Ovarian reserve in patients with FMR1 gene premutation and the role of fertility preservation.

INTRODUCTION: Women with premutation (PM) of the FMR1 gene may suffer from reduced ovarian reserve or even premature ovarian insufficiency (POI). We studied hormonal and ultrasound ovarian reserve, fertility and fertility preservation outcomes in these patients. PATIENTS AND METHOD: Retrospective cohort study of 63 female FMR1 premutation carriers. RESULTS: Sixty-three female patients bearing an FMR1 premutation were included. Median age was 30 years [26.5-35]. Median number of CGG triplets was 83 [77.2-92]. Before diagnosis of PM, 19 women (30%) had had in all 35 pregnancies, resulting in 20 births, including 7 affected children. After diagnosis of PM, 17 women (26.1%) had in all 23 pregnancies, at a median age of 34.5 years [32.2-36.0]: 2 after preimplantation genetic diagnosis, 3 after oocyte donation, 18 spontaneously, and 5 ending in medical termination for Fragile X syndrome. Thirty-three patients (52.4%) had POI diagnosis (median age, 30 years [27-34]) with median FSH level 84 IU/L [50.5-110] and median AMH level 0.08 ng/ml [0.01-0.19]. After POI diagnosis, 8 women had in all 9 pregnancies: 3 following oocyte donation, and 6 spontaneous in 5 women (15.1%). Eight of the 9 pregnancies resulted in a live birth (including 2 affected children) and 1 in medical termination for trisomy 13. The median age of the 30 patients without POI was 31 years [25.2-35.0]. Thirteen women (20.6%) underwent fertility preservation, at a median age of 29 years [24-33]: FSH 7.7 IU/L [6.8-9.9], AMH 1.1 ng/mL [0.95-2.1], antral follicle count 9.5 [7.7-14.7]. A median 15 oocytes [10-26] were cryopreserved in a median 2 cycles [1-3]. At the time of writing, no oocytes had yet been thawed for in vitro fertilization. CONCLUSIONS: This study shows the importance of early fertility preservation after diagnosis of FMR1 premutation in women, due to early deterioration of ovarian reserve. Genetic counseling is essential in these patients, as spontaneous pregnancies are not uncommon, even in cases of impaired ovarian reserve, and can lead to birth of affected children.

Different Nuclear Architecture in Human Sperm According to Their Morphology.

Human sperm parameters serve as a first step in diagnosing male infertility, but not in determining the potential for successful pregnancy during assisted reproductive technologies (ARTs) procedures. Here, we investigated the relationship between sperm head morphology at high magnification, based on strict morphologic criteria, and the nuclear architecture analyzed by fluorescence in situ hybridization (FISH). We included five men. Two of them had an elevated high-magnification morphology score of 6 points (Score 6) indicating high fertility potential, whereas three had a low score of 0 points (Score 0), indicating low fertility potential. We used FISH to study the inter-telomeric distance and the chromosomal territory area of chromosome 1 (Chr. 1). We then compared these two parameters between subjects with high and low scores. FISH data analysis showed that the inter-telomeric distance (ITD) and chromosomal territory area (CTA) of Chr. 1 were significantly higher in subjects with low scores (score 0) than high scores (score 6). Our results suggest that (i) there is a link between nuclear architecture and sperm head abnormalities, particularly vacuoles; and (ii) it is possible to select spermatozoa with normal nuclear architecture, which might indirectly explain the positive ART outcomes observed with this technique.

Skewed X-chromosome inactivation drives the proportion of DNAAF6-defective airway motile cilia and variable expressivity in primary ciliary dyskinesia.

BACKGROUND: Primary ciliary dyskinesia (PCD) is a rare airway disorder caused by defective motile cilia. Only male patients have been reported with pathogenic mutations in X-linked DNAAF6, which result in the absence of ciliary dynein arms, whereas their heterozygous mothers are supposedly healthy. Our objective was to assess the possible clinical and ciliary consequences of X-chromosome inactivation (XCI) in these mothers. METHODS: XCI patterns of six mothers of male patients with DNAAF6-related PCD were determined by DNA-methylation studies and compared with their clinical phenotype (6/6 mothers), as well as their ciliary phenotype (4/6 mothers), as assessed by immunofluorescence and high-speed videomicroscopy analyses. The mutated X chromosome was tracked to assess the percentage of cells with a normal inactivated DNAAF6 allele. RESULTS: The mothers' phenotypes ranged from absence of symptoms to mild/moderate or severe airway phenotypes, closely reflecting their XCI pattern. Analyses of the symptomatic mothers' airway ciliated cells revealed the coexistence of normal cells and cells with immotile cilia lacking dynein arms, whose ratio closely mirrored their XCI pattern. CONCLUSION: This study highlights the importance of searching for heterozygous pathogenic DNAAF6 mutations in all female relatives of male PCD patients with a DNAAF6 defect, as well as in females consulting for mild chronic respiratory symptoms. Our results also demonstrate that about one-third-ranging from 20% to 50%-normal ciliated airway cells sufficed to avoid severe PCD, a result paving the way for gene therapy.

Chromosomal segregation analysis and HOST-based sperm selection in a complex reciprocal translocation carrier.

INTRODUCTION: Complex chromosomal rearrangements (CCRs) involve two or more chromosomes and at least three breakpoints. Due to their complexity, they are associated with a high number of unbalanced gametes, whose fertilization is often incompatible with viable fetal development. Preimplantation genetic diagnosis (PGD) is usually offered to those patients and typically shows modest results considering the high number of unbalanced embryos. We previously showed that a sperm selection process using the hypo-osmotic swelling test (HOST) allows for an 83% reduction in the proportion of unbalanced spermatozoa (US) in male rearrangements carriers. This is the first report of the use of this procedure in a CCR carrier. CASE DESCRIPTION: We report on the case of a 36-year-old male t(4;7;14)(q12;p21;q11.2) carrier who presented to our center for infertility. Sperm fluorescent in situ hybridization showed an 88% proportion of unbalanced spermatozoa. After hypo-osmotic incubation and selection of spermatozoa with a specific flagellar conformation, the proportion of unbalanced spermatozoa dropped to 15%. DISCUSSION: In the present case, we show that it is possible to select chromosomally balanced prior to in vitro fertilization in male CCR carriers. This technique has the potential of increasing the proportion of euploid embryos and therefore the chances of healthy pregnancy and birth.

Unraveling a case of 46,XY DSD due to 17ß-Hydroxysteroid Dehydrogenase type 3 mutations at the age of 49.

17-ß Hydroxysteroid dehydrogenase type 3 (17ß-HSD3) is an enzyme transforming Delta 4 androstenedione into testosterone. It is involved in the early development of the male genital tract. In this case report, we describe a 46,XY Difference of Sexual Development (DSD) individual with a female phenotype, primary amenorrhea, facial dysmorphia and mental retardation. Gene sequencing using a panel of genes involved in DSD revealed two heterozygous loss-of-function mutations in the HSD17B3 enzyme. Furthermore, a microarray analysis revealed a 37Mb segmental 3p duplication and a recurrent 16p13.11 microduplication. The large 3p duplication is responsible for her mental retardation and her facial dysmorphia. Interestingly, HSD17B3 mutations were identified only in adulthood, at the age of 49. Furthermore, the patient's severe mental retardation and facial dysmorphia are due to genetic abnormalities different from the ones involved in her DSD.

Prevalence and characteristics of gonadoblastoma in a retrospective multi-center study with follow-up investigations of 70 patients with Turner syndrome and a 45,X/46,XY karyotype.

Introduction: A gonadectomy is currently recommended in patients with Turner syndrome (TS) and a 45,X/46,XY karyotype, due to a potential risk of gonadoblastoma (GB). However, the quality of evidence behind this recommendation is low. Objective: This study aimed to evaluate the prevalence of GB, its characteristics, as well as its risk factors, according to the type of Y chromosomal material in the karyotype. Methods: Our study within French rare disease centers included patients with TS and a 45,X/46,XY karyotype, without ambiguity of external genitalia. Clinical characteristics of the patients, their age at gonadectomy, and gonadal histology were recorded. The regions of the Y chromosome, the presence of TSPY regions, and the percentage of 45,X/46,XY mosaicism were evaluated. Results: A total of 70 patients were recruited, with a median age of 29.5 years (21.0-36.0) at the end of follow-up. Fifty-eight patients had a gonadectomy, at a mean age of 15 ± 8 years. GB was present in nine cases. Two were malignant, which were discovered at the age of 14 and 32 years, without metastases. Neither the percentage of XY cells within the 45,X/46,XY mosaicism nor the number of TSPY copies was statistically different in patients with or without GB (P = 0.37). However, the entire Y chromosome was frequent in patients with GB (6/9). Conclusions: In our study, including a large number of patients with 45,X/46,XY TS, the prevalence of gonadoblastoma is 12.8%. An entire Y chromosome appears as the main risk factor of GB and should favor early gonadectomy. Significant statement: About 10% of patients with TS have a karyotype containing Y chromosomal material: 45,X/46,XY. Its presence is related to the risk of GB. Therefore, a prophylactic gonadectomy is currently recommended in such patients. However, the quality of evidence is low. Our objective was to evaluate the prevalence of GB according to the type of Y-chromosomal material. We found a prevalence of GB of 12.8% in a cohort of 70 TS patients. No sign of hyperandrogenism was observed. The entire Y chromosome was the most frequent type of Y-material in patients with GB. As the prognosis of these tumors was good, a delay of surgery might be discussed.

Whole exome sequencing in a cohort of familial premature ovarian insufficiency cases reveals a broad array of pathogenic or likely pathogenic variants in 50% of families.

OBJECTIVE: To study the diagnostic yield, including variants in genes yet to be incriminated, of whole exome sequencing (WES) in familial cases of premature ovarian insufficiency (POI). DESIGN: Cross-sectional study. SETTING: Endocrinology and reproductive medicine teaching hospital departments. PATIENTS: Familial POI cases were recruited as part of a nationwide multicentric cohort. A total of 36 index cases in 36 different families were studied. Fifty-two relatives were available, including 25 with POI and 27 affected who were nonaffected. Karyotype analysis, FMR1 screening, single nucleotide polymorphism array analysis, and WES were performed in all subjects. INTERVENTIONS: None. MAIN OUTCOME MEASURES: The primary outcome was a molecular etiology, as diagnosed by karyotype, FMR1 screening, single nucleotide polymorphism array, and WES. RESULTS: A likely molecular etiology (pathogenic or likely pathogenic variant) was identified in 18 of 36 index cases (50% diagnostic yield). In 12 families, we found a pathogenic or likely pathogenic variant in a gene previously incriminated in POI, and in 6 families, we found a pathogenic or likely pathogenic variant in new candidate genes. Most of the variants identified were located in genes involved in cell division and meiosis (n = 11) or DNA repair (n = 4). CONCLUSIONS: The genetic etiologic diagnosis in POI allows for genetic familial counseling, anticipated pregnancy planning, and ovarian tissue preservation or oocyte preservation. Identifying new genes may lead to future development of therapeutics in reproduction based on disrupted molecular pathways. CLINICAL TRIAL REGISTRATION NUMBER: NCT 01177891.

Neurodevelopmental phenotype in 36 new patients with 8p inverted duplication-deletion: Genotype-phenotype correlation for anomalies of the corpus callosum.

Inverted duplication deletion 8p [invdupdel(8p)] is a complex and rare chromosomal rearrangement that combines a distal deletion and an inverted interstitial duplication of the short arm of chromosome 8. Carrier patients usually have developmental delay and intellectual disability (ID), associated with various cerebral and extra-cerebral malformations. Invdupdel(8p) is the most common recurrent chromosomal rearrangement in ID patients with anomalies of the corpus callosum (AnCC). Only a minority of invdupdel(8p) cases reported in the literature to date had both brain cerebral imaging and chromosomal microarray (CMA) with precise breakpoints of the rearrangements, making genotype-phenotype correlation studies for AnCC difficult. In this study, we report the clinical, radiological, and molecular data from 36 new invdupdel(8p) cases including three fetuses and five individuals from the same family, with breakpoints characterized by CMA. Among those, 97% (n = 32/33) of patients presented with mild to severe developmental delay/ID and 34% had seizures with mean age of onset of 3.9 years (2 months-9 years). Moreover, out of the 24 patients with brain MRI and 3 fetuses with neuropathology analysis, 63% (n = 17/27) had AnCC. We review additional data from 99 previously published patients with invdupdel(8p) and compare data of 17 patients from the literature with both CMA analysis and brain imaging to refine genotype-phenotype correlations for AnCC. This led us to refine a region of 5.1 Mb common to duplications of patients with AnCC and discuss potential candidate genes within this region.

Molecular Profiling of Spermatozoa Reveals Correlations between Morphology and Gene Expression: A Novel Biomarker Panel for Male Infertility.

Choosing spermatozoa with an optimum fertilizing potential is one of the major challenges in assisted reproductive technologies (ART). This selection is mainly based on semen parameters, but the addition of molecular approaches could allow a more functional evaluation. To this aim, we used sixteen fresh sperm samples from patients undergoing ART for male infertility and classified them in the high- and poor-quality groups, on the basis of their morphology at high magnification. Then, using a DNA sequencing method, we analyzed the spermatozoa methylome to identify genes that were differentially methylated. By Gene Ontology and protein-protein interaction network analyses, we defined candidate genes mainly implicated in cell motility, calcium reabsorption, and signaling pathways as well as transmembrane transport. RT-qPCR of high- and poor-quality sperm samples allowed showing that the expression of some genes, such as AURKA, HDAC4, CFAP46, SPATA18, CACNA1C, CACNA1H, CARHSP1, CCDC60, DNAH2, and CDC88B, have different expression levels according to sperm morphology. In conclusion, the present study shows a strong correlation between morphology and gene expression in the spermatozoa and provides a biomarker panel for sperm analysis during ART and a new tool to explore male infertility.

Position statement on the diagnosis and management of premature/primary ovarian insufficiency (except Turner Syndrome).

Premature ovarian insufficiency (POI) is a rare pathology affecting 1-2% of under-40 year-old women, 1 in 1000 under-30 year-olds and 1 in 10,000 under-20 year-olds. There are multiple etiologies, which can be classified as primary (chromosomal, genetic, auto-immune) and secondary or iatrogenic (surgical, or secondary to chemotherapy and/or radiotherapy). Despite important progress in genetics, more than 60% of cases of primary POI still have no identifiable etiology; these cases are known as idiopathic POI. POI is defined by the association of 1 clinical and 1 biological criterion: primary or secondary amenorrhea or spaniomenorrhea of>4 months with onset before 40 year of age, and elevated follicle-stimulating hormone (FSH)>25IU/L on 2 assays at>4 weeks' interval. Estradiol level is low, and anti-Müllerian hormone (AMH) levels have usually collapsed. Initial etiological work-up comprises auto-immune assessment, karyotype, FMR1 premutation screening and gene-panel study. If all of these are normal, the patient and parents may be offered genome-wide analysis under the "France Génomique" project. The term ovarian insufficiency suggests that the dysfunction is not necessarily definitive. In some cases, ovarian function may fluctuate, and spontaneous pregnancy is possible in around 6% of cases. In confirmed POI, hormone replacement therapy is to be recommended at least up to the physiological menopause age of 51 years. Management in a rare diseases center may be proposed.

Detailed cell-level analysis of sperm nuclear quality among the different hypo-osmotic swelling test (HOST) classes.

PURPOSE: We studied the quality differences between the different hypo-osmotic swelling test (HOST) classes, as measured by criteria of DNA fragmentation, DNA decondensation, and nuclear architecture. The aim was to find particular HOST classes associated with good-quality metrics, which may be potentially used in ICSI (intra-cytoplasmic sperm injection). METHODS: Ten patients from the Department of Reproductive Medicine at Tenon Hospital (Paris, France) were included. Their semen samples were collected and divided into two fractions: one was incubated in a hypo-osmotic solution as per HOST protocol and sorted by sperm morphology, and a second was incubated without undergoing the HOST protocol to serve as an unsorted baseline. Three parameters were assessed: DNA fragmentation (TUNEL assay), DNA decondensation (chromomycin A3 assay), and nuclear architecture (FISH, with telomeric and whole chromosome painting probes). The different HOST classes were evaluated for these three parameters, and statistical analysis was performed for each class versus the unsorted non-HOST-treated sperm. Results with p<0.05 were considered statistically significant. RESULTS: For each of the parameters evaluated, we found significant differences between HOST-selected spermatozoa and non-selected spermatozoa. Overall, spermatozoa of HOST classes B and B+ exhibited the highest quality based on four metrics (low DNA fragmentation, low DNA decondensation, short inter-telomeric distance, and small chromosome 1 territory area), while spermatozoa of HOST classes A and G exhibited the poorest quality by these metrics. CONCLUSION: In addition to their pathophysiological interest, our results open possibilities of sperm selection prior to ICSI, which may allow for optimization of reproductive outcomes in heretofore unstudied patient populations.

Recurrence of an early postzygotic rescue of an inherited unbalanced translocation resulting in mosaic segmental uniparental isodisomy of chromosome 11q in siblings.

Balanced translocations are associated with a risk of transmission of unbalanced chromosomal rearrangements in the offspring. Such inherited chromosomal abnormalities are typically non-mosaic as they are present in the germline. We report the recurrence in two siblings of a mosaicism for a chromosomal rearrangement inherited from their asymptomatic father who carried a balanced t(2;11)(q35;q25) translocation. Both siblings exhibited a similar phenotype including intellectual disability, dysmorphic features, kyphoscoliosis, and cervical spinal stenosis. Karyotyping, fluorescence in situ hybridization and SNP array analysis of blood lymphocytes of both siblings identified two cell lines: one carrying a 2q35q37.3 duplication and a 11q25qter deletion (~90% cells), and one carrying an 11q uniparental isodisomy of maternal origin (~10% cells). We hypothesize that these mosaics were related to a postzygotic rescue mechanism which unexpectedly recurred in both siblings.

SRY-negative 46,XX testicular/ovotesticular DSD: Long-term outcomes and early blockade of gonadotropic axis.

OBJECTIVE: SRY-negative 46,XX testicular and ovotesticular disorders/differences of sex development (T/OTDSD) represent a very rare and unique DSD condition where testicular tissue develops in the absence of a Y chromosome. To date, very few studies have described the phenotype, clinical and surgical management and long-term outcomes of these patients. Particularly, early blockade of the gonadotropic axis in patients raised in the female gender to minimize postnatal androgenization has never been reported. DESIGN: Retrospective description of sixteen 46,XX T/OTDSD patients. RESULTS: Sixteen 46,XX SRY-negative T/OTDSD were included. Most (12/16) were diagnosed in the neonatal period. Sex of rearing was male for six patients and female for ten, while the clinical presentation varied, with an external masculinization score from 1 to 10. Five patients raised as girl were successfully treated with GnRH analog to avoid virilization during minipuberty. Ovotestes/testes were found bilaterally for 54% of the patients and unilaterally for the others (with a contralateral ovary). Gonadal surgery preserved appropriate tissue in the majority of cases. Spontaneous puberty occurred in two girls and one boy, while two boys required hormonal induction of puberty. One of the girls conceived spontaneously and had an uneventful pregnancy. DNA analyses (SNP-array, next-generation sequencing and whole-exome sequencing) were performed. A heterozygous frameshit mutation in the NR2F2 gene was identified in one patient. CONCLUSIONS: This study presents a population of patients with 46,XX SRY-negative T/OTDSD. Early blockade of gonadotropic axis appears efficient to reduce and avoid further androgenization in patients raised as girls.

Simple FISH-based evaluation of spermatic nuclear architecture shows an abnormal chromosomal organization in balanced chromosomal rearrangement carriers.

INTRODUCTION: Interphasic DNA has a constant three-dimensional conformation, which is particularly striking for spermatic DNA, with distinct chromosomal territories and a constant chromosomal conformation. We hypothesized that this organization is fragile, and that an excess or a lack of chromosomal segments could hinder the genomic structure as a whole. METHODS: Five human male chromosomal translocation carriers and five controls were included. Spermatic DNA spatial organization was studied, in both balanced and unbalanced spermatozoa, with two-dimensional fluorescent in situ hybridization (FISH) via analysis of chromosomes not implicated in the cases' translocations, compared to that of normal controls. Two parameters were studied: the distance between the two telomeric ends of chromosome 1, and the area of the chromosomal territories of chromosomes 1 and 17. RESULTS: Sperm FISH analysis of rearrangement carriers revealed changes in the nuclear architecture compared to that of controls. Inter-telomeric distance and chromosomal territories areas were both significantly increased. DISCUSSION: We show that an excess or lack of chromosomal segments can hinder the normal spatial nuclear architecture in sperm. These results show that nuclear architecture is a fragile assembly, and that local chromosomal abnormalities may impact the nucleus as a whole. This suggests a potential avenue for selection of spermatozoa prior to in vitro fertilization, not only in rearrangement carriers but also in the infertile population at large. Furthermore, we suggest that 2D-FISH could possibly be a useful tool in assessing spermatic nuclear organization in a way to evaluate male fertility.

A 14q distal chromoanagenesis elucidated by whole genome sequencing.

Chromoanagenesis represents an extreme form of genomic rearrangements involving multiple breaks occurring on a single or multiple chromosomes. It has been recently described in both acquired and rare constitutional genetic disorders. Constitutional chromoanagenesis events could lead to abnormal phenotypes including developmental delay and congenital anomalies, and have also been implicated in some specific syndromic disorders. We report the case of a girl presenting with growth retardation, hypotonia, microcephaly, dysmorphic features, coloboma, and hypoplastic corpus callosum. Karyotype showed a de novo structurally abnormal chromosome 14q31qter region. Molecular characterization using SNP-array revealed a complex unbalanced rearrangement in 14q31.1-q32.2, on the paternal chromosome 14, including thirteen interstitial deletions ranging from 33 kb to 1.56 Mb in size, with a total of 4.1 Mb in size, thus suggesting that a single event like chromoanagenesis occurred. To our knowledge, this is one of the first case of 14q distal deletion due to a germline chromoanagenesis. Genome sequencing allowed the characterization of 50 breakpoints, leading to interruption of 10 genes including YY1 which fit with the patient's phenotype. This precise genotyping of breaking junction allowed better definition of genotype-phenotype correlations.

Pathogenic variants in the DEAH-box RNA helicase DHX37 are a frequent cause of 46,XY gonadal dysgenesis and 46,XY testicular regression syndrome.

PURPOSE: XY individuals with disorders/differences of sex development (DSD) are characterized by reduced androgenization caused, in some children, by gonadal dysgenesis or testis regression during fetal development. The genetic etiology for most patients with 46,XY gonadal dysgenesis and for all patients with testicular regression syndrome (TRS) is unknown. METHODS: We performed exome and/or Sanger sequencing in 145 individuals with 46,XY DSD of unknown etiology including gonadal dysgenesis and TRS. RESULTS: Thirteen children carried heterozygous missense pathogenic variants involving the RNA helicase DHX37, which is essential for ribosome biogenesis. Enrichment of rare/novel DHX37 missense variants in 46,XY DSD is highly significant compared with controls (P value = 5.8 × 10-10). Five variants are de novo (P value = 1.5 × 10-5). Twelve variants are clustered in two highly conserved functional domains and were specifically associated with gonadal dysgenesis and TRS. Consistent with a role in early testis development, DHX37 is expressed specifically in somatic cells of the developing human and mouse testis. CONCLUSION: DHX37 pathogenic variants are a new cause of an autosomal dominant form of 46,XY DSD, including gonadal dysgenesis and TRS, showing that these conditions are part of a clinical spectrum. This raises the possibility that some forms of DSD may be a ribosomopathy.

Association of Maternal First Trimester Serum Levels of Free Beta Human Chorionic Gonadotropin and Hypospadias: A Population Based Study.

PURPOSE: Human chorionic gonadotropin stimulates fetal testosterone production and contributes to normal development of male genitalia. Using population based data we hypothesized that differences in maternal free beta human chorionic gonadotropin may be associated with hypospadias. MATERIALS AND METHODS: Data were obtained from the Paris Registry of Congenital Malformations (REMAPAR) (2011 to 2016). The initial study population included 3,172 pregnant women who gave birth to a singleton live born male infant with a congenital malformation. After exclusion of cases with unknown beta human chorionic gonadotropin and those with chromosomal or genetic abnormalities, the study population included 194 boys with isolated hypospadias and 1,075 controls. For cases with operative notes (125) we obtained data on type (proximal/distal) of hypospadias. Using quantile regression we compared median values of multiple of median beta human chorionic gonadotropin measured for first trimester Down syndrome screening (10th to 13th gestational weeks) for overall as well as by type of hypospadias vs controls. We also considered possible effects of placental dysfunction (maternal age, intrauterine growth retardation and preterm births) as potential confounding factors. RESULTS: Overall the median beta human chorionic gonadotropin multiple of median was comparable for women who had an infant with hypospadias vs controls (0.99 vs 0.95, p=0.3). However, proximal hypospadias was associated with a statistically significant higher median multiple of median than distal hypospadias or unspecified (1.49 vs 0.92 vs 1.05, p=0.02). The estimates were comparable after adjustment for placental dysfunction. CONCLUSIONS: Our findings support the hypothesis that an alteration in maternal beta human chorionic gonadotropin levels is associated with hypospadias. However, this association appears to be limited to proximal hypospadias.