Molecular cytogenetic analysis of inv dup(15) chromosomes, using probes specific for the Prader-Willi/Angelman syndrome region: clinical implications.

Twenty-seven cases of inverted duplications of chromosome 15 (inv dup [15]) were investigated by FISH with two DNA probes specific for the Prader-Willi syndrome/Angelman syndrome (PWS/AS) region on proximal 15q. Sixteen of the marker chromosomes displayed two copies of each probe, while in the remaining 11 markers no hybridization was observed. A significant association was found between the presence of this region and an abnormal phenotype (P < .01). This is the largest study to date of inv dup(15) chromosomes, that uses molecular cytogenetic methods and is the first to report a significant association between the presence of a specific chromosomal region in such markers and an abnormal phenotype.     

Molecular cytogenetic study of patients with Pallister-Killian syndrome

The Pallister-Killian syndrome (PKS) is characterized by tissue limited chromosomal mosaicism, i.e. the presence of a supernumerary metacentric chromosome [i(12p)] often confined to skin fibroblasts while the karyotype of cultured lymphocytes is normal. In the present study, chromosome painting by chromosomal in situ suppression (CISS) hybridization and interphase cytogenetic procedures employing biotinylated or digoxigenin labelled probes was carried out. These probes comprised a chromosome 12 specific library (LA 12NSO1) and chromosome 12 centromere specific -satellite (pSP12-1). They were used to analyse and quantify the presence of i(12p) in lymphocytes, granulocytes/monocytes, skin fibroblasts and buccal mucosal cells from five patients and one aborted fetus with PKS, and ten normal donors. CISS hybridization on mitotic skin fibroblasts reliably indicated the presence of i(12p) cells, even when metaphases of poor quality were included in the analysis. Two of the five patients showed i(12p) in a small proportion (0.5%) of the cultured lymphocytes too. The interphase cytogenetics procedure did not reveal the isochromosome in lymphocytes or granulocytes/monocytes in any of the patients. Two of the six patients had a twofold increase in the number of buccal mucosal cells with three hybridization signals over control values. However, for mucosal cells, methodological improvements are required. For cytogenetic diagnosis of PKS, cultured fibroblasts subjected to chromosome painting by CISS hybridization with a chromosome 12 specific library probe are recommended.     

A molecular and cytogenetic study in Finnish Prader-Willi patients

The Prader-Willi syndrome (PWS) is a developmental disorder caused by a deficiency of paternal contributions, arising from differently sized deletions, uniparental disomy or rare imprinting mutations, in the chromosome region 15q11–q13. We studied 41 patients with suspected PWS and their parents using cytogenetic and molecular techniques. Of the 27 clinically typical PWS patients, 23 (85%) had a molecular deletion that could be classified into four size categories. Only 15 of them (71%) could be detected cytogenetically. Maternal uniparental heterodisomy was observed in four cases. The rest of the patients showed no molecular defects including rare imprinting mutations. In our experience, the use of the methylation test with the probe PW71 (D15S63), together with the probe hN4HS (SNRPN), which distinguishes between a deletion and uniparental disomy, is the method of choice for the diagnosis of PWS.     

Molecular characterization of two proximal deletion breakpoint regions in both Prader-Willi and Angelman syndrome patients.

Prader-Willi syndrome (PWS) and Angelman syndrome (AS) are distinct mental retardation syndromes caused by paternal and maternal deficiencies, respectively, in chromosome 15q11-q13. Approximately 70% of these patients have a large deletion of approximately 4 Mb extending from D15S9 (ML34) through D15S12 (IR10). To further characterize the deletion breakpoints proximal to D15S9, three new polymorphic microsatellite markers were developed that showed observed heterozygosities of 60%-87%. D15S541 and D15S542 were isolated from YAC A124A3 containing the D15S18 (IR39) locus. D15S543 was isolated from a cosmid cloned from the proximal right end of YAC 254B5 containing the D15S9 (ML34) locus. Gene-centromere mapping of these markers, using a panel of ovarian teratomas of known meiotic origin, extended the genetic map of chromosome 15 by 2-3 cM toward the centromere. Analysis of the more proximal S541/S542 markers on 53 Prader-Willi and 33 Angelman deletion patients indicated two classes of patients: 44% (35/80) of the informative patients were deleted for these markers (class I), while 56% (45/80) were not deleted (class II), with no difference between PWS and AS. In contrast, D15S543 was deleted in all informative patients (13/48) or showed the presence of a single allele (in 35/48 patients), suggesting that this marker is deleted in the majority of PWS and AS cases. These results confirm the presence of two common proximal deletion breakpoint regions in both Prader-Willi and Angelman syndromes and are consistent with the same deletion mechanism being responsible for paternal and maternal deletions. One breakpoint region lies between D15S541/S542 and D15S543, with an additional breakpoint region being proximal to D15S541/S542.     

Prader-Willi syndrome: clinical genetics, cytogenetics and molecular biology

Prader-Willi syndrome (PWS) is a neurodevelopmental disorder that arises from lack of expression of paternally inherited genes known to be imprinted and located in the chromosome 15q11-q13 region. PWS is considered the most common syndromal cause of life-threatening obesity and is estimated at 1 in 10,000 to 20,000 individuals. A de novo paternally derived chromosome 15q11-q13 deletion is the cause of PWS in about 70% of cases, and maternal disomy 15 accounts for about 25% of cases. The remaining cases of PWS result either from genomic imprinting defects (microdeletions or epimutations) of the imprinting centre in the 15q11-q13 region or from chromosome 15 translocations. Here, we describe the clinical presentation of PWS, review the current understanding of causative cytogenetic and molecular genetic mechanisms, and discuss future directions for research.     

Clinical and cytogenetic studies of the Prader-Willi syndrome: Evidence of phenotype-karyotype correlation

Summary Twenty-seven patients with the presumed diagnosis of Prader-Willi syndrome (PWS) were studied clinically and cytogenetically. The patients were classified into three study groups on the basis of their clinical pictures: group 1 with 12 children meeting the strict diagnostic criteria for PWS; group 2 with nine floppy infants and young children, aged 3 years or less, without obesity and hyperphagia; and group 3 with six older children in whom some characteristic features of the syndrome were absent. High-resolution GTG banding of prometaphase chromosomes revealed del(15)(q11.1;q12) in eleven and t(15;15)(qterp11.2::q12qter) in one of the twelve group 1 patients. In group 2, four patients had del(15)(q11.1;q12), one had t(15;15)(qterp11.1::q13qter), and the remaining four had normal karyotypes. The deleted segment common to the 17 patients with the chromosome aberrations was confined to subband 15q11.2. On the other hand, all six group 3 patients had normal karyotypes. These findings indicated that when strictly defined PWS is absolutely correlated with chromosome 15 aberrations, i.e, there is a positive phenotype-karyotype correlation, and that the aberrations are etiologically related to the syndrome. Parental origin of the deleted chromosome was determined in seven patients, with OFQ-heteromorphisms being used as hereditary markers. The deleted chromosome originated from the paternal chromosome 15 in six patients and from the maternal 15 in one.     

Molecular characterization of Prader-Willi syndrome by real-time PCR

Prader-Willi syndrome (PWS) is a contiguous gene syndrome caused by the loss of function of genes situated within the 15q11-q13 region. The loss of function arises as a result of paternally derived mutations complemented by maternal imprinting. The molecular events underlying the disorder include interstitial deletions (70%), uniparental disomy (UPD) (25%), imprinting center defects (<5%), and rarely chromosomal translocations (<1%). The standard diagnosis of PWS is based on clinical observations and genetic investigations involving DNA methylation studies and fluorescence in situ hybridization (FISH) analysis. The absence of a paternal methylation pattern within 15q11 is sufficient for a diagnosis of PWS, and FISH analyses are used for the additional categorization of patients as either deletion or nondeletion. The main limitation of these investigations is that they neither determine the size of the molecular deletions nor permit detection of individuals with microdeletions in the PWS imprinting center that regulates imprinting in this region. We have designed and implemented a real-time PCR assay using genomic DNA and SYBR green I intercalating dye to determine the size of the chromosomal deletions in patients with PWS. This has been successfully performed on genomic DNA isolated from both peripheral blood leukocytes and buccal epithelial cells. Through this assay, the two common deletion classes in PWS were observed, and all results were 100% concordant with previous FISH assays performed on the same patients.     

Prader-Willi syndrome: genetic tests and clinical findings

Here we describe the genetic studies performed in 53 patients with the suspected diagnosis of Prader-Willi syndrome (PWS). PWS is characterized by neonatal hypotonia, hypogonadism, delayed psychomotor development, hyperphagia, obesity, short stature, small hands and feet, learning disabilities, and obsessive-compulsive behavior. Through the methylation analysis of the SNRPN gene, microsatellite studies of loci mapped within and outside the PWS/AS region, and fluorescence in situ hybridization (FISH) study, we confirmed the diagnosis in 35 patients: 27 with a paternal deletion, and 8 with maternal uniparental disomy (UPD). The clinical comparisons between deleted and UPD patients indicated that there were no major phenotype differences, except for a lower birth length observed in the UPD children. Our sample was composed of more girls than boys; UPD patients were diagnosed earlier than the deleted cohort (2(10/12) s. 7(9/12) years); and, in the deleted group, the boys were diagnosed earlier than the girls (5(2/12) vs. 7(8/12) years, respectively).     

Mixed gonadal dysgenesis: a syndrome of broad clinical, cytogenetic and histopathologic spectrum.

Mixed gonadal dysgenesis (MGD) is an abnormality of sexual differentiation (ASD), which encompasses an heterogeneous group of different gonadal and phenotypic abnormalities. This study describes the main clinical features found in 16 patients with MGD, relating the clinical presentation with cytogenetic evaluation and histopathological findings. For purpose of this study, MGD was considered in those patients who fulfilled the following diagnostic criteria: 1) müllerian and/or wolfflan derivatives; 2) any of the following gonadal characteristics: a) bilateral intrabdominal or scrotal immature testicular tissue; b) intrabdominal or scrotal immature testicular tissue with contralateral streak gonad. Patients were selected from an ASD study which was carried out in Medical Genetic Unit of University of Zulia (UGM-LUZ), Maracaibo, Venezuela, from 1980 to 1997. The following information was extracted from the medical history at UGM-LUZ: age, gender which patient was reared, clinical presentation, cytogenetic evaluation, laparoscopic findings and gonadal biopsy. Sixteen patients fulfilled the diagnostic criteria and ranged in age from 1.2 to 39.4 years with an average of 12.65 years. Only 5 patients were reared as males. Twelve patients consulted for genital ambiguity. Chromosomal evaluation was as following: 8 patients with 45,X/46,XY mosaicism: 5 had a 46,XY normal male karyotype and the remaining patients: 46,XX; 46,XX/46,XY and 45,X/46,Xi(Xq) karyotypes, respectively. All patients showed müllerian derivatives and occasionally wolffian derivatives. Gonadal tumors were present in 2 patients. Molecular studies of genes that govern gonadal development are necessary for a better understanding of the wide heterogeneity present in MGD.     

Psychological profile and behavioural characteristics in 12 patients with Prader-Willi syndrome.

In the present study medical, psychological and behavioural aspects in 12 patients with Prader-Willi syndrome (PWS), aged between 13 months and 28 years are presented. In half of the patients the diagnosis of PWS was made before the age of one year. The contribution of cytogenetic investigations with the detection of a 15q11 deletion in half of the PWS patients is discussed. In the evaluation of the intellectual performances IQ levels were spread between 45 and 95 with a mean IQ of 54. A specific behavioral profile in all patients with characteristic fluctuations with age was observed. Rapid changes in behaviour increased with age and after puberty, mostly related with withholding of food. The present study reinforces the importance of early diagnosis in the PWS with adequate and intelligible information towards the parents. It may prevent the dramatic obesity which leads to severe physical problems and psychological burdens in PWS adolescents and adults.     

Microduplication 22q11.2, an emerging syndrome: clinical, cytogenetic, and molecular analysis of thirteen patients

Chromosome 22, particularly band 22q11.2, is predisposed to rearrangements due to misalignments of low-copy repeats (LCRs). DiGeorge/velocardiofacial syndrome (DG/VCFS) is a common disorder resulting from microdeletion within the same band. Although both deletion and duplication are expected to occur in equal proportions as reciprocal events caused by LCR-mediated rearrangements, very few microduplications have been identified. We have identified 13 cases of microduplication 22q11.2, primarily by interphase fluorescence in situ hybridization (FISH). The size of the duplications, determined by FISH probes from bacterial artificial chromosomes and P(1) artificial chromosomes, range from 3-4 Mb to 6 Mb, and the exchange points seem to involve an LCR. Molecular analysis based on 15 short tandem repeats confirmed the size of the duplications and indicated that at least 1 of 15 loci has three alleles present. The patients' phenotypes ranged from mild to severe, sharing a tendency for velopharyngeal insufficiency with DG/VCFS but having other distinctive characteristics, as well. Although the present series of patients was ascertained because of some overlapping features with DG/VCF syndromes, the microduplication of 22q11.2 appears to be a new syndrome.     

Hypopigmentation in the Prader-Willi syndrome.

Cutaneous and ocular pigmentation were evaluated in 29 individuals with the Prader-Willi syndrome (PWS). Criteria for hypopigmentation included the presence of type I or II skin, the lightest skin type in the family by history, and iris translucency on globe transillumination. On the basis of these criteria, 48% of the PWS individuals were hypopigmented. The presence of hypopigmentation correlated with a small interstitial deletion on the proximal long arm of chromosome 15; however, this deletion was also found in individuals who did not meet the full criteria for hypopigmentation. Hairbulb tyrosinase activity and glutathione content, as well as urine cysteinyldopa excretion, were low in PWS individuals with and without hypopigmentation and did not separate these two groups. We conclude that hypopigmentation is found in a significant proportion of individuals with PWS and that the hypopigmentation may be associated with a deletion of the long arm of chromosome 15. The mechanism for the hypopigmentation is unknown.     

Two 48,XXYY patients: clinical, cytogenetic and molecular aspects

Two 48,XXYY males, a young and an adult patient, have been clinically and molecularly analysed. Clinical findings seem less severe in the young patient. This clinical difference could be mainly due to the age of the younger patient or, alternatively, the different pattern of X-inactivation observed in the two patients could play a role in the degree of the clinical manifestations.     

Del (10)p autosomal deletion syndrome: clinical, cytogenetic and gene marker studies.

A 46,XX,del(10)p13 karyotype (Paris Conference, 1971) was identified in a 5-year-old Negro girl with mental and growth retardation, brachy- and trigonocephaly, downward slanting palpebral fissures, hypotelorism, epicanthal folds, ptosis, strabismus, dysplastic nose, high-arched palate, microdontia, small low-set posteriorly rotated ears, asymmetrical thorax, wide-spaced nipples, and minor abnormalities of hands and feet. Both parents and a brother had normal karyotypes. Expression of more than 50 polymorphic gene loci determining blood groups, serum proteins and red cell enzymes was studied. The results did not permit localization of a gene locus on the deleted segment of chromosome 10. The proposita was heterozygous for the Rh and MN blood groups and for the red cell enzymes adenosine deaminase, glutamate pyruvate transaminase and esterase D. These gene loci are thereby excluded from region 10p13 yields 10pter.     

The Prader-Willi syndrome and the Angelman syndrome

The Prader-Willi syndrome and the Angelman syndrome are characterised by a complex clinical and behavioural phenotype resulting from loss of paternal or maternal expression, respectively, of genes located on the human chromosome 15q11-13. Different molecular mechanisms leading to this imbalance have been identified, including microdeletions, intragenic mutations, uniparental disomy and imprinting centre defects. Low copy repeat gene clusters are known to flank the 15q11-13 microdeletion. They predispose to unequal crossing-over events resulting in the deletion. Involvement of multiple disease genes is strongly suspected and traditional positional cloning techniques as well as animal models are used to identify the involved genes. In this review we include the present state of art and a delineation of future approach to study the candidate genes in these two syndromes.