Persistent Müllerian duct syndrome due to anti-Müllerian hormone receptor 2 microdeletions: a diagnostic challenge.

The persistent Müllerian duct syndrome (PMDS) is defined by the persistence of Müllerian derivatives in an otherwise normally virilized 46,XY male. It is usually caused by mutations in either the anti-Müllerian hormone (AMH) or AMH receptor type 2 (AMHR2) genes. We report the first cases of PMDS resulting from a microdeletion of the chromosomal region 12q13.13, the locus of the gene for AMHR2. One case involved a homozygous microdeletion of five exons of the AMHR2 gene. In the second case, the whole AMHR2 gene was deleted from the maternally inherited chromosome. The patient's paternal allele carried a stop mutation, which was initially thought to be homozygous by Sanger sequencing. Diagnostic methods are discussed, with an emphasis on comparative genomic hybridization and targeted massive parallel sequencing.

De novo variants in CNOT3 cause a variable neurodevelopmental disorder.

As a result of exome-based sequencing work performed by the DDD study, de novo variants in CNOT3 have emerged as a newly recognised cause of a developmental disorder. This paper describes molecular and clinical details of 16 probands with developmental disorders and de novo CNOT3 variants. It is the first such description of the developmental phenotype associated with CNOT3 variants. Eight of these cases were discovered as part of the DDD study, while the other eight were found as a result of large-scale sequencing work performed by other groups. A highly specific phenotype was not recognised in these 16 cases. The most consistent phenotypic features seen in subjects with de novo variants in CNOT3 were hypotonia, relatively small stature, developmental delay, behavioural problems and intellectual disability. There is no easily recognisable facial phenotype, but some common dysmorphic features such as anteverted nares, thin upper lip and low set eyebrows were shared among some of the probands. Haploinsufficiency appears to be the most likely mechanism of action, with eight cases found to have protein-truncating variants. Of the other eight cases (all missense variants), three share an amino acid substitution at the same position which may therefore represent an important functional domain.

Hereditary persistence of alpha-fetoprotein (HPAF P): review of the literature.

Alpha-fetoprotein (AFP) serum levels are raised in several clinical conditions, ranging from non-pathological conditions to malignancies. Hereditary persistence of alpha-fetoprotein (HPAFP) is a rare benign disorder with elevated AFP levels. HPAFP is described as a benign autosomal dominantly inherited condition which is not associated with any clinical disability or additional symptoms. In the past 28 years, only 19 families have been described; due to this unfamiliarity with HPAFP, elevated AFP levels are never attributed to HPAFP. However, undiagnosed HPAFP can result in inappropriate and unnecessary treatment decisions. Therefore, HPAFP should be taken into consideration in patients with unexplained elevated AFP levels, and especially in patients with urological disorders.

Does the genetic and familial background of males undertaking ICSI affect the outcome?

PURPOSE: To investigate whether the success rate of ICSI is (1) related to the etiology of infertility or (2) adversely affected by a family history of potential genetic disorders. METHODS: All men with an ICSI indication in our hospital between 1994 and 2005 were included in our cohort study. Data on the ICSI process, etiology of infertility, and family history were collected. ICSI success rates of infertility subgroups and a subgroup with a positive family history were compared to a group with unknown etiology and a negative family history. RESULTS: There was no significant difference in clinical pregnancy or delivery rates between the subgroups. Couples achieving a pregnancy underwent significantly more ICSI cycles compared to couples not achieving a pregnancy. CONCLUSION: Our results suggest that the success rate of ICSI treatment is not related to the cause of infertility or a family history positive for potential genetic disorders.

Variant Klinefelter syndrome 47,X,i(X)(q10),Y and normal 46,XY karyotype in monozygotic adult twins.

Klinefelter syndrome (KS; 47, XXY) is characterized by increased body height, hypergonadotrophic hypogonadism, and infertility. We describe a patient with a variant KS (47,X,i(Xq),Y) who has a twin brother with a 46,XY karyotype. Molecular studies showed that the twins were monozygotic. The presence of an isochromosome Xq in one of two monozygotic twins allows precise investigation of its phenotypic effect. The patient was somewhat shorter (3.5 cm) and had a smaller volume of the testes (8 vs. 18 ml) as compared to his twin brother. Furthermore he had increased gonadotrophin levels and an extreme oligoasthenoteratozoospermia (OAT). These data support the view that genes on Xp cause increased body height and genes on Xq cause infertility in KS. To our knowledge this is the first report on a heterokaryotypic monozygotic twin with a variant KS.

Vesico-ureteral reflux in children with prenatally detected hydronephrosis: a systematic review.

OBJECTIVE: To investigate the value of prenatally detected hydronephrosis (PNH) as a prognostic factor for vesico-ureteral reflux (VUR). METHODS: The MEDLINE database was searched for articles on PNH and VUR published between 1980 and 2004. A total of 18 studies were identified and reviewed for various aspects. Results were separated for primary and/or secondary VUR whenever possible, because of the different underlying pathogenic mechanisms. RESULTS: There was considerable variation between the different studies with respect to methodology and study design. One of the main discrepancies was the way in which postnatal abnormalities were ascertained: by postnatal ultrasound, voiding cystourethrogram (VCUG) alone, or combined or sequential ultrasound and VCUG. Taking these limitations into account, the published data showed there to be a mean prevalence of 15% for postnatal primary VUR after PNH. Of all patients with PNH, 53% had no postnatal anomalies, whereas 29% had other anomalies, such as duplex collecting systems. CONCLUSIONS: Of all infants with PNH, 15% had primary VUR proven postnatally and 53% had no other anomalies detected. We suggest a standardized protocol for future studies, to enable better comparison of follow-up protocols. Published by John Wiley & Sons, Ltd.

An amino-terminal DAX1 (NROB1) missense mutation associated with isolated mineralocorticoid deficiency.

CONTEXT: Mutations in DAX1 (dosage-sensitive sex reversal-adrenal hypoplasia congenita critical region on the X chromosome gene 1; NR0B1) cause X-linked adrenal hypoplasia congenita, a disease characterized by primary adrenal failure, testicular dysgenesis, and gonadotropin deficiency. Most DAX1 mutations are deletions, nonsense, or frameshift mutations that markedly impair its transcriptional activity. Missense mutations have been restricted to the carboxy-terminal domain and are associated with more variable clinical phenotypes. OBJECTIVE: The objective was to identify novel clinical phenotypes associated with DAX1 missense mutations. PATIENTS AND DESIGN: We investigated the genetic basis of isolated mineralocorticoid deficiency in a patient who carries a unique missense mutation (W105C) in the amino-terminal region of DAX1. RESULTS: The W105C DAX1 mutation in the proband was present in three asymptomatic hemizygous males, but it was not detected in the general population. Using in vitro studies of DAX1 expression and function in transfected cells, we demonstrate that the mutant DAX1 protein exhibits mild loss of function, whether studied for genes it represses or for genes it activates. Structure-function studies suggest that the W105C and other mutations in the aminoterminus are compensated by the presence of repeated LXXLL motifs that mediate DAX1 interactions with other proteins. CONCLUSIONS: We describe the first missense mutation in the aminoterminus of DAX1 and conclude that mutations in this region may be partially compensated by redundant functional domains. Mild DAX1 mutations may be a cause of isolated mineralocorticoid deficiency.

Is there an influence of X-chromosomal imprinting on the phenotype in Klinefelter syndrome? A clinical and molecular genetic study of 61 cases.

Studies on Turner syndrome suggested the presence of X-chromosomal-imprinted genes involved in social and verbal cognition. Imprinted genes on autosomes were shown to affect growth. Could imprinting of such genes on the X chromosome also influence psychomotor development and growth in men with Klinefelter syndrome (KS), who have a supernumerary X? We recorded anthropometric and psychomotor development parameters for 61 males with KS (age range 2-56 years). In 54 cases, we were able to assess intelligence quotient (IQ) and found that impaired speech - and motor developmental problems were reported significantly more often in the paternal X - than in the maternal X group (P = 0.02). We found some significant (P < 0.05) increased body size parameters in the paternal X group, which concurs with data reporting a growth promoting influence of paternally derived genes. Our results suggest X-chromosomal imprinting occurs in males with KS.

[From gene to disease; congenital adrenal hypoplasia and the DAX-1 gene]. / Van gen naar ziekte; congenitale bijnierschorshypoplasie en het DAX-1-gen.

Congenital adrenal hypoplasia is an X-linked disorder resulting in adrenocortical deficiency, failure to complete puberty due to hypogonadotrophic hypogonadism, and infertility. The disease is caused by mutations in the DAX-1 gene. The DAX-1 protein is a transcription inhibitor; it represses the transcription of other, as yet mostly unknown, genes. Mutation analysis can confirm a clinical diagnosis of congenital adrenal hypoplasia. An early diagnosis might prevent critical damage due to an adrenal crisis in an undiagnosed patient. Molecular testing can be used for carrier detection and genetic counselling.

Kinetics of meiosis in azoospermic males: a joint histological and cytological approach.

We have developed a protocol for the identification of aberrant chromosome behavior during human male meiosis up to metaphase of the secondary spermatocyte. Histological evaluation by the Johnsen score of a testicular biopsy was combined with immunofluorescence of first meiotic prophase spermatocytes, using antibodies against synaptonemal complex protein 3 (SYCP3) and the product of the ataxia telangiectasia and rad3-related gene (ATR). This combination enables accurate meiotic prophase substaging and the identification of pachytene spermatocytes with asynapsis. Furthermore, we also investigated the competence of late pachytene primary spermatocytes to complete the first meiotic division up to metaphase and of secondary spermatocytes to transform into metaphase by an in vitro challenge with okadaic acid (OA). We tested this protocol on five males with normal Johnsen scores that presented with obstructive azoospermia, five males with low Johnsen scores and non-obstructive azoospermia and six vasectomized control males of proven fertility and normal Johnsen scores. In all azoospermics, the profiling of meiotic prophase stages by immunofluorescence increases the resolving power of the Johnsen score. In both obstructive and non-obstructive azoospermic patients, relatively more leptotene meiotic prophase stages were counted compared to the controls. In non-obstructive azoospermics, a marked heterogeneity in spermatogenesis was found, after combining the results of all three approaches, pointing at functional mosaicism of the germinal epithelium. Asynaptic pachytene spermatocytes were rarely encountered. Also, when first meiotic metaphase could be induced by OA, chiasma counts were normal. In none of the non-obstructive azoospermic males did the pattern of spermatogenesis resemble that of knock-out mouse azoospermics. We conclude that this combined histological and cytological approach enables a detailed phenotypic classification of infertile males, at a level comparable to that applied for male-sterile knock-out mice with a meiotic defect. This may facilitate the identification of candidate genes for human male infertility.

Diploid/triploid mosaicism in dysmorphic patients.

Diploid/triploid mosaicism is a dysmorphology syndrome consisting of mental retardation, truncal obesity, body and/or facial asymmetry, growth retardation, hypotonia, a small phallus, malformed low-set ears and micrognathia. In 75% of the cases, the blood karyotype is normal and the diagnosis can only be established after analysis of cultured fibroblasts. This chromosome abnormality may therefore be underdiagnosed. This paper focuses on the identification of mentally retarded and dysmorphic patients with diploid/triploid mosaicism. Detailed clinical description of well-defined patients may help in deciding if a skin biopsy for karyotyping of fibroblasts should be taken. Three new cases are presented, in which DNA marker analysis showed that the extra set of chromosomes in each case was derived from the mother. We present a review of 25 cases described in the literature and we discuss the inclusion of a second polar body into an early diploid embryo as the most likely mechanism.

Split hand/split foot, iris/choroid coloboma, hypospadias and subfertility: a new developmental malformation syndrome?

This paper presents a patient with the following malformations: split hand and split foot on the left side, a hypoplastic fifth ray of the right hand and a hypoplastic first ray of the right foot with a small cleft between the first and second ray; eye abnormalities which consist of a complete iris coloboma of the left eye in an atypical position (cranio-temporal) and a coloboma of the choroid in the right eye; a glandular hypospadias and terato-zoospermia. Since split hand/split foot can be caused by mutations in the p63 gene, mutation analysis of this gene was performed. However, sequencing analysis did not reveal a mutation. This malformation complex may represent a new syndrome.

Short stature as the only presenting feature in a patient with an isodicentric (Y)(q11.23) and gonadoblastoma. A clinical and molecular cytogenetic study.

A 13-year-old phenotypically female patient presented with short stature (height SDS -2.6), but without any Turner stigmata or other dysmorphic features. Chromosome analysis showed mosaicism for an isodicentric (idic) (Y)(q11.23) containing cell line and a 45,X cell line. Subsequent gonadectomy revealed a left streak ovary and a right ovary of abnormal appearance, which on histological examination appeared to contain a gonadoblastoma. DNA analysis showed that the proposed critical region of the gonadoblastoma locus on the Y chromosome was contained within the patient's idic (Y). Conclusion. The case described here shows that patients with 45,X/46,X, isodicentric (Yp) mosaicism and a female phenotype (1) can lack external virilisation but still have a gonadoblastoma and (2) do not necessarily have Turner stigmata but can present with only short stature. This case also underlines the importance of karyotyping patients with unexplained short stature to enable gonadectomy if Y-derived material is detected.

Analysis of spermatozoa from seven ICSI males with constitutional sex chromosomal abnormalities by fluorescent in situ hybridization.

PURPOSE: The objective was to estimate the risk for subfertile males with a constitutional sex chromosomal abnormality of transmitting such a chromosome abnormality to their children, conceived by intracytoplasmic sperm injection (ICSI). METHODS: Semen samples were obtained from seven severely oligospermic ICSI candidates. Six of them had a numerical sex chromosomal abnormality, including mosaic 45,X/46,XY, mosaic 46,XY/47, XXY, 47,XXY (Klinefelter's syndrome), and 47,XYY. One male had a structural abnormality, namely, an inversion of the Y chromosome. The semen was studied by three-color fluorescent in situ hybridization (FISH) with probes specific for chromosomes 18,X, and Y. RESULTS: Chromosomal aneuploidy rates of any of the three chromosomes were significantly higher than the aneuploidy rates observed in three control samples but comparable to the rates observed in 10 ICSI candidates with oligoasthenoteratozoospermia (OAT) and a normal constitutional karyotype. CONCLUSIONS: Our data indicate that males with (mosaic) sex chromosomal abnormalities have no higher risk of producing offspring with a sex chromosomal abnormality by ICSI than OAT males with a normal karyotype.

Severe phenotypes associated with inactive ring X chromosomes.

Mental retardation and congenital malformations in individuals with small ring X chromosomes are often due to the functional disomy that results from failure of these chromosomes to undergo X inactivation. Such chromosomes either lack the XIST locus or do not express it. We have carried out genetic analysis of the ring X chromosomes from two girls with a 45,X/46,X,r(X) karyotype, mental retardation, and a constellation of abnormalities characteristic of the severe phenotype due to X disomy. In each case the ring X chromosome included an intact XIST locus which was expressed; the breakpoints were distal to DXS128, and therefore outside the XIC region; transcription analysis of alleles at the androgen receptor locus confirmed that these were inactive chromosomes. The characteristics of the XIST RNA were similar to the wild-type. Additional studies in cultured fibroblasts showed a second ring in a small percentage of the cells. The association of severe phenotype with an inactive X chromosome most likely reflects the presence of a second ring X chromosome which was active at least in some tissues during embryogenesis, but is no longer prominent in the tissues we analyzed.

[Genetics in medical practice after 2000]. / Genetica in de medische praktijk na 2000.

Within a few years the 80,000 human genes will be identified which will increase our understanding of the genetic aspects of a large number of diseases, not only the relatively rare monogenic diseases but also the more frequent multifactorial diseases such as atherosclerosis, thrombosis and schizophrenia. Pharmacogenomics will lead to particular medication that is tuned to the genetic variations of the patient. Presently this increase in knowledge in the area of DNA diagnosis and genetic counselling will lead to a continuous increase in requests for assays. The possibilities for application will depend on new technological developments such as the DNA array technology and automation. To meet the increasing number of requests for genetic counselling, genetic nurses will have to play an important role. Moreover, collaboration between clinical geneticists and other specialists will have to be further intensified.

Reproductive decisions of men with microdeletions of the Y chromosome: the role of genetic counselling.

Couples dealing with microdeletions of the Y chromosome have to make decisions about their reproductive future. Do they opt for intracytoplasmic sperm injection (ICSI), artificial insemination with donor insemination (AID) or no treatment? We analysed this decision in 28 couples and investigated the role of the counsellor and the counselling process on the final decision of the couple. Ten counsellors from six fertility clinics in The Netherlands and Belgium were interviewed about their genetic counselling of couples dealing with microdeletions. The answers to the questionnaire were converted to 11 dichotomous variables. Of the 1627 tested men in the six centres, 37 (2.3%) had a microdeletion in the AZFc region, a subregion of the AZF region on the Y chromosome important for normal spermatogenesis. The decisions of 28 of them could be analysed. Most couples chose ICSI (79%). The remaining couples chose donor insemination (7%) or refrained from treatment (14%). Several variables, including the counselling procedure, the counsellor and the available treatments in the fertility centre, influenced the decision of the couple. In conclusion, most couples dealing with microdeletions in the AZF region choose ICSI. Several aspects of the process of genetic counselling appear to be related to the final decision.

Sperm analysis in a subfertile male with a Y;16 translocation, using four-color FISH.

Sperm analysis was performed in a male with oligoasthenoteratozoospermia (OAT) and a reciprocal t(Y;16) (q11. 21;q24), using four-color FISH. Intracytoplasmic sperm injection (ICSI) treatment in this patient had resulted in the birth of one chromosomally balanced and two chromosomally normal children. To assess the risk of having a chromosomally unbalanced conception after ICSI, morphologically normal spermatozoa were studied with a set of probes allowing detection of all segregation variants. There were 51% normal or balanced sperm cells. The fraction of sperm products resulting from alternate and adjacent I segregation was 87%, 12% were products of 3:1 disjunction, and the other 1% had other types of aneuploidy. If morphologically abnormal cells were also included in the FISH analysis, nearly 90% of all the spermatozoa were unbalanced. We conclude that although the majority of males with a Y/autosome translocation are infertile due to azoospermia, our patient produces sufficient morphologically and chromosomally normal spermatozoa to have chromosomally normal or balanced offspring after ICSI. Assuming that ICSI with an unbalanced spermatozoon from this patient would result in a nonviable embryo in many cases, the combination of in vitro and subsequent in vivo selection probably results in a risk of unbalanced offspring of much less than 50%. Hence, FISH studies on the sperm of translocation carriers are useful for estimating the risk of having unbalanced offspring after ICSI and in understanding the mechanisms underlying infertility in such carriers.