Pathological characteristics of SRY-negative 38,XX-DSD pigs: A family case report.

Disorders of sex development (DSD) are congenital conditions characterized by atypical development of chromosomes, gonads, or anatomical sex. XX-DSD pigs disrupt the production of high-quality breeding pigs and impede the advancement of the pig industry. However, the etiology of XX-DSD pigs remains unclear. Systematic reports on the genetic and pathological characteristics of prepubescent XX-DSD pigs in familial contexts are sparse. This study aimed to investigate the genetic and pathological features of one-month-old XX-DSD pigs within a familial context and to provide phenotypic information to elucidate the pathogenic mechanisms of XX-DSD pigs. The findings revealed that inbreeding within the XX-DSD family may contribute to the pathogenesis of XX-DSD pigs. All XX-DSD pigs in the family had a chromosomal sex of female and were male pseudohermaphrodites. The degree of masculinization of the reproductive organs varied among XX-DSD pigs, demonstrating phenotypic heterogeneity. HE staining showed that the testes of prepubescent XX-DSD pigs contained vesicles in the seminiferous tubules, with or without vestigial germ cells. Ultrastructural analyses indicated that sertoli cells, leydig cells and germ cells in the testes of XX-DSD pigs exhibited pathological damage, confirming impaired testicular function. Immunofluorescence staining revealed high expression of SRY-box transcription factor 9 (SOX9) in XX-DSD pig testicular tissues, while forkhead box L2 (FOXL2) was minimally expressed. Disordered secretion of reproductive hormones in prepubescent XX-DSD pigs indicated abnormal hypothalamic-pituitary-gonadal axis (HPGA) function. This study elucidates the genetic and pathological characteristics of prepubescent XX-DSD pigs in familial case, providing valuable insights for further exploration of the pathogenic mechanisms underlying XX-DSD.

Unusual sex chromosomal DSD in a domestic Shorthair cat with a 37,X/38,XY mosaic karyotype.

BACKGROUND: Sex chromosome abnormalities associated with disorders of sexual development (DSD) are rarely described in cats, mainly due to the lack of chromossome studies that precisely reveal the condition. Genetic approaches are therefore required in order to detect sex chromossomes abnormalities as variations in the number and structure of chromosomes, or the presence of a second cell line as mosaicim or chimerism. CASE PRESENTATION: A male Shorthair cryptorchid cat was presented with clinical signs of anorexia, tenesmus and hyperthermia. Ultrasonography revealed a fluid-filled structure, with approximately 1 cm in diameter, adjacent to the descending colon. Computed tomography evidenced a tubular structure, ventral to the descending colon and caudal to the bladder, which extended cranially, through two branches. Histopathological evaluation confirmed the presence of two atrophic uterine horns and one hypoplastic testicle with epididymis at the end of one of the uterine horns. The end of the other uterine horn was attached to a structure composed by a mass of adipocytes. Cytogenetic analysis revealed a mosaic 37,X/38,XY karyotype. The two cell lines were found in 15% and 85% of the lymphocytes, respectively. Genetic analysis confirmed the presence of SRY and ZFY genes in blood and hair bulbs, and revealed a marked reduction in SRY expression in the testicle. Additionally, this case presented exceptionally rare features, such as a Leydig' cell tumour and a chronic endometritis in both uterine horns. CONCLUSIONS: Complete imaging workup, cytogenetic analysis and SRY gene expression should be systematically realized, in order to properly classify disorders of sexual development (DSD) in cats.

FDXR variants cause adrenal insufficiency and atypical sexual development.

Genetic defects affecting steroid biosynthesis cause cortisol deficiency and differences of sex development; among these defects are recessive mutations in the steroidogenic enzymes CYP11A1 and CYP11B, whose function is supported by reducing equivalents donated by ferredoxin reductase (FDXR) and ferredoxin. So far, mutations in the mitochondrial flavoprotein FDXR have been associated with a progressive neuropathic mitochondriopathy named FDXR-related mitochondriopathy (FRM), but cortisol insufficiency has not been documented. However, patients with FRM often experience worsening or demise following stress associated with infections. We investigated 2 female patients with FRM carrying the potentially novel homozygous FDXR mutation p.G437R with ambiguous genitalia at birth and sudden death in the first year of life; they presented with cortisol deficiency and androgen excess compatible with 11-hydroxylase deficiency. In addition, steroidogenic FDXR-variant cell lines reprogrammed from 3 patients with FRM fibroblasts displayed deficient mineralocorticoid and glucocorticoid production. Finally, Fdxr-mutant mice allelic to the severe p.R386W human variant showed reduced progesterone and corticosterone production. Therefore, our comprehensive studies show that human FDXR variants may cause compensated but possibly life-threatening adrenocortical insufficiency in stress by affecting adrenal glucocorticoid and mineralocorticoid synthesis through direct enzyme inhibition, most likely in combination with disturbed mitochondrial redox balance.

Testicular differentiation in 46,XX DSD: an overview of genetic causes.

In mammals, the development of male or female gonads from fetal bipotential gonads depends on intricate genetic networks. Changes in dosage or temporal expression of sex-determining genes can lead to differences of gonadal development. Two rare conditions are associated with disruptions in ovarian determination, including 46,XX testicular differences in sex development (DSD), in which the 46,XX gonads differentiate into testes, and 46,XX ovotesticular DSD, characterized by the coexistence of ovarian and testicular tissue in the same individual. Several mechanisms have been identified that may contribute to the development of testicular tissue in XX gonads. This includes translocation of SRY to the X chromosome or an autosome. In the absence of SRY, other genes associated with testis development may be overexpressed or there may be a reduction in the activity of pro-ovarian/antitesticular factors. However, it is important to note that a significant number of patients with these DSD conditions have not yet recognized a genetic diagnosis. This finding suggests that there are additional genetic pathways or epigenetic mechanisms that have yet to be identified. The text will provide an overview of the current understanding of the genetic factors contributing to 46,XX DSD, specifically focusing on testicular and ovotesticular DSD conditions. It will summarize the existing knowledge regarding the genetic causes of these differences. Furthermore, it will explore the potential involvement of other factors, such as epigenetic mechanisms, in developing these conditions.

[Clinicopathological analysis of gonadal differentiation of sex development disorder].

Objective: To investigate pathological features and differential diagnosis in the gonads with disorder of sex development. Methods: Thirty-six cases of clinically diagnosed hermaphroditism with gonadal biopsy in the Department of Pathology, the Seventh Medical Center of People's Liberation Army General Hospital from April 2007 to July 2021, were collected. All biopsy pathological sections were reviewed, and the gonadal cases with abnormal pathological morphology were screened out. The clinical and imaging data and karyotype of these cases were reviewed. Additional immunohistochemical staining was performed and relevant literature was reviewed. Results: Seven cases of ovotesticular disorder of sex development (OTDSD) were identified, which were characterized by the presence of testicular and ovarian differentiation in the same individual. All patients were under 15 years old and presented with abnormal appearance of external genitalia, and the ratio of male to female was 2∶5. Ultrasonography showed testicular structure in all female patients and cryptorchidism in all male patients. The most common karyotype was 46, XX. One case with undifferentiated gonadal tissue (UGT) and one case with streak gonads were screened out. UGT germ cells were neither in seminiferous tubules nor in follicles, but randomly distributed in an ovarial-type interstitial background, sometimes accompanied by immature sex cords. Streak gonads resembled UGT without germ cells. FOXL2 was positive in granulosa cells, but negative in Sertoli cells. SOX9 expression was opposite. OCT4 was weakly positively/negatively expressed in oocytes and positively expressed in the germ nuclei of UGT. Conclusions: Four differentiation patterns need to be identified in the gonadal biopsy: ovarian differentiation, testicular differentiation, undifferentiated gonadal tissue and streak gonad. The positive expression of SOX9 indicates testicular differentiation, while the positive expression of FOXL2 confirms ovarian differentiation, and the expression of both markers in the same tissue indicates ovotestis differentiation. It is very important to identify UGT, because that has a high probability of developing into gonadoblastoma in the future.

[Clinical, genetic, and pathological analysis in 165 children with disorders of sex development]. / 165ä¾‹å„¿ç«¥æ€§å‘è‚²å¼‚å¸¸ç–¾ç— çš„ä¸´åºŠã€é—ä¼ å­¦ä¸Žç— ç†å­¦åˆ†æž.

OBJECTIVES: To investigate the clinical phenotypes, genetic characteristics, and pathological features of children with disorders of sex development (DSD). METHODS: A retrospective analysis was conducted on epidemiological, clinical phenotype, chromosomal karyotype, gonadal pathology, and genotype data of 165 hospitalized children with DSD at Children's Hospital of Hebei Province and Tangshan Maternal and Child Health Hospital from August 2008 to December 2022. RESULTS: Among the 165 children with DSD, common presenting symptoms were short stature (62/165, 37.6%), clitoromegaly (33/165, 20.0%), cryptorchidism (28/165, 17.0%), hypospadias (24/165, 14.5%), and skin pigmentation abnormalities/exteriorized pigmented labia majora (19/165, 11.5%). Chromosomal karyotype analysis was performed on 127 cases, revealing 36 cases (28.3%) of 46,XX DSD, 34 cases (26.8%) of 46,XY DSD, and 57 cases (44.9%) of sex chromosome abnormalities. Among the sex chromosome abnormal karyotypes, the 45,X karyotype (11/57, 19%) and 45,X/other karyotype mosaicism (36/57, 63%) were more common. Sixteen children underwent histopathological biopsy of gonadal tissues, resulting in retrieval of 25 gonadal tissues. The gonadal tissue biopsies revealed 3 cases of testes, 3 cases of dysplastic testes, 6 cases of ovaries, 11 cases of ovotestes, and 1 case each of streak gonad and agenesis of gonads. Genetic testing identified pathogenic/likely pathogenic variants in 23 cases (23/36, 64%), including 12 cases of 21-hydroxylase deficiency congenital adrenal hyperplasia caused by CYP21A2 pathogenic variants. CONCLUSIONS: Short stature, clitoromegaly, cryptorchidism, hypospadias, and skin pigmentation abnormalities are common phenotypes in children with DSD. 45,X/other karyotype mosaicism and CYP21A2 compound heterozygous variants are major etiological factors in children with DSD. The most commonly observed gonadal histopathology in children with DSD includes ovotestes, ovaries, and testes/dysgenetic testes.

Duplication of SOX3 in an SRY-negative 46,XX male with prostatic utricle: case report and literature review.

BACKGROUND: 46,XX male disorders of sex development are rare. Approximately 80% of cases of testicular tissue differentiation may be due to translocation of SRY to the X chromosome or an autosome. SRY-negative 46,XX males show overexpression of pro-testis genes, such as SOX9 and SOX3, or failure of pro-ovarian genes, such as WNT4 and RSPO1, which induces testis differentiation, however, almost all testicles exhibit dysgenesis. Following inadequate exposure to androgens during the embryo stage, remnants of the Mullerian duct and incomplete closure of the urogenital sinus lead to enlargement of prostatic utricles. This condition is associated with proximal hypospadias and disorders of sex development. Many cases are asymptomatic, but show increased rates of postoperative complications and surgical failure. CASE PRESENTATION: A 5-year-old Chinese boy with scrotal hypospadias and bilateral cryptorchidism with prostatic utricles was presented. Gonadal histology showed ovo-testicular tissue on the right side and testicular tissue on the left side; all testicular tissue exhibited dysgenesis. Furthermore, chromosome karyotype analysis revealed 46,XX and, the presence of SRY was ruled out by polymerase chain reaction analysis. Whole-genome analysis showed the boy has a 1.4-Mb duplication in the Xq27.1q27.2 region (arr[hg19]Xq27.1q27.2:139585794-140996652) involving SOX3. No SOX3 duplication was observed in the parents, who had a normal phenotype. CONCLUSIONS: We report the first case of an SRY-negative 46 XX male with prostatic utricle caused by SOX3 duplication. SOX3 duplication may cause sex reversal, and all 46,XX SRY-negative males should be screened for SOX3 mutations. Gonadal biopsy is recommended to evaluate ovarian and testicular tissue development. Testicular dysgenesis and low exposure to male hormones during fetal development can lead to enlarged prostatic utricles. Thus endoscopic examination should be performed preoperatively to detect prostatic utricles in SRY-negative 46,XX males to determine the surgical plan and reduce postoperative complications.

Gonadal Development in Smallmouth Bass (Micropterus Dolomieu) Reared in the Absence and Presence of 17-α-Ethinylestradiol.

Testicular oocytes in wild adult bass (Micropterus spp.) are considered a potential indication of exposure to estrogenic compounds in municipal, agricultural, or industrial wastewater. However, our ability to interpret links between testicular oocyte occurrence in wild fish species and environmental pollutants is limited by our understanding of normal and abnormal gonadal development. We previously reported low-to-moderate testicular oocyte prevalence (7%-38%) among adult male bass collected from Minnesota waters with no known sources of estrogenic compounds. In the present study, two experiments were conducted in which smallmouth bass (Micropterus dolomieu) fry were exposed to control water or 17-α-ethinylestradiol (EE2) during gonadal differentiation, then reared in clean water for an additional period. Histological samples were evaluated at several time points during the exposure and grow-out periods, and the sequence and timing of gonadal development in the presence of estrogen were compared with that of control fish. Testicular oocytes were not observed in any control or EE2-exposed fish. Among groups exposed to 1.2 or 5.1 ng/L EE2 in Experiment 1 or 3.0 ng/L EE2 in Experiment 2, ovaries were observed in 100% of fish up to 90 days after exposure ceased, and approximately half of those ovaries had abnormal characteristics, suggesting that they likely developed in sex-reversed males. Groups exposed to 0.1, 0.4, or 1.0 ng/L in Experiment 2 developed histologically normal ovaries and testes in proportions not significantly different from 1:1. These findings suggest that, while presumably able to cause sex reversal, juvenile exposure to EE2 may not be a unique cause of testicular oocytes in wild bass, although the long-term outcomes of exposure are unknown. Environ Toxicol Chem 2022;41:1416-1428. © 2022 SETAC. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.

Are NR5A1 Variations a Frequent Cause of 46,XX Ovotesticular Disorders of Sex Development? Analysis from a Single Center and Systematic Review.

INTRODUCTION: Ovotesticular disorder of sex development (OT-DSD) is a rare condition defined by concomitance of testicular tissue and ovarian tissue (containing follicles) in the same individual. In SRY-negative 46,XX OT-DSD, the presence of testicular tissue may be due to variations in NR5A1. Our aims were to search for NR5A1 variants in SRY-negative 46,XX OT-DSD patients and to perform a systematic review on the contribution of NR5A1 variations to 46,XX OT-DSD. METHODS: Sanger sequencing of NR5A1 was performed in seven SRY-negative 46,XX OT-DSD patients: five simplex cases and two with another sibling with a 46,XX DSD. Systematic review of original studies on NR5A1 sequencing of 46,XX OT-DSD patients was performed according to PRISMA-P guideline. Case reports were selected for analysis of clinical features. Individuals with NR5A1-associated testicular DSD were not included. RESULTS: Sanger sequencing of NR5A1 did not reveal pathogenic variants among our patients. Our cohort was included in this systematic review with seven other articles, totalizing fifty-six 46,XX OT-DSD patients investigated by Sanger or whole-exome sequencing. From them, three NR5A1 pathogenic variants were identified (5% of the cases). Clinical analysis of these 3 cases and 5 case reports revealed: predominance of ovotestis (13/16 gonads) and bilateral OT-DSD (5/8 cases). CONCLUSION: The etiology of most 46,XX OT-DSD cases remains elusive, highlighting the importance of a deeper molecular investigation.

Adult acampomelic campomelic dysplasia and disorders of sex development due to a reciprocal translocation involving chromosome 17q24.3 upstream of the SOX9 gene.

Balanced chromosomal rearrangements with a breakpoint located upstream of the sex determining region Y-box 9 (SOX9) gene on chromosome 17q24.3 are associated with skeletal abnormalities, campomelic dysplasia (CMPD), or acampomelic campomelic dysplasia (ACMPD). We report on a female patient with a reciprocal translocation of t (11; 17) (p15.4; q24.3), who was diagnosed with acampomelic campomelic dysplasia. The 34-year-old Japanese patient presented with distinct skeletal abnormalities, profound intellectual disability, and female phenotype despite the presence of Y chromosome and the sex determining region Y (SRY) gene. Her menarche started at 33 years and 4 months after hormone therapy of estrogen therapy followed by estrogen progesterone therapy. By conducting whole genome sequencing followed by Sanger sequencing validation, we determined the precise breakpoint positions of the reciprocal translocation, one of which was located 203 kb upstream of the SOX9 gene. Considering the phenotypic variations previously reported among the CMPD/ACMPD patients with a chromosomal translocation in the vicinity of SOX9, the identified translocation was concluded to be responsible for all major phenotypes observed in the patient.

Sex Differences in the Immune System Become Evident in the Perinatal Period in the Four Core Genotypes Mouse.

We have used the four core genotypes (FCG) mouse model, which allows a distinction between effects of gonadal secretions and chromosomal complement, to determine when sex differences in the immune system first appear and what influences their development. Using splenic T cell number as a measure that could be applied to neonates with as yet immature immune responses, we found no differences among the four genotypes at postnatal day 1, but by day 7, clear sex differences were observed. These sex differences were unexpectedly independent of chromosomal complement and similar in degree to gonadectomized FCG adults: both neonatal and gonadectomized adult females (XX and XY) showed 2-fold the number of CD4+ and 7-fold the number of CD8+ T cells versus their male (XX and XY) counterparts. Appearance of this long-lived sex difference between days 1 and 7 suggested a role for the male-specific perinatal surge of testicular testosterone. Interference with the testosterone surge significantly de-masculinized the male CD4+, but not CD8+ splenic profile. Treatment of neonates demonstrated elevated testosterone limited mature cell egress from the thymus, whereas estradiol reduced splenic T cell seeding in females. Neonatal male splenic epithelium/stroma expressed aromatase mRNA, suggesting capacity for splenic conversion of perinatal testosterone into estradiol in males, which, similar to administration of estradiol in females, would result in reduced splenic T cell seeding. These sex steroid effects affected both CD4+ and CD8+ cells and yet interference with the testosterone surge only significantly de-masculinized the splenic content of CD4+ cells. For CD8+ cells, male cells in the thymus were also found to express one third the density of sphingosine-1-phosphate thymic egress receptors per cell compared to female, a male characteristic most likely an indirect result of Sry expression. Interestingly, the data also support a previously unrecognized role for non-gonadal estradiol in the promotion of intra-thymic cell proliferation in neonates of both sexes. Microarray analysis suggested the thymic epithelium/stroma as the source of this hormone. We conclude that some immune sex differences appear long before puberty and more than one mechanism contributes to differential numbers and distribution of T cells.

Clinical phenotype and management of individuals with mosaic monosomy X with Y chromosome material stratified by genital phenotype.

Individuals mosaic for monosomy X and a cell line with Y chromosome material can have genitalia that appear phenotypical female, male, or ambiguous. Those with this karyotype and typical female genitalia are diagnosed with Turner syndrome; however, this definition specifically excludes those with genitalia other than typical female. There is limited information on whether medical and neurodevelopmental risks are similar among individuals with monosomy X and Y chromosome material across genital phenotypes. This multicenter retrospective study compared comorbidities and clinical management in individuals with monosomy X and Y material and male/ambiguous genitalia to those with typical female genitalia. Electronic medical records for all patients with monosomy X and Y material (n = 76) at two large U.S. pediatric centers were abstracted for predetermined data and outcomes. Logistic regression was used to compare the two phenotypic groups adjusting for site and duration of follow-up. The male/ambiguous genitalia group was just as likely to have congenital heart disease (RR 1.0, 95%CI [0.5-1.9]), autoimmune disease (RR 0.6 [0.2-1.3]), and neurodevelopmental disorders (RR 1.4 [0.8-1.2]) as those with female genitalia. Despite similar risks, they were less likely to receive screening and counseling. In conclusion, individuals with monosomy X and Y chromosome material have similar medical and neurodevelopmental risks relative to individuals with Turner syndrome regardless of genitalia, but there are notable differences in clinical management.

Novel variants in the stem cell niche factor WNT2B define the disease phenotype as a congenital enteropathy with ocular dysgenesis.

WNT2B is a member of the Wnt family, a group of signal transduction proteins involved in embryologic development and stem cell renewal and maintenance. We recently reported homozygous nonsense variants in WNT2B in three individuals with severe, neonatal-onset diarrhea, and intestinal failure. Here we present a fourth case, from a separate family, with neonatal diarrhea associated with novel compound heterozygous WNT2B variants. One of the two variants was a frameshift variant (c.423del [p.Phe141fs]), while the other was a missense change (c.722 G > A [p.G241D]) that we predict through homology modeling to be deleterious, disrupting post-translational acylation. This patient presented as a neonate with severe diet-induced (osmotic) diarrhea and growth failure resulting in dependence on parenteral nutrition. Her gastrointestinal histology revealed abnormal cellular architecture particularly in the stomach and colon, including oxyntic atrophy, abnormal distribution of enteroendocrine cells, and a paucity of colonic crypt glands. In addition to her gastrointestinal findings, she had bilateral corneal clouding and atypical genital development later identified as a testicular 46,XX difference/disorder of sexual development. Upon review of the previously reported cases, two others also had anterior segment ocular anomalies though none had atypical genital development. This growing case series suggests that variants in WNT2B are associated with an oculo-intestinal (and possibly gonadal) syndrome, due to the protein's putative involvement in multiple developmental and stem cell maintenance pathways.

Gene expression and functional abnormalities in XX/Sry Leydig cells.

The SRY gene induces testis development even in XX individuals. However, XX/Sry testes fail to produce mature sperm, due to the absence of Y chromosome carrying genes essential for spermatogenesis. XX/Sry Sertoli cells show abnormalities in the production of lactate and cholesterol required for germ cell development. Leydig cells are essential for male functions through testosterone production. However, whether XX/Sry adult Leydig cells (XX/Sry ALCs) function normally remains unclear. In this study, the transcriptomes from XY and XX/Sry ALCs demonstrated that immediate early and cholesterogenic gene expressions differed between these cells. Interestingly, cholesterogenic genes were upregulated in XX/Sry ALCs, although downregulated in XX/Sry Sertoli cells. Among the steroidogenic enzymes, CYP17A1 mediates steroid 17α-hydroxylation and 17,20-lyase reaction, necessary for testosterone production. In XX/Sry ALCs, the latter reaction was selectively decreased. The defects in XX/Sry ALCs, together with those in the germ and Sertoli cells, might explain the infertility of XX/Sry testes.

A large cohort of disorders of sex development and their genetic characteristics: 6 novel mutations in known genes.

INTRODUCTION: Disorders of sex development (DSD) constitutes a group of congenital conditions that affect urogenital differentiation and are associated with chromosomal, gonadal and phenotypic sex abnormalities. OBJECTIVE: To evaluate the clinical and genetic features of childhood DSD cases. MATERIALS AND METHODS: DSD patients followed up between the years of 2002-2018 were evaluated in terms of their complaints, demographic, clinical features and genetic diagnoses. RESULTS: Out of 289 patients, 143(49.5%) were classified as 46XY DSD, 62(21.5%) as 46XX DSD and 84(29%) as sex chromosomal DSD. Genetic diagnosis was achieved in 150 patients (51.9%). The distribution of the molecular diagnosis of the 46XY DSD patients were; 12 (26.6%) SRD5A2, 10 (22.2%) AR, 7 (15.5%) HSD17B3, 3 (6.6%) WT-1, 2 (4.4%) AMHR2, 2 (4.4%) AMH, 2 (4.4%) LHCGR, 2 (4.4%) HSD3B2, 1 (2.2%) NR5A1, 1 (2.2%) CYP17A1 and 1 (2.2%) SRY mutation. Fifty (80.6%) of the 46XX DSD patients received a diagnosis with clinical and laboratory findings. Twenty-four (38.7%) of them were 21-hydroxylase deficiency, 9(14.5%) Rokitansky-Küster-Hauser Syndrome, 4 (6.5%) 11-ß hydroxylase deficiency, 3 (4.8%) gonadal dysgenesis and 2 (3.2%) aromatase deficiency. In 46XX group pathogenic mutations were detected in 21(33.8%) of the patients. Eighty-four (29%) patients were diagnosed as sex chromosomal disorder. Of these 66 (78.5%) were Turner Syndrome, 6 (7.2%) Klinefelter Syndrome and 10 (11.9%) mix gonadal dysgenesis. Gender re-assignment was decided in 11 patients. Malignant and pre-invasive lesions was diagnosed in 8 (2.7%) patients. CONCLUSION: Many of DSD's are clinically similar and etiology of numerous of them still cannot be established. A multi-disciplinary approach and new rapid genetic diagnostic methods are needed in the process from diagnosis to gender assignment and follow-up.

Molecular genetics of disorders of sex development in a highly consanguineous population.

Consanguinity increases the risk of hereditary diseases including disorders of sex development (DSD). There are minimal data on DSD in the highly consanguineous population of Saudi Arabia. This study reports the molecular genetics of a series of patients with different types of DSD. METHODS: We enrolled 77 patients from 47 families with DSD. DNA was isolated from peripheral leucocytes. Genes of interest were amplified by polymerase chain reaction and subsequently sequenced. RESULTS: Overall, 77 patients from 47 families (44 of them are consanguineous) had a total of 29 mutations; 16 of them were described before and 13 were novel mutations. The most common condition was 5-α reductase (SRD5A2) deficiency (25 patients from 18 families) and the most common mutation was a splice site mutation in intron 1 (c.282-2A>G). The next most common condition was 11-ß hydroxylase (CYP11B1) deficiency where 19 patients from 10 families had 8 mutations (7 of them are novel). Other mutations affected CYP17A1 with 2 novel and 2 known mutations in 7 patients; HSD3B2 with 2 known mutations in 11 patients of 4 families; StAR with 1 novel and 1 known mutations in 4 patients; NR0B1 with 1 novel mutation in 2 siblings; HSD17B3 with 1 known mutation in 3 siblings; LHCGR with 1 novel mutation in 2 siblings; and AR with 1 novel and 3 known mutations in 4 unrelated patients. CONCLUSION: In the highly consanguineous and homogeneous population of Saudi Arabia, SRD5A2 and CYP11B1 deficiencies are common causes of DSDs. Other DSDs occur less frequently but often with novel mutations.

Survey of disorders of sex development in a large cohort of patients with diverse Mendelian phenotypes.

Disorders of sex development (DSD) are congenital conditions with atypical development of chromosomal, gonadal, or anatomical sex. The estimated incidence ranges from 1 in 4,500-5,500 for strictly defined "ambiguous genitalia" to 1 in 300 or higher when a broader definition is implemented. In this study, we aim to define DSD phenotypes encountered in a large heterogeneous cohort of molecularly characterized Mendelian disorders in a single center. Data were retrieved for patients with documented abnormal genitalia based on the 2006 consensus criteria. Out of 149 patients (129 families) with compatible human phenotype ontology, 76 patients (68 families) had an identified genetic cause and were included in our analysis. Potentially causal variants were identified in 42 genes, and two patients had a dual molecular diagnosis. Six genes have no associated phenotype in OMIM (PIANP, CELSR2, USP2, FAM179B, TXNDC15, and CCDC96). Thirteen genes have non-DSD OMIM phenotypes, thus we are expanding their phenotype to include DSD. We also highlight how certain disorders are under-recognized despite their established DSD phenotype in OMIM, especially CTU2-related DREAM-PL syndrome and TSPYL1-related sudden infant death with dysgenesis of the testes syndrome. In conclusion, this study of a large heterogeneous Mendelian cohort expands the list of genes and disorders beyond those classically DSD-linked.

Heterozygous deletion of Sox9 in mouse mimics the gonadal sex reversal phenotype associated with campomelic dysplasia in humans.

Heterozygous mutations in the human SOX9 gene cause the skeletal malformation syndrome campomelic dysplasia which in 75% of 46, XY individuals is associated with male-to-female sex reversal. Although studies in homozygous Sox9 knockout mouse models confirmed that SOX9 is critical for testis development, mice heterozygous for the Sox9-null allele were reported to develop normal testes. This led to the belief that the SOX9 dosage requirement for testis differentiation is different between humans, which often require both alleles, and mice, in which one allele is sufficient. However, in prior studies, gonadal phenotypes in heterozygous Sox9 XY mice were assessed only by either gross morphology, histological staining or analyzed on a mixed genetic background. In this study, we conditionally inactivated Sox9 in somatic cells of developing gonads using the Nr5a1-Cre mouse line on a pure C57BL/6 genetic background. Section and whole-mount immunofluorescence for testicular and ovarian markers showed that XY Sox9 heterozygous gonads developed as ovotestes. Quantitative droplet digital PCR confirmed a 50% reduction of Sox9 mRNA as well as partial sex reversal shown by an upregulation of ovarian genes. Our data show that haploinsufficiency of Sox9 can perturb testis development in mice, suggesting that mice may provide a more accurate model of human disorders/differences of sex development than previously thought.

Homozygous p.Val89Leu plays an important pathogenic role in 5α-reductase type 2 deficiency patients with homozygous p.Arg246Gln in SRD5A2.

OBJECTIVE: To evaluate the pathogenic role of a few benign variants and hypomorphic pathogenic variants in SRD5A2. DESIGN AND METHODS: We retrospectively analyzed phenotypes and genotypes in 23 Indian patients with genetically proven steroid 5α-reductase 2 (SRD5A2) deficiency. The interactions of the SRD5A2 enzymes resulting due to the most common benign variant (p.Val89Leu), the most common (hypomorphic) pathogenic variant (p.Arg246Gln) and the double variants (p.Val89Leu and p.Arg246Gln) in SRD5A2 were compared with that of the wild type (WT) enzyme by molecular dynamics (MD) simulation. RESULTS: The majority (n = 19, 82.61%) of patients presented for atypical genitalia and had male gender identity (16/20). Including the two novel ones (p.Leu83Pro, p.Ala28Leufs*103), a total of nine different pathogenic variants were observed. p.Arg246Gln was the most common pathogenic variant (n = 12). Homozygous p.Arg246Gln (n = 9) variant was associated with milder undervirilization (Sinnecker score of ≤3a: 8/9 vs 6/14, P = 0.04) and had concurrent homozygous p.Val89Leu in all patients. Interestingly, asymptomatic fathers of two index patients were homozygous for p.Arg246Gln which questioned the pathogenicity of the variation as a sole factor. Unlike all symptomatic homozygous p.Arg246Gln patients who were also homozygous for p.Val89Leu, asymptomatic homozygous p.Arg246Gln fathers were heterozygous for p.Val89Leu. On MD simulation SRD5A2 p.Val89Leu-Testeosterone (T) and SRD5A2 p.Arg246Gln-T complexes, but not SRD5A2 p.Val89Leu and p.Arg246Gln-T complex, demonstrated close interaction between NADPH and T as that of SRD5A2 WT-T. CONCLUSIONS: p.Arg246Gln may not be pathogenic as a sole variation even in the homozygous state; additional contribution of homozygous p.Val89Leu variant may be essential for the pathogenicity of p.Arg246Gln in SRD5A2.

Natural human chimeras: A review.

The term chimera has been borrowed from Greek mythology and has a long history of use in biology and genetics. A chimera is an organism whose cells are derived from two or more zygotes. Recipients of tissue and organ transplants are artificial chimeras. This review concerns natural human chimeras. The first human chimera was reported in 1953. Natural chimeras can arise in various ways. Fetal and maternal cells can cross the placental barrier so that both mother and child may become microchimeras. Two zygotes can fuse together during an early embryonic stage to form a fusion chimera. Most chimeras remain undetected, especially if both zygotes are of the same genetic sex. Many are discovered accidently, for example, during a routine blood group test. Even sex-discordant chimeras can have a normal male or female phenotype. Only 28 of the 50 individuals with a 46,XX/46,XY karyotype were either true hermaphrodites or had ambiguous genitalia. Blood chimeras are formed by blood transfusion between dizygotic twins via the shared placenta and are more common than was once assumed. In marmoset monkey twins the exchange via the placenta is not limited to blood but can involve other tissues, including germ cells. To date there are no examples in humans of twin chimeras involving germ cells. If human chimeras are more common than hitherto thought there could be many medical, social, forensic, and legal implications. More multidisciplinary research is required for a better understanding of this fascinating subject.