Molecular characterization of 39 de novo sSMC: contribution to prognosis and genetic counselling, a prospective study.

Small supernumerary marker chromosomes (sSMCs) are structurally abnormal chromosomes that cannot be characterized by karyotype. In many prenatal cases of de novo sSMC, the outcome of pregnancy is difficult to predict because the euchromatin content is unclear. This study aimed to determine the presence or absence of euchromatin material of 39 de novo prenatally ascertained sSMC by array-comparative genomic hybridization (array-CGH) or single nucleotide polymorphism (SNP) array. Cases were prospectively ascertained from the study of 65,000 prenatal samples [0.060%; 95% confidence interval (CI), 0.042-0.082]. Array-CGH showed that 22 markers were derived from non-acrocentric markers (56.4%) and 7 from acrocentic markers (18%). The 10 additional cases remained unidentified (25.6%), but 7 of 10 could be further identified using fluorescence in situ hybridization; 69% of de novo sSMC contained euchromatin material, 95.4% of which for non-acrocentric markers. Some sSMC containing euchromatin had a normal phenotype (31% for non-acrocentric and 75% for acrocentric markers). Statistical differences between normal and abnormal phenotypes were shown for the size of the euchromatin material (more or less than 1 Mb, p = 0.0006) and number of genes (more or less than 10, p = 0.0009). This study is the largest to date and shows the utility of array-CGH or SNP array in the detection and characterization of de novo sSMC in a prenatal context.

A new highly penetrant form of obesity due to deletions on chromosome 16p11.2.

Obesity has become a major worldwide challenge to public health, owing to an interaction between the Western 'obesogenic' environment and a strong genetic contribution. Recent extensive genome-wide association studies (GWASs) have identified numerous single nucleotide polymorphisms associated with obesity, but these loci together account for only a small fraction of the known heritable component. Thus, the 'common disease, common variant' hypothesis is increasingly coming under challenge. Here we report a highly penetrant form of obesity, initially observed in 31 subjects who were heterozygous for deletions of at least 593 kilobases at 16p11.2 and whose ascertainment included cognitive deficits. Nineteen similar deletions were identified from GWAS data in 16,053 individuals from eight European cohorts. These deletions were absent from healthy non-obese controls and accounted for 0.7% of our morbid obesity cases (body mass index (BMI) >or= 40 kg m(-2) or BMI standard deviation score >or= 4; P = 6.4 x 10(-8), odds ratio 43.0), demonstrating the potential importance in common disease of rare variants with strong effects. This highlights a promising strategy for identifying missing heritability in obesity and other complex traits: cohorts with extreme phenotypes are likely to be enriched for rare variants, thereby improving power for their discovery. Subsequent analysis of the loci so identified may well reveal additional rare variants that further contribute to the missing heritability, as recently reported for SIM1 (ref. 3). The most productive approach may therefore be to combine the 'power of the extreme' in small, well-phenotyped cohorts, with targeted follow-up in case-control and population cohorts.

Sudden death: ethical and legal problems of post-mortem forensic genetic testing for hereditary cardiac diseases.

Hereditary non-structural diseases such as catecholaminergic polymorphic ventricular tachycardia (CPVT), long QT, and the Brugada syndrome as well as structural disease such as hypertrophic cardiomyopathy (HCM) and arrhythmogenic right ventricular cardiomyopathy (ARVC) cause a significant percentage of sudden cardiac deaths in the young. In these cases, genetic testing can be useful and does not require proxy consent if it is carried out at the request of judicial authorities as part of a forensic death investigation. Mutations in several genes are implicated in arrhythmic syndromes, including SCN5A, KCNQ1, KCNH2, RyR2, and genes causing HCM. If the victim's test is positive, this information is important for relatives who might be themselves at risk of carrying the disease-causing mutation. There is no consensus about how professionals should proceed in this context. This article discusses the ethical and legal arguments in favour of and against three options: genetic testing of the deceased victim only; counselling of relatives before testing the victim; counselling restricted to relatives of victims who tested positive for mutations of serious and preventable diseases. Legal cases are mentioned that pertain to the duty of geneticists and other physicians to warn relatives. Although the claim for a legal duty is tenuous, recent publications and guidelines suggest that geneticists and others involved in the multidisciplinary approach of sudden death (SD) cases may, nevertheless, have an ethical duty to inform relatives of SD victims. Several practical problems remain pertaining to the costs of testing, the counselling and to the need to obtain permission of judicial authorities.

[Genetic dermatology clinics: a practical approach]. / La consultation génétique en dermatologie: une approche pratique.

There are numerous genodermatoses and different types of transmission. Recent technological progress of molecular genetics allows to confirm or specify increasingly the clinical diagnosis and to better define the risk of recurrency. A close collaboration of dermatologists and geneticists has been established at CHUV since several years. By means of clinical examples we illustrate the organisation and procedures of this pluridisciplinary consultation which aims to optimize the clinical management of rare genetic diseases.

Confirmation of the origin of NISCH syndrome.

Neonatal ichthyosis-sclerosing cholangitis (NISCH) syndrome, a rare autosomal recessive ichthyosis syndrome characterized by scalp hypotrichosis, scarring alopecia, ichthyosis, and sclerosing cholangitis, was described for the first time in 2002. It is caused by a mutation in the gene coding for the tight junction protein claudin-1. Only four patients carrying the same mutation of the CLDN1 gene have been described until now. We report a patient presenting with the clinical characteristics of NISCH syndrome and carrying a novel mutation in the CLDN1 gene.

[Clinical and genetics aspects of Alport syndrome]. / Aspects cliniques et génétiques du syndrome d'Alport.

Microscopic haematuria of glomerular origin, without known aetiology, should raise the suspicion of Alport Syndrome IASI in children as well as in adults. The genetic mutations causing AS lie in the genes encoding for the alpha3, alpha4 and alpha5 chains of the collagen type IV, the main constituent of glomerular basement membranes (GBM). The various mutations and modes of transmission of the disease account for the heterogeneous clinical presentations. No specific treatment of AS is currently available. However, a better understanding of the GBM's ultrastructure, in particular of type IV collagen, will hopefully enable the identification of novel therapeutic targets.

SPG3A is the most frequent cause of hereditary spastic paraplegia with onset before age 10 years.

Seven families with six different SPG3A mutations were identified among 106 with autosomal dominant hereditary spastic paraplegia (HSP). Two mutations were novel (T162P, C375R). SPG3A was twice as frequent as SPG4 in patients with onset before age 10 years (31.8%). Later onset was not observed. The phenotype was pure HSP, but disease duration was longer than in non-SPG3A/SPG4 patients, leading ultimately to greater handicap.

Segregation of chromosomes in sperm of Robertsonian translocation carriers.

Robertsonian translocations are the most frequent structural chromosomal abnormalities in humans and can affect fertility, with various degrees of sperm alterations in men; or the pregnancy outcome of the carriers. The studies on meiotic segregation of chromosomes in sperm of Robertsonian translocation males find a majority of normal or balanced spermatozoa for the chromosomes related to the translocation (mean 85.42%; range 60-96.60%). Furthermore, recent studies suggest an interchromosomal effect. Studies on spermatozoa from translocation carriers, and in mouse models help the comprehension of the meiotic segregation mechanisms. Results of meiotic segregation analysis in man could be integrated in genetic counselling especially when assisted reproductive technology is required.

Absence of anti-Müllerian hormone (AMH) and M2A immunoreactivities in Sertoli cell-only syndrome and maturation arrest with and without AZF microdeletions.

BACKGROUND: Some genes identified in the AZF locus are expressed only in germinal cells; others are ubiquitous. AZF microdeletions seem to occur at the earliest stages of ontogenetic development, and one might therefore assume that Sertoli cells preserve some immature characteristics and that their immunophenotype may be modified by the existence of a molecular defect. MATERIALS AND METHODS: Two immunohistological markers of Sertoli cell immaturity [anti-Müllerian hormone (AMH) and M2A] were tested in two histopathological groups (maturation arrest at spermatocyte I stage and Sertoli cell-only syndrome). We analysed 68 testicular samples obtained from 39 patients with non-obstructive azoospermia associated or not with AZF microdeletions. RESULTS: The absence of M2A and AMH immunoreactivity in adult gonads was observed without any correlation to spermatogenetic impairment or molecular deficit in the AZF region. In the samples of these two series, Sertoli cells showed a mature phenotype for AMH and M2A markers. CONCLUSIONS: In patients with AZF microdeletions, the genotype-phenotype correlations seem to be more complex than has been suggested previously; more detailed characterization of the immunohistochemical phenotype associated with the molecular defect may be useful in understanding the spermatogenic failure mechanism.

French multi-centric study of 2000 amniotic fluid interphase FISH analyses from high-risk pregnancies and review of the literature.

This prospective and multi-centric study confirms the accuracy and the limitations of interphase FISH and shows that any cytogenetics laboratory can perform this technique. With regard to the technical approach, we think that slides must be examined by two investigators, because the scoring may be subjective. The main problem with the AneuVysion kit concerns the alpha satellite probes, and especially the chromosome 18 probe, which is sometimes very difficult to interpret because of the high variability of the size of the spots, and this may lead to false negative and uninformative cases. The best solution would be to replace these probes by locus-specific probes. Concerning clinical management, we offer interphase FISH only in very high-risk pregnancies or/and at late gestational age because of the cost of the test. We think that an aberrant FISH result can be used for a clinical decision when it is associated with a corresponding abnormal ultrasound scan. In other cases, most of the time, we prefer to wait for the standard karyotype.

[Repetitive acute pancreatitis in a late-diagnosed cystic fibrosis: prevention of relapses by octreotide in the long term]. / Poussées à répétition de pancréatite aiguë dans une mucoviscidose à révélation tardive: prévention par octréotide au long cours.

We report on the case of a 35 year-old woman who was initially admitted for acute pancreatitis in october 1995. The patient was suffering from asthma (since childhood) and diffuse abdominal pain (since adolescence). The diagnosis of cystic fibrosis was made fortuitously during a sterility evaluation. After extensive etiological screening the acute pancreatitis was considered to be a manifestation of the cystic fibrosis. Despite therapy with pancreatic enzymes, the patient continued to suffer from chronic abdominal pain. High intake of analgesics was required. Until December 1995, the patient was repeatedly admitted for episodes of acute pancreatitis. In January 1996, we initiated a preventive treatment with subcutaneous octreotide between 100 and 200 microgram, three time a day. Thereafter, there were fewer episodes of pancreatitis and the consumption of analgesics decreased. Side effects of octreotide were intermittent diarrhea and development of cholelithiasis that was complicated by biliary migration in November 1998. In June 1999, the prolonged-release form of the molecule was given without modification of the efficacy.

Meiotic segregation analysis by FISH investigation of spermatozoa of a 46,Y,der(X),t(X;Y)(qter-->p22::q11-->qter) carrier.

Chromosome analysis performed on a 30-year-old man revealed a 46,Y,der(X),t(X;Y)(qter-->p22::q11-->qter) karyotype, confirmed by fluorescence in situ hybridization (FISH). The man was of short stature, and no mental retardation was noticed; genitalia and testes were normal, as were the patient's FSH, LH, and testosterone blood levels. Sperm analysis showed azoospermia at the time of the first sampling and severe oligozoospermia, with 125,000 spermatozoa/milliliter, at the time of the second sampling. The sperm gonosomal complement of this patient and of a 46,XY donor were analyzed using multicolor FISH with X- and Y-chromosome probes. Our results clearly indicated that germinal cells carrying the translocation are able to complete the meiotic process by producing spermatozoa compatible with normal embryonic development, with more than 80% of the spermatozoa having either a Y chromosome or a der(X); however, a high level of spermatozoa with gonosomal disomies was observed. We also found a significant increase in the frequency of autosomal disomies in the carrier, which would suggest an interchromosomal effect. All previously reported cases in adult males were associated with azoospermia; testicular histological studies, performed in patients carrying the same X;Y translocation, showed spermatogenetic arrest after pachytene. To our knowledge, this is the first molecular analysis of the gonosomal complement in spermatozoa of men with a t(X;Y)(qter-->p22::q11-->qter).

Adult hypophosphatasia. Current aspects.

Hypophosphatasia is an inborn metabolic disorder in which abnormally low levels of the enzyme nonspecific alkaline phosphatase result in defective skeletal and dental mineralization (rickets, fractures, dental abnormalities) and in accumulation of the enzyme substrates (phosphoethanolamine, pyridoxal-5'phosphate and inorganic pyrophosphate). The build-up of inorganic pyrophosphate promotes the development of articular chondrocalcinosis. There are several forms of hypophosphatasia, with wide variations in severity. We report the case of a 53-year-old man with typical manifestations of moderate adulthood hypophosphatasia. Investigations in his family found the disease in a sister and two children. He had two autosomal mutations, which were transmitted recessively. Several mutations of the alkaline phosphatase gene have been identified. The genotype is correlated with the phenotype: some mutations are associated with milder forms and others with more severe forms of the disease.

AZFa deletions in Sertoli cell-only syndrome: a retrospective study.

Lack of data on the genotype-phenotype relationship in cases of AZF microdeletions is due to the limited number of histological investigations in human male infertility cases. We investigated the possibility of retrospective detection of Yq11 microdeletions by using DNA extracted from diagnostic testicular biopsies. We used histological criteria to select two series of material: 22 biopsies with Sertoli cell-only syndrome and 14 biopsies with maturation arrest at the spermatocyte I stage. Two markers, DFFRY and DAZ, were tested by nested polymerase chain reaction (PCR) in the two series. In the Sertoli cell-only syndrome series, we found four deletions affecting the DFFRY gene (18.2%). In the second series, no deletions were detected. Two conclusions may be considered, although the number of specimens analysed is limited: (i) the frequency of deletions observed in Sertoli cell-only syndrome allows us to suggest that deletion in the AZFa region may be involved in this pathology; and (ii) retrospective studies may yield some additional elements in our search for eventual genotype-phenotype relationships.

Fifteen new mutations (-195C>T, L-12X, 298-2A>G, T117N, A159T, R229S, 997+2T>A, E274X, A331T, H364R, D389G, 1256delC, R433H, N461I, C472S) in the tissue-nonspecific alkaline phosphatase (TNSALP) gene in patients with hypophosphatasia.

Hypophosphatasia is a rare inherited disorder characterized by defective bone mineralization and deficiency of serum and liver/bone/kidney-type alkaline phosphatase (L/B/K ALP) activity. We report the characterization of tissue-nonspecific alkaline phosphatase (TNSALP) gene mutations in a series of 12 families affected by severe or mild hypophosphatasia. Twenty distinct mutations were found, 5 of which were previously reported. Nine of the 15 new mutations were missense mutations (T117N, A159T, R229S, A331T, H364R, D389G, R433H, N461I, and C472S). The others were 2 nonsense mutations (L-12X and E274X), one single nucleotide deletion (1256delC), 2 mutations affecting splicing (298-2A>G, 997+2T>A), and a mutation in the major transcription start site (-195C>T). Hum Mutat 15:293, 2000.

Fortuitous detection of uniparental isodisomy of chromosome 6.

Uniparental isodisomy is defined as the inheritance of two copies of the same parental chromosome and can result in defects when it produces homozygosity for a recessive mutation or in the presence of imprinting. We describe the detection of a chromosome 6 uniparental isodisomy in a 9 year old girl, discovered during a search for an HLA identical sib. HLA typing, erythrocyte phenotyping, and genotypes of microsatellite polymorphisms were compatible with a paternal isodisomy of chromosome 6, with normal biparental origin of the other chromosomes. Paternal cells were not responsive to the patient's cells in mixed lymphocyte cultures. This fortuitous detection of a chromosome 6 isodisomy suggests that cases of chromosome 6 UPD may not be deleterious and may therefore go undetected.

Molecular analysis by fluorescence in situ hybridization of a prenatally detected de novo complex chromosomal rearrangement t(2q;3p;4q;13q).

We report one case of de novo complex chromosomal rearrangement (CCR) t(2q;3p;4q;13q) with at least five chromosomal breakpoints. This CCR was detected prenatally at 22 weeks of gestation, when mild echographic indications were disclosed during a routine examination in a female with no family history of congenital abnormalities. This observation clearly illustrates what the fluorescence in situ hybridization (FISH) technique can offer to the analysis of such rearrangements, together with standard cytogenetic techniques. No chromosomal imbalance was cytologically proved. Nevertheless, the status of the infant at birth and the disorders that he exhibited during the following months demonstrate once again that even in the absence of alarming ultrasonographic verifications and even if standard and molecular cytogenetics do not allow us to confirm evident chromosomal imbalances, genetic counselling in the case of prenatally detected de novo CCR must remain cautious.