Formin-mediated nuclear actin at androgen receptors promotes transcription.

Steroid hormone receptors are ligand-binding transcription factors essential for mammalian physiology. The androgen receptor (AR) binds androgens mediating gene expression for sexual, somatic and behavioural functions, and is involved in various conditions including androgen insensitivity syndrome and prostate cancer1. Here we identified functional mutations in the formin and actin nucleator DAAM2 in patients with androgen insensitivity syndrome. DAAM2 was enriched in the nucleus, where its localization correlated with that of the AR to form actin-dependent transcriptional droplets in response to dihydrotestosterone. DAAM2 AR droplets ranged from 0.02 to 0.06 µm3 in size and associated with active RNA polymerase II. DAAM2 polymerized actin directly at the AR to promote droplet coalescence in a highly dynamic manner, and nuclear actin polymerization is required for prostate-specific antigen expression in cancer cells. Our data uncover signal-regulated nuclear actin assembly at a steroid hormone receptor necessary for transcription.

Disruption of the topologically associated domain at Xp21.2 is related to 46,XY gonadal dysgenesis.

BACKGROUND: Duplications at the Xp21.2 locus have previously been linked to 46,XY gonadal dysgenesis (GD), which is thought to result from gene dosage effects of NR0B1 (DAX1), but the exact disease mechanism remains unknown. METHODS: Patients with 46,XY GD were analysed by whole genome sequencing. Identified structural variants were confirmed by array CGH and analysed by high-throughput chromosome conformation capture (Hi-C). RESULTS: We identified two unrelated patients: one showing a complex rearrangement upstream of NR0B1 and a second harbouring a 1.2 Mb triplication, including NR0B1. Whole genome sequencing and Hi-C analysis revealed the rewiring of a topological-associated domain (TAD) boundary close to NR0B1 associated with neo-TAD formation and may cause enhancer hijacking and ectopic NR0B1 expression. Modelling of previous Xp21.2 structural variations associated with isolated GD support our hypothesis and predict similar neo-TAD formation as well as TAD fusion. CONCLUSION: Here we present a general mechanism how deletions, duplications or inversions at the NR0B1 locus can lead to partial or complete GD by disrupting the cognate TAD in the vicinity of NR0B1. This model not only allows better diagnosis of GD with copy number variations (CNVs) at Xp21.2, but also gives deeper insight on how spatiotemporal activation of developmental genes can be disrupted by reorganised TADs causing impairment of gonadal development.

A Novel Look at Dosage-Sensitive Sex Locus Xp21.2 in a Case of 46,XY Partial Gonadal Dysgenesis without NR0B1 Duplication.

A region of 160 kb at Xp21.2 has been defined as dosage-sensitive sex reversal (DSS) and includes the NR0B1 gene, considered to be the candidate gene involved in XY gonadal dysgenesis if overexpressed. We describe a girl with 46,XY partial gonadal dysgenesis carrying a 297 kb duplication at Xp21.2 upstream of NR0B1 initially detected by chromosomal microarray analysis. Fine mapping of the breakpoints by whole-genome sequencing showed a tandem duplication of TASL (CXorf21), GK and partially TAB3, upstream of NR0B1. This is the first description of an Xp21.2 duplication upstream of NR0B1 associated with 46,XY partial gonadal dysgenesis.

In vitro functional characterization of the novel DHH mutations p.(Asn337Lysfs*24) and p.(Glu212Lys) associated with gonadal dysgenesis.

In humans, mutations of Desert Hedgehog gene (DHH) have been described in patients with 46,XY gonadal dysgenesis (GD), associated or not with polyneuropathy. In this study, we describe two patients diagnosed with GD, both harboring novel DHH compound heterozygous mutations p.[Tyr176*];[Asn337Lysfs*24] and p.[Tyr176*];[Glu212Lys]. To investigate the functional consequences of p.(Asn337Lysfs*24) and p.(Glu212Lys) mutations, located within the C-terminal part of DHh on auto-processing, we performed in vitro cleavage assays of these proteins in comparison with Drosophila melanogaster Hedgehog (Hh). We found that p.(Glu212Lys) mutation retained 50% of its activity and led to a partially abolished DHh auto-processing. In contrast, p.(Asn337Lysfs*24) mutation resulted in a complete absence of auto-proteolysis. Furthermore, we found a different auto-processing profile between Drosophila Hh and human DHh, which suggests differences in the processing mechanism between the two species. Review of the literature shows that proven polyneuropathy and GD is associated with complete disruption of DHh-N, whereas disruption of the DHh auto-processing is only described with GD. We propose a model that may explain the differences between Schwann and Leydig cell development by autocrine versus paracrine DHh signaling. To our knowledge, this is the first study investigating the effect of DHH mutations on DHh in vitro auto-processing.

Functional characterization of five NR5A1 gene mutations found in patients with 46,XY disorders of sex development.

Steroidogenic factor-1 (SF1), encoded by the NR5A1 gene, is a key regulator of steroidogenesis and reproductive development. NR5A1 mutations described in 46,XY patients with disorders of sex development (DSD) can be associated with a range of conditions of phenotypes; however, the genotype-phenotype correlation remains elusive in many cases. In the present study, we describe the impact of five NR5A1 variants (three novel: p.Arg39Cys, p.Ser32Asn, and p.Lys396Argfs*34; and two previously described: p.Cys65Tyr and p.Cys247*) on protein function, identified in seven patients with 46,XY DSD. In vitro functional analyses demonstrate that NR5A1 mutations impair protein functions and result in the DSD phenotype observed in our patients. Missense mutations in the DNA binding domain and the frameshift mutation p.Lys396Argfs*34 lead to both, markedly affected transactivation assays, and loss of DNA binding, whereas the mutation p.Cys247* retained partial transactivation capacity and the ability to bind a consensus SF1 responsive element. SF1 acts in a dose-dependent manner and regulates a cascade of genes involved in the sex determination and steroidogenesis, but in most cases reported so far, still lead to a sufficient adrenal steroidogenesis and function, just like in our cases, in which heterozygous mutations are associated to 46,XY DSD with intact adrenal steroid biosynthesis.

Persistence of foetal testicular features in patients with defective androgen signalling.

OBJECTIVE: Congenital defects of androgen synthesis or action in 46,XY individuals can result in impaired virilisation, despite the apparent testicular development. In a recent case, report of a young adult with complete androgen insensitivity syndrome (CAIS), tumourous gonadal tissue was shown to express HSD17B3 in Sertoli cells (SCs) and not in Leydig cells (LCs). This expression pattern differs from the typical adult human testis and resembles a foetal mouse testis, suggesting an underlying testicular development and function defect. Here, we investigate the effect of altered androgen signalling in gonads from five 46,XY individuals with defects in androgen synthesis or action. METHODS: Gonadal tissue sections from four patients with CAIS, one with CYP17A1 deficiency, and one control were immunostained for LC developmental and steroidogenic markers. The expression of some of these markers during development was investigated by reanalysing previously published single-cell RNA sequencing (scRNA-seq) data from normal human testicular tissues. RESULTS: All gonadal tissues from the patients show an exclusive expression of HSD17B3 in SCs and an expression of the foetal/immature LC marker DLK1 in a subset of LCs, suggesting an androgen-dependent differentiation defect of adult SCs and LCs. Furthermore, reanalysis of scRNA-seq data reveals an expression of HSD17B3 in foetal and neonatal SCs that is downregulated in adult SCs. CONCLUSIONS: Androgen signalling may affect the differentiation of adults, but possibly not foetal SCs or LCs, and may induce a shift of testosterone production from the tubular compartment in the foetal phase to the interstitial compartment in the adult phase.

DHX37 and NR5A1 Variants Identified in Patients with 46,XY Partial Gonadal Dysgenesis.

The group of disorders known as 46,XY gonadal dysgenesis (GD) is characterized by anomalies in testis determination, including complete and partial GD (PGD) and testicular regression syndrome (TRS). Several genes are known to be involved in sex development pathways, however approximately 50% of all cases remain elusive. Recent studies have identified variants in DHX37, a gene encoding a putative RNA helicase essential in ribosome biogenesis and previously associated with neurodevelopmental disorders, as a cause of PGD and TRS. To investigate the potential role of DHX37 in disorders of sexual development (DSD), 25 individuals with 46,XY DSD were analyzed and putative pathogenic variants were found in four of them. WES analyses were performed on these patients. In DHX37, the variant p.(Arg308Gln), recurrent associated with DSD, was identified in one patient; the p.(Leu467Val), predicted to be deleterious, was found together with an NR5A1 loss-of-function variant in patient 2; and, the p.(Val999Met) was identified in two unrelated patients, one of whom (patient 3) also carried a pathogenic NR5A1 variant. For both patients carrying DHX37 and NR5A1 pathogenic variants, a digenic inheritance is suggested. Our findings support the importance of DHX37 variants as a cause of disorders of sex development, implying a role in testis development.

Clinical, Biochemical, and Molecular Characterization of Indian Children with Clinically Suspected Androgen Insensitivity Syndrome.

This study describes the clinical, biochemical, and molecular characteristics of Indian children with 46,XY DSD and suspected androgen insensitivity syndrome (AIS). Fifty children (median age 3.0 years, range 0-16.5 years) with 46,XY DSD and a suspected diagnosis of AIS were enrolled. Sanger sequencing was performed to identify pathogenic variants in the androgen receptor (AR) gene and to study genotype-phenotype correlations. All 5 (100%) patients with CAIS and 14/45 (31%) patients with PAIS had pathogenic/likely pathogenic variants in the AR gene (overall, 14 different variants in 19 patients; 38.8%). There was no significant difference in clinical (cryptorchidism, hypospadias, or external masculinizing score) or biochemical parameters (gonadotropins and testosterone) between patients with or without pathogenic variants. However, patients with AIS were more likely to have a positive family history, be assigned female gender at birth, and present with gynaecomastia at puberty. Three novel pathogenic/likely pathogenic variants, including one splice donor site variant c.2318+1G>A, one frameshift variant p.H790Lfs*40, and one missense variant p.G821E, were identified in 3 patients with CAIS. The missense variant p.G821E was predicted as deleterious, damaging, disease-causing, and likely functionally inactive by in silico analysis and protein modelling study. Two previously not reported pathogenic/likely pathogenic variants, including p.R386H and p.G396R, were identified in patients with PAIS. This study contributes in expanding the spectrum of pathogenic variants in the AR gene in patients with AIS. Only 31% patients with a provisional diagnosis of PAIS had pathogenic variants in the AR gene, suggesting other possible mechanisms or candidate genes may be responsible for such a phenotypic presentation.

Pubertal development in 46,XY patients with NR5A1 mutations.

PURPOSE: Mutations in the NR5A1 gene, encoding the transcription factor Steroidogenic Factor-1, are associated with a highly variable genital phenotype in patients with 46,XY differences of sex development (DSD). Our objective was to analyse the pubertal development in 46,XY patients with NR5A1 mutations by the evaluation of longitudinal clinical and hormonal data at pubertal age. METHODS: We retrospectively studied a cohort of 10 46,XY patients with a verified NR5A1 mutation and describe clinical features including the external and internal genitalia, testicular volumes, Tanner stages and serum concentrations of LH, FSH, testosterone, AMH, and inhibin B during pubertal transition. RESULTS: Patients who first presented in early infancy due to ambiguous genitalia showed spontaneous virilization at pubertal age accompanied by a significant testosterone production despite the decreased gonadal volume. Patients with apparently female external genitalia at birth presented later in life at pubertal age either with signs of virilization and/or absence of female puberty. Testosterone levels were highly variable in this group. In all patients, gonadotropins were constantly in the upper reference range or elevated. Neither the extent of virilization at birth nor the presence of Müllerian structures reliably correlated with the degree of virilization during puberty. CONCLUSION: Patients with NR5A1 mutations regardless of phenotype at birth may demonstrate considerable virilization at puberty. Therefore, it is important to consider sex assignment carefully and avoid irreversible procedures during infancy.

MYRF: A New Regulator of Cardiac and Early Gonadal Development-Insights from Single Cell RNA Sequencing Analysis.

De novo variants in the myelin regulatory factor (MYRF), a transcription factor involved in the differentiation of oligodendrocytes, have been linked recently to the cardiac and urogenital syndrome, while familiar variants are associated with nanophthalmos. Here, we report for the first time on a patient with a de novo stop-gain variant in MYRF (p.Q838*) associated with Scimitar syndrome, 46,XY partial gonadal dysgenesis (GD) and severe hyperopia. Since variants in MYRF have been described in both 46,XX and 46,XY GD, we assumed a role of MYRF in the early development of the bipotential gonad. We used publicly available single cell sequencing data of human testis and ovary from different developmental stages and analysed them for MYRF expression. We identified MYRF expression in the subset of coelomic epithelial cells at stages of gonadal ridge development in 46,XX and 46,XY individuals. Differential gene expression analysis revealed significantly upregulated genes. Within these, we identified CITED2 as a gene containing a MYRF binding site. It has been shown that Cited2-/- mice have gonadal defects in both testis and ovary differentiation, as well as defects in heart development and establishment of the left-right axis. This makes MYRF a potential candidate as an early regulator of gonadal and heart development via upregulation of the transcriptional cofactor CITED2.

Can Non-Coding NR5A1 Gene Variants Explain Phenotypes of Disorders of Sex Development?

INTRODUCTION: NR5A1 is an essential transcription factor that regulates several target genes involved in reproduction and endocrine function. Pathogenic variants in this gene are responsible for a wide spectrum of disorders/differences of sex development (DSD). METHODS: The molecular study involved Sanger sequencing, in vitro assays, and whole exome sequencing (WES). RESULTS: Four variants were identified within the NR5A1 non-coding region in 3 patients with 46,XY DSD. In vitro analyses showed that promoter activity was affected in all cases. WES revealed variants in SRA1, WWOX, and WDR11 genes. DISCUSSION/CONCLUSION: Evaluation of clinical and phenotypic significance of variants located in a non-coding region of a gene can be complex, and little is known regarding their association with DSD. Nevertheless, based on the important region for interaction with cofactors essential to promote appropriated sex development and on our in vitro results, it is feasible to say that an impact on gene expression can be expected and that this may be correlated with the DSD pathophysiology presented in our patients. Considering the number of cases that remain elusive after screening for the well-known DSD related genes, we emphasize the importance of a careful molecular analysis of NR5A1 non-coding region which is commonly neglected and might explain some idiopathic DSD cases.

Metabolic effects of estradiol versus testosterone in complete androgen insensitivity syndrome.

PURPOSE: To study differences in metabolic outcomes between testosterone and estradiol replacement in probands with complete androgen insensitivity syndrome (CAIS). METHODS: In this multicentre, double-blind, randomized crossover trial, 26 women with CAIS were included of whom 17 completed the study. After a two-months run in phase with estradiol, probands either received transdermal estradiol followed by crossover to transdermal testosterone or vice versa. After six months, differences in lipids, fasting glucose, insulin, hematocrit, liver parameters and blood pressure between the treatment phases were investigated. RESULTS: Linear mixed models adjusted for period and sequence did not reveal major group differences according to treatment for the investigated outcomes. In each treatment group, there were however significant uniform changes in BMI and cholesterol. BMI increased significantly, following six months of estradiol ( + 2.7%; p = 0.036) as well as testosterone treatment ( + 2.8%; p = 0.036). There was also a significant increase in total ( + 10.4%; p = 0.001) and LDL-cholesterol ( + 29.2%; p = 0.049) and a decrease in HDL-cholesterol (-15.8%; p < 0.001) following six months of estradiol as well as six months of testosterone treatment (total cholesterol: + 14.6%; p = 0.008; LDL-cholesterol: + 39.1%; p = 0.005, HDL-cholesterol: -15.8%; p = 0.004). Other parameters remained unchanged. CONCLUSION: Transdermal estradiol as well as testosterone treatment in women with CAIS results in worsening in lipid profiles. Given the relatively small sample size, subtle group differences in other metabolic parameters may have remained undetected.

Functional characterization of GNAS mutations found in patients with pseudohypoparathyroidism type Ic defines a new subgroup of pseudohypoparathyroidism affecting selectively Gsα-receptor interaction.

Pseudohypoparathyroidism type Ia (PHPIa) is caused by GNAS mutations leading to deficiency of the α-subunit of stimulatory G proteins (Gsα) that mediate signal transduction of G protein-coupled receptors via cAMP. PHP type Ic (PHPIc) and PHPIa share clinical features of Albright hereditary osteodystrophy (AHO); however, in vitro activity of solubilized Gsα protein is normal in PHPIc but reduced in PHPIa. We screened 32 patients classified as PHPIc for GNAS mutations and identified three mutations (p.E392K, p.E392X, p.L388R) in four unrelated families. These and one novel mutation associated with PHPIa (p.L388P) were introduced into a pcDNA3.1(-) expression vector encoding Gsα wild-type and expressed in a Gsα-null cell line (Gnas(E2-/E2-) ). To investigate receptor-mediated cAMP accumulation, we stimulated the endogenous expressed ß(2) -adrenergic receptor, or the coexpressed PTH or TSH receptors, and measured the synthesized cAMP by RIA. The results were compared to receptor-independent cholera toxin-induced cAMP accumulation. Each of the mutants associated with PHPIc significantly reduced or completely disrupted receptor-mediated activation, but displayed normal receptor-independent activation. In contrast, PHPIa associated p.L388P disrupted both receptor-mediated activation and receptor-independent activation. We present a new subgroup of PHP that is caused by Gsα deficiency and selectively affects receptor coupling functions of Gsα.

In vitro functional characterization of androgen receptor gene mutations at arginine p.856 of the ligand-binding-domain associated with androgen insensitivity syndrome.

Androgens are critical for male sex differentiation. Their actions are mediated by the androgen receptor (AR). Mutations disrupting AR function result in the androgen insensitivity syndrome (AIS). In this study, we identified in a patient with complete AIS, a novel AR mutation p.R856L. To investigate the functional properties of p.R856L, we performed functional studies. In comparison, we have characterized two already described mutations: p.R856H and p.R856C. We used a model composed of two different promoters fused to a reporter gene, two cell lines, and showed that all mutations were able to transactivate the (ARE)2-TATA promoter expressed in CHO cells more highly. Moreover, we confirmed the pathogenicity of the p.R856L and p.R856C mutations, and their associations with complete AIS. In contrast, the p.R856H mutation, which is associated with a spectrum of AIS phenotypes, showed less severe transcriptional constraints. Altogether, our studies allowed us to better characterize arginine residue at p.R856 position.

Validation of a next-generation sequencing (NGS) panel to improve the diagnosis of X-linked hypophosphataemia (XLH) and other genetic disorders of renal phosphate wasting.

BACKGROUND: Hypophosphataemic rickets (HR) comprise a clinically and genetically heterogeneous group of conditions, defined by renal-tubular phosphate wasting and consecutive loss of bone mineralisation. X-linked hypophosphataemia (XLH) is the most common form, caused by inactivating dominant mutations in PHEX, a gene encompassing 22 exons located at Xp22.1. XLH is treatable by anti-Fibroblast Growth Factor 23 antibody, while for other forms of HR such as therapy may not be indicated. Therefore, a genetic differentiation of HR is recommended. OBJECTIVE: To develop and validate a next-generation sequencing panel for HR with special focus on PHEX. DESIGN AND METHODS: We designed an AmpliSeq gene panel for the IonTorrent PGM next-generation platform for PHEX and ten other HR-related genes. For validation of PHEX sequencing 50 DNA-samples from XLH-patients, in whom 42 different mutations in PHEX and 1 structural variation have been proven before, were blinded, anonymised and investigated with the NGS panel. In addition, we analyzed one known homozygous DMP1 mutation and two samples of HR-patients, where no pathogenic PHEX mutation had been detected by conventional sequencing. RESULTS: The panel detected all 42 pathogenic missense/nonsense/splice-site/indel PHEX-mutations and in one the known homozygous DMP1 mutation. In the remaining two patients, we revealed a somatic mosaicism of a PHEX mutation in one; as well as two variations in DMP1 and a very rare compound heterozygous variation in ENPP1 in the second patient. CONCLUSIONS: This developed NGS panel is a reliable tool with high sensitivity and specificity for the diagnosis of XLH and related forms of HR.

CYP17A1 deficient XY mice display susceptibility to atherosclerosis, altered lipidomic profile and atypical sex development.

CYP17A1 is a cytochrome P450 enzyme with 17-alpha-hydroxylase and C17,20-lyase activities. CYP17A1 genetic variants are associated with coronary artery disease, myocardial infarction and visceral and subcutaneous fat distribution; however, the underlying pathological mechanisms remain unknown. We aimed to investigate the function of CYP17A1 and its impact on atherosclerosis in mice. At 4-6 months, CYP17A1-deficient mice were viable, with a KO:Het:WT ratio approximating the expected Mendelian ratio of 1:2:1. All Cyp17a1 knockout (KO) mice were phenotypically female; however, 58% were Y chromosome-positive, resembling the phenotype of human CYP17A1 deficiency, leading to 46,XY differences/disorders of sex development (DSD). Both male and female homozygous KO mice were infertile, due to abnormal genital organs. Plasma steroid analyses revealed a complete lack of testosterone in XY-KO mice and marked accumulation of progesterone in XX-KO mice. Elevated corticosterone levels were observed in both XY and XX KO mice. In addition, Cyp17a1 heterozygous mice were also backcrossed onto an Apoe KO atherogenic background and fed a western-type diet (WTD) to study the effects of CYP17A1 on atherosclerosis. Cyp17a1 x Apoe double KO XY mice developed more atherosclerotic lesions than Apoe KO male controls, regardless of diet (standard or WTD). Increased atherosclerosis in CYP17A1 XY KO mice lacking testosterone was associated with altered lipid profiles. In mice, CYP17A1 deficiency interferes with sex differentiation. Our data also demonstrate its key role in lipidomic profile, and as a risk factor in the pathogenesis of atherosclerosis.

Disorders of sex development expose transcriptional autonomy of genetic sex and androgen-programmed hormonal sex in human blood leukocytes.

BACKGROUND: Gender appears to be determined by independent programs controlled by the sex-chromosomes and by androgen-dependent programming during embryonic development. To enable experimental dissection of these components in the human, we performed genome-wide profiling of the transcriptomes of peripheral blood mononuclear cells (PBMC) in patients with rare defined "disorders of sex development" (DSD, e.g., 46, XY-females due to defective androgen biosynthesis) compared to normal 46, XY-males and 46, XX-females. RESULTS: A discrete set of transcripts was directly correlated with XY or XX genotypes in all individuals independent of male or female phenotype of the external genitalia. However, a significantly larger gene set in the PBMC only reflected the degree of external genital masculinization independent of the sex chromosomes and independent of concurrent post-natal sex steroid hormone levels. Consequently, the architecture of the transcriptional PBMC-"sexes" was either male, female or even "intersex" with a discordant alignment of the DSD individuals' genetic and hormonal sex signatures. CONCLUSION: A significant fraction of gene expression differences between males and females in the human appears to have its roots in early embryogenesis and is not only caused by sex chromosomes but also by long-term sex-specific hormonal programming due to presence or absence of androgen during the time of external genital masculinization. Genetic sex and the androgen milieu during embryonic development might therefore independently modulate functional traits, phenotype and diseases associated with male or female gender as well as with DSD conditions.

Spectrum of Pathogenic Variants in SRD5A2 in Indian Children with 46,XY Disorders of Sex Development and Clinically Suspected Steroid 5α-Reductase 2 Deficiency.

The aim of this study was to assess the prevalence of pathogenic variants in the SRD5A2 gene in children with 46,XY disorders of sex development (DSD) with normal to high serum testosterone levels and absence of Müllerian structures on imaging and to evaluate the genotype-phenotype correlation. Seventy-five patients with 46,XY DSD and probable clinical diagnosis of 5α-reductase 2 deficiency or androgen insensitivity syndrome were enrolled. Genetic analysis was done for pathogenic variants in SRD5A2, and the genotype-phenotype correlation was studied. As a result, 10 pathogenic or likely pathogenic biallelic variants in SRD5A2, either homozygous or compound heterozygous, were identified in 25 of 75 (33.3%) patients. The hCG stimulated testosterone: dihydrotestosterone (T:DHT) ratio was elevated in all patients with pathogenic variants in SRD5A2 and in nearly 90% of those without pathogenic variants in SRD5A2 in whom this was assessed. The missense pathogenic variant p.R246Q was a hotspot. One novel pathogenic variant p.Y178*, and a variant p.F194I, not previously reported in patients with 5α-reductase 2 deficiency, were identified. The missense variant p.F194I was predicted as deleterious and damaging by in silico analysis and as likely to reduce the enzyme activity by protein modeling. In conclusion, pathogenic variants in SRD5A2 can be detected in a wide spectrum of Indian patients with 46,XY DSD. Molecular genetic analysis should be considered as a first-line test as the T:DHT ratio lacks specificity and a hotspot variant is present in a vast majority.

Clinical Findings and Follow-Up of 46,XY and 45,X/46,XY Testicular Dysgenesis.

Historically, the terms partial (PGD) and mixed gonadal dysgenesis (MGD) have been used to describe incomplete testicular differentiation in individuals with 46,XY or 45,X/46,XY karyotypes, respectively. However, it is currently unclear to what extent clinical features actually differ between these individuals. The aim of this study was to compare clinical, laboratory, and histological findings in these 2 groups. Patients with testicular dysgenesis seen in our service between 1989 and 2013 were selected. Sixty-one patients met the inclusion criteria. Individuals with 46,XY and 45,X/46,XY karyotypes were compared regarding genital features, gonadal histology and function, growth, and associated conditions. Twenty-five had mosaicism with a 45,X cell line (MGD), while a 46,XY karyotype (PGD) was found in 36 cases belonging to 32 families. Mutations in NR5A1, WT1, and SRY genes associated with testicular dysgenesis were found in 12 families. There were no significant differences regarding parental consanguinity, degree of external androgenization, gonadal location, histology, and function, and associated conditions. However, in the MGD group, the presence of a uterus, lower birth weight and length, and short stature were more often observed. Therefore, the use of histological features to classify PDG and MGD should be abandoned and replaced by classification based on karyotype.

Reduced Androgen Receptor Expression in Genital Skin Fibroblasts From Patients With 45,X/46,XY Mosaicism.

CONTEXT: Molecular mechanisms causing the broad phenotypic diversity of external masculinization in individuals with 45,X/46,XY mosaicism are poorly understood. OBJECTIVE: Analysis of androgen receptor (AR) expression and function as a putative influencing factor for the genital phenotype in patients with 45,X/46,XY mosaicism. DESIGN: Measurement of AR mRNA expression levels, AR activity [DHT-mediated APOD (apolipoprotein D) induction] and cellular 45,X/46,XY ratios in genital skin fibroblasts from individuals with 45,X/46,XY mosaicism and male reference individuals, and determination of the external virilization scale from individuals with 45,X/46,XY mosaicism. SETTING: University hospital endocrine research laboratory. Patients or Other Participants: 30 genital skin fibroblast cultures (GFs) from male reference individuals and 15 GFs from individuals with 45,X/46,XY mosaicism. INTERVENTION: None. MAIN OUTCOME MEASURES: Determination of AR mRNA expression and AR activity in male reference GFs and 45,X/46,XY GFs and correlation of the obtained data with the cellular 45,X/46,XY ratios and the patients' external virilization scale. RESULTS: In 6 of 15 45,X/46,XY GFs, AR mRNA expression and AR activity were significantly lower compared with those in the 46,XY reference GFs. In this subgroup of reduced AR mRNA expression, a positive trend was seen between AR mRNA expression and the percentage of XY-positive cells. Furthermore, we found a positive correlation between AR activity and the external virilization scale in the 15 45,X/46,XY GF samples (P = 0.03). CONCLUSION: Our results suggest that AR expression and AR activity might influence the phenotypic variability seen in patients with 45,X/46,XY mosaicism.