of BWS have provided important data with respect to epigenotype/genotype-phenotype correlations; for example, alterations of Domain 1 are associated with the highest risk for tumor development, specifically Wilms' tumor. Further, the elucidation of the molecular basis for monozygotic twinning in BWS defined a critical period for imprint maintenance during pre-implantation embryonic development. In the future, such molecular studies in BWS will permit enhanced medical management and targeted genetic counseling.

Unusual prenatal presentation of Beckwith-Wiedemann syndrome.

When Beckwith-Wiedemann syndrome (BWS) is detected prenatally, it is usually on the basis of macroglossia, exomphalos or enlarged kidneys. We describe a case that presented as gross hepatomegaly and a suspected enlarged pancreas at 20 weeks' gestation, with none of the usual features.

Oppositely imprinted genes p57(Kip2) and igf2 interact in a mouse model for Beckwith-Wiedemann syndrome.

Beckwith-Wiedemann syndrome (BWS) is a clinically variable disorder characterized by somatic overgrowth, macroglossia, abdominal wall defects, visceromegaly, and an increased susceptibility to childhood tumors. The disease has been linked to a large cluster of imprinted genes at human chromosome 11p15.5. A subset of BWS patients has been identified with loss-of-function mutations in p57(KIP2), a maternally expressed gene encoding a G(1) cyclin-dependent kinase inhibitor. Some patients display loss of imprinting of IGF2, a fetal-specific growth factor that is paternally expressed. To understand how the same disease can result from misregulation of two linked, but unrelated, genes, we generated a mouse model for BWS that both harbors a null mutation in p57(Kip2) and displays loss of Igf2 imprinting. These mice display many of the characteristics of BWS, including placentomegaly and dysplasia, kidney dysplasia, macroglossia, cleft palate, omphalocele, and polydactyly. Some, but not all, of the phenotypes are shown to be Igf2 dependent. In two affected tissues, the two imprinted genes appear to act in an antagonistic manner, a finding that may help explain how BWS can arise from mutations in either gene.

Nephrogenic rests, nephroblastomatosis, and associated lesions of the kidney.

The fetal kidney is formed by the development of nephrons from fetal metanephric blastema surrounding the ureteric bud. The fetal renal tissue matures into normal renal parenchyma during gestation, but, occasionally, fetal tissue persists into infancy as microscopic foci called nephrogenic rests. Nephrogenic rests are found in approximately 1% of infant kidneys at autopsy. Nephrogenic rests are associated with an increased risk of Wilms tumor, and it is theorized that nephrogenic rests undergo neoplastic change into Wilms tumor. Fortunately, this transformation occurs in less than 1% of young children with nephrogenic rests. Nephrogenic rests are associated with many syndromes, including Beckwith-Wiedemann syndrome, hemihypertrophy, and sporadic aniridia. Children with identifiable syndromes, once diagnosed, should be screened for the development of Wilms tumor. Nephrogenic rests are associated with other lesions such as multilocular cystic nephroma and multicystic dysplasia, usually without malignant complications.

Prenatal detection of a congenital pancreatic cyst and Beckwith-Wiedemann syndrome.

We report a case of congenital pancreatic cyst detected prenatally by ultrasound in a fetus with evidence for a diagnosis of Beckwith-Wiedemann syndrome (BWS). Neonatal hypoglycaemia was prevented. The cyst was managed by internal drainage. This is the second reported case of BWS associated with pancreatic cystic dysplasia and the first time that this association has been detected prenatally. Differential diagnosis of cystic abdominal lesions occurring in utero should take pancreatic cysts into consideration. This case suggests that pancreatic cysts should be included in the BWS phenotype.

Transactivation of Igf2 in a mouse model of Beckwith-Wiedemann syndrome.

The gene IGF2, which encodes a fetal insulin-like growth factor, is imprinted, so only one of two parental copies of the gene is expressed. The altered expression of IGF2 has been implicated in Beckwith-Wiedemann syndrome, a human fetal overgrowth syndrome, which is characterized by overgrowth of several organs and an increased risk of developing childhood tumours. We have introduced Igf2 transgenes into the mouse genome by using embryonic stem cells, which leads to transactivation of the endogenous Igf2 gene. The consequent overexpression of Igf2 results in most of the symptoms of Beckwith-Wiedemann syndrome, including prenatal overgrowth, polyhydramnios, fetal and neonatal lethality, disproportionate organ overgrowth including tongue enlargement, and skeletal abnormalities. These phenotypes establish Igf2 overexpression as a key determinant of Beckwith-Wiedemann syndrome.

The cell type-specific IGF2 expression during early human development correlates to the pattern of overgrowth and neoplasia in the Beckwith-Wiedemann syndrome.

Overstimulation by insulin-like growth factor II is implied in several overgrowth conditions and childhood cancers. We have therefore studied spatial and temporal expression patterns of the insulin-like growth factor II gene (IGF2) and the insulin-like growth factor type 1 receptor gene during normal human development (5.5 to 23.0 weeks postfertilization). The set of cell types with the most abundant IGF2 expression correlated strikingly to the organomegaly and tumor predisposition of the Beckwith-Wiedemann syndrome. Intrauterine growth and postnatal organ weights of a prematurely born child with a full-blown syndrome are presented. The cell type-specific IGF2 expression of these organs and of multifocal Wilms' tumors from two other children affected by the Beckwith-Wiedemann syndrome were also studied. The results clarify and extend previous findings concerning human prenatal IGF2 expression and are consistent with a short range overstimulatory role of locally produced IGF II ensuing after the first trimester in the Beckwith-Wiedemann syndrome.

Somatic mosaicism for partial paternal isodisomy in Wiedemann-Beckwith syndrome: a post-fertilization event.

Genomic imprinting has been implicated in the aetiology of an overgrowth cancer-prone syndrome, the Wiedemann-Beck-with syndrome (WBS). We have demonstrated uniparental disomy (UPD) for paternal chromosome 11p markers in 5 out of 25 sporadic cases (20%). Delineation of the extent of the disomy region may help in understanding the mechanism and the stage, meiotic or mitotic, of disomy formation in this disease and in associated tumours. Our current studies in WBS patients with seventeen 11p and one 11q markers reveal paternal isodisomy, not heterodisomy, in the five cases. For one case we demonstrate unambiguously that partial isodisomy for 11p and somatic mosaicism for UPD resulted from a post-fertilization event. The restriction of isodisomy to part of 11p in another case, and somatic mosaicism for UPD in three other cases, suggest a mitotic recombinational event that must have occurred after fertilization. Mosaic phenotypes reflect the timing of their origin and the fate of the cells involved, as well as the cell-specific pattern of imprinting. Somatic mosaicism for UPD in four cases may thus explain the incomplete forms of WBS. The association of hemihypertrophy in sporadic WBS and even some cases of isolated hemihypertrophy. This is in agreement with a recent report of paternal isodisomy for 11p markers in a patient with hemihypertrophy, Wilms' tumour and adrenocortical carcinoma. Moreover, the risk of developing a tumour seems higher (50%) for patients with paternal 11p UPD than for WBS patients in general (7.5%).

Placentomegaly with massive hydrops of placental stem villi, diploid DNA content, and fetal omphaloceles: possible association with Beckwith-Wiedemann syndrome.

Marked placental hydrops is generally associated with hydatidiform mole. Diagnosis of hydatidiform mole requires both villous hydrops and trophoblast hyperplasia. This report describes four cases with massive hydrops of placental stem villi without associated trophoblast hyperplasia. All four had diploid DNA content by flow cytometry. Fetal omphalocele was present in three; and one had diagnostic Beckwith-Wiedemann syndrome (BWS). In two others, there were pathologic features suggestive of BWS. The fourth fetus had multiple anomalies by ultrasound; autopsy examination of the fragmented fetus failed to disclose additional pathology. The association of massive placental hydrops involving stem villi, fetal omphalocele, and diploid DNA content is unusual. These fetal and placental findings may suggest possible BWS in some cases and allow for antenatal diagnosis of affected fetuses, clinical evaluation of additional family members, and planning for neonatal care.