XX Male Patients Research Articles

46,XX males with congenital adrenal hyperplasia: a clinical and biochemical description.

Congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (21OHD) or 11-hydroxylase deficiency (11OHD) is characterized by underproduction of cortisol and overproduction of adrenal androgens. These androgens lead to a variable degree of virilization of the female external genitalia in 46,XX individuals. Especially in developing countries, diagnosis is often delayed and 46,XX patients might be assigned as males. This study aims to describe the clinical and biochemical characteristics of a unique cohort of untreated male-reared 46,XX classic CAH patients from Indonesia and discusses treatment challenges. Nine untreated classic CAH patients with 46,XX genotype and 21OHD (n=6) or 11OHD (n=3), aged 3-46 years old, were included. Biometrical parameters, clinical characteristics, and biochemical measurements including glucocorticoids, renin, androgens, and the pituitary-gonadal axis were evaluated. All patients had low early morning serum cortisol concentrations (median 89 nmol/L) without significant increase after ACTH stimulation. Three patients with salt wasting 21OHD reported one or more periods with seizures and/or vomiting in their past until the age of 6, but not thereafter. The remaining patients reported no severe illness or hospitalization episodes, despite their decreased capacity to produce cortisol. In the 21OHD patients, plasma renin levels were elevated compared to the reference range, and in 11OHD patients renin levels were in the low-normal range. All adult patients had serum testosterone concentrations within the normal male reference range. In 21OHD patients, serum 11-oxygenated androgens comprised 41-60% of the total serum androgen concentrations. Glucocorticoid treatment was offered to all patients, but they refused after counseling as this would reduce their endogenous androgen production and they did not report complaints of their low cortisol levels. We describe a unique cohort of untreated classic 46,XX male CAH patients without overt clinical signs of cortisol deficiency despite their cortisol underproduction and incapacity to increase cortisol levels after ACTH stimulation. The described adolescent and adult patients produce androgen levels within or above the normal male reference range. Glucocorticoid treatment will lower these adrenal androgen concentrations. Therefore, in 46,XX CAH patients reared as males an individual treatment approach with careful counseling and clear instructions is needed.

MP01-11 IN VITRO PROPAGATION OF UNDIFFERENTIATED SPERMATOGONIA ISOLATED FROM A 46, XX MALE TESTIS

You have accessJournal of UrologyCME1 Apr 2023MP01-11 IN VITRO PROPAGATION OF UNDIFFERENTIATED SPERMATOGONIA ISOLATED FROM A 46, XX MALE TESTIS Mark Xu, Aaron Bradshaw, Harpreet Gosal, Abinav Udaiyar, Omar Abdelaal, Kimberly Stogner-Underwood, Stuart Howards, Tamer Yalcinkaya, Anthony Atala, and Hooman Sadri-Ardekani Mark XuMark Xu More articles by this author , Aaron BradshawAaron Bradshaw More articles by this author , Harpreet GosalHarpreet Gosal More articles by this author , Abinav UdaiyarAbinav Udaiyar More articles by this author , Omar AbdelaalOmar Abdelaal More articles by this author , Kimberly Stogner-UnderwoodKimberly Stogner-Underwood More articles by this author , Stuart HowardsStuart Howards More articles by this author , Tamer YalcinkayaTamer Yalcinkaya More articles by this author , Anthony AtalaAnthony Atala More articles by this author , and Hooman Sadri-ArdekaniHooman Sadri-Ardekani More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003212.11AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: The SRY-positive 46, XX genotype is a testicular disorder of sex development found in 1/20,000 males. Patients are phenotypically male with small testes, variable hypogonadism, and azoospermia. Current literature has not yet documented spermatogonia in 46 XX male patients. This study aimed to identify and propagate spermatogonia in vitro in a 46, XX male. METHODS: A 28-year-old Caucasian male was initially suspected of having Klinefelter Syndrome (XXY), but karyotype and FISH analysis found a 46, XX genotype with a Y chromosome translocation containing the SRY gene onto the short distal arm of the X chromosome. He underwent experimental bilateral microscopic testicular biopsy and tissue preservation under IRB protocol. Fresh tissue was sent for histopathologic analysis using H&E staining and immunostaining of undifferentiated spermatogenic markers, including PGP 9.5/UCHL1 and ZBTB16. Biopsy portions from bilateral testes were used for characterization. Digital PCR and Real-time Quantitative (RT-Q) PCR were performed with ZBTB16 (undifferentiated spermatogonia), STAR (Leydig cell), CD34 (Peritubular cell), and SOX9 (Sertoli cell) target primers on testis samples from both sides. Isolated cells from the right testicular biopsy were cultured in the StemPro medium for four weeks. RT-QPCR was performed with target primers for undifferentiated spermatogonia (ZBTB16, UCHL1, THY1, CD9, ITGA6, ITGB1), Sertoli (SOX9, GATA4, CYP19a1), Leydig (STAR, CALB2), and peritubular cells (CD34). Digital PCR was performed using ZBTB16 target primer to quantify undifferentiated spermatogonia. Flow cytometry on cultured cells identified the spermatogonia stem cell (SSC) population as negative for HLA-ABC and positive for CD9 and CD49f. RESULTS: Testes exhibited Leydig cell hyperplasia, seminiferous tubule hyalinization like in Klinefelter Syndrome, and tubules filled mainly with Sertoli cells. Rare cells in seminiferous tubules were weakly positive for PGP9.5 and ZBTB16, representing undifferentiated spermatogonia. Digital and RT-QPCRs of testis tissue and cultured cells found evidence of all major testicular somatic cells and undifferentiated spermatogonia. Digital PCR for ZBTB16 identified 7.61% of cultured cells as undifferentiated spermatogonia and 0.71% as SSCs. Flow cytometry identified 2.52% of cultured cells as SSCs. CONCLUSIONS: This is the first study suggesting the presence of SSCs in 46 XX male testes. Future work will focus on further characterization of cultured testicular cells and In Vitro or In Vivo differentiation of SSCs from XX male testis. Source of Funding: WFIRM © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e6 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Mark Xu More articles by this author Aaron Bradshaw More articles by this author Harpreet Gosal More articles by this author Abinav Udaiyar More articles by this author Omar Abdelaal More articles by this author Kimberly Stogner-Underwood More articles by this author Stuart Howards More articles by this author Tamer Yalcinkaya More articles by this author Anthony Atala More articles by this author Hooman Sadri-Ardekani More articles by this author Expand All Advertisement PDF downloadLoading ...

Identification of an SRY-negative 46,XX infertility male with a heterozygous deletion downstream of SOX3 gene

BackgroundA male individual with a karyotype of 46,XX is very rare. We explored the genetic aetiology of an infertility male with a kayrotype of 46,XX and SRY negative.MethodsThe peripheral blood sample was collected from the patient and subjected to a few genetic testing, including chromosomal karyotyping, azoospermia factor (AZF) deletion, short tandem repeat (STR) analysis for AMELX, AMELY and SRY, fluorescence in situ hybridization (FISH) with specific probes for CSP 18/CSP X/CSP Y/SRY, chromosomal microarray analysis (CMA) for genomic copy number variations(CNVs), whole-genome analysis(WGA) for genomic SNV&InDel mutation, and X chromosome inactivation (XCI) analysis.ResultsThe patient had a karyotype of 46,XX. AZF analysis showed that he missed the AZF region (including a, b and c) and SRY gene. STR assay revealed he possessed the AMELX in the X chromosome, but he had no the AMELY and SRY in the Y chromosome. FISH analysis with CSP X/CSP Y/SRY showed only two X centromeric signals, but none Y chromosome and SRY. The above results of the karyotype, FISH and STR analysis did not suggest a Y chromosome chimerism existed in the patient's peripheral blood. The result of the CMA indicated a heterozygous deletion with an approximate size of 867 kb in Xq27.1 (hg19: chrX: 138,612,879–139,480,163 bp), located at 104 kb downstream of SOX3 gene, including F9, CXorf66, MCF2 and ATP11C. WGA also displayed the above deletion fragment but did not present known pathogenic or likely pathogenic SNV&InDel mutation responsible for sex determination and development. XCI assay showed that he had about 75% of the X chromosome inactivated.ConclusionsAlthough the pathogenicity of 46,XX male patients with SRY negative remains unclear, SOX3 expression of the acquired function may be associated with partial testis differentiation of these patients. Therefore, the CNVs analysis of the SOX3 gene and its regulatory region should be performed routinely for these patients.

Status of SOX9 in the testesof SRY negative and positive azoospermic males

In mammals, after expression of SRY in the sexually indifferent gonad, several genes encoding transcription factors and growth factors induced spermatogenesis and androgen production in testes. However, the etiology of SRY-negative XX men and pathologic difference of seminiferous tubules between SRY positive and negative men were unclear. We investigated several genes involved in gonad differentiation in XX male patients.

Molecular and cytogenetic characterization of six 46, XX males due to translocations between the short arms of X and Y chromosomes

To characterize molecular and cytogenetic abnormalities in six 46, XX males, and to investigate the clinical manifestations and underlying mechanisms in such patients. Clinical data of six XX male patients were collected. Karyotyping, multiple polymerase chain reaction (PCR) and fluorescence in situ hybridization (FISH) were utilized to detect and locate the sex determining region (SRY) gene. PCR and FISH showed that all patients were SRY-positive XX males. All patients have their SRY gene located at the tip of derivative X chromosomes, which have resulted from translocation between short arms of X and Y chromosomes. High resolution karyotyping at 550-750 band level has revealed that the translocation breakpoints were at Xp22.33 and Yp11.2 in three patients. In the remaining patients, the breakpoints were either at Xp22.32 and Yp11.31 or Xp22.31 and Yp11.2. The breakpoints at Xp22.32, Xp22.31 and Yp11.31 were rarely reported. Genotype-phenotype correlation analysis indicated that the clinical manifestations were age-specific. Four adult patients have come to clinical attention due to infertility, with typical features including azoospermia and testis dysgenesis, whereas poorly developed secondary sexual characteristics and short stature were main complaints of adolescence patients, and short stature was the sole symptom in a child patient. Combined karyotyping, PCR and FISH are important for the analysis of XX males. Particularly, high resolution karyotyping is valuable for the refinement of chromosome breakpoints and detailed analysis of genotype-phenotype correlation.

The investigation of the relationship between the phenotypes of 46, XX males and the SRY gene

To investigate the relationship between the phenotypes in XX male patients and the sex determining region(SRY) gene. Multiple polymerase chain reactions were carried out in 6 male patients with karyotype of 46, XX, and then the PCR products were sequenced directly. Three cases of male infertility were positive for the SRY gene without evident malformation in their extra genitalia, while 3 cases with testes were negative for the SRY gene, with evident malformation in their extra genitalia. The SRY gene is key in sex determination and development, yet there might be other important genes involved.

Up‐regulation of SOX9 in human sex‐determining region on the Y chromosome (SRY)‐negative XX males

In mammals, gonadal sex is normally determined by the presence or absence of the Y chromosome gene SRY. After expression of SRY in the sexually indifferent gonad, a number of genes encoding transcription factors and growth factors implicated in testis differentiation start to show male-specific expression. However, in XX males, these genes must be up-regulated in the absence of SRY, but the aetiology of SRY-negative XX maleness remains unclear. We examined the expression of representative gonad marker genes in SRY-negative XX male testes. RT-PCR and immunohistochemical studies revealed that SOX9, DAX-1, Ad4BP/SF-1, WT-1, GATA-4 and MIS were expressed in testicular tissues of SRY-negative XX males. Expression levels of SOX9 in testes of these patients averaged 1.9-fold higher than in normal XY testes, while expression levels of Ad4BP/SF-1, DAX-1 and MIS were lower in the SRY-negative XX testes than in XY testes. All XX patients were found to carry two copies of the SOX9 gene per diploid genome as do normal XX females and XY males. The XX male patients also carried two copies of the DAX-1 gene as do normal XX females, while normal XY males carry a single DAX-1 gene. Our data suggest that lesions affecting SOX9 expression are the key factor in sex determination in SRY-negative XX males, and that the decreased expression of Ad4BP/SF-1, DAX-1 and MIS contribute to their clinical features.

Varones XX: clínica y frecuencia en la consulta de esterilidad

Introduction Sexual differentiation is not always determined by chromosomal sex. Thus, there are some men with XX sex chromosomes and some women with XY chromosomes. This disassociation between gonadal sex and chromosomal sex is usually accompanied by reproductive problems and consequently this abnormality is more frequently observed in infertility clinics. Objective To assess the incidence of XX males consulting for male infertility, as well as the potential clinical implications of this abnormality. Material and methods We reviewed 4 XX men from a population of 4,560 men who attended the andrology clinic because of primary reproductive sterility. All 4 patients underwent seminal study, physical examination and gonadal determinations of plasma gonadotropins, testosterone, estradiol and inhibin B. In addition, FISH analysis was performed to detect the possible presence of the SRY gene. Results In all patients, the seminogram showed the presence of azoospermia, which was associated with hypospermia in 3 patients. The 4 patients had increased plasma levels of gonadotropin and inhibin B levels were undetectable. Testosterone and estradiol were found within the acceptable range of normality. Clinical examination only revealed the presence of testicular atrophy in all patients. The SRY gene was detected in all 4 patients and was located on the X chromosome in 3 of the patients and on chromosome 1 in the fourth patient. Conclusions The frequency of XX males in our infertility clinic was 0.087% (4/4,560), representing an incidence nearly 20 times higher than that expected in a general population (0.005%; 1/20,000). All 4 XX male patients were sterile as a result of azoospermia. Despite the dissociation between gonadal sex and chromosomal sex, clinical examination only revealed testicular atrophy, with no other phenotypic alterations.

Clinical, Endocrinological, and Epigenetic Features of the 46,XX Male Syndrome, Compared with 47,XXY Klinefelter Patients

The 46,XX male syndrome represents a rare, poorly characterized form of male hypogonadism. The objective of the study was to distinguish the 46,XX male syndrome from the more frequent 47,XXY-Klinefelter syndrome in regard to clinical, hormonal, and epigenetic features. This was a case-control study. The study was conducted at a university-based reproductive medicine and andrology institution. Eleven SRY-positive 46,XX males were compared with age-matched controls: 101 47,XXY Klinefelter patients, 78 healthy men, and 157 healthy women [latter all heterozygous for androgen receptor (AR) alleles]. There were no interventions. There was a comparison of phenotype, endocrine profiles, and X-chromosomal inactivation patterns of AR alleles. The 46,XX males were significantly smaller than Klinefelter patients or healthy men, resembling female controls in height and weight. The incidence of maldescended testes was significantly higher than that in Klinefelter patients and controls. Gynecomastia was more frequent in comparison with controls, whereas there was a nonsignificant trend in comparison with Klinefelter patients. All XX males were infertile and most were hypogonadal. The inactivation patterns of AR alleles in XX males were significantly more skewed than in Klinefelter patients and women. Seven of 10 heterozygous XX male patients displayed an extreme skewing of more than 80% with no preference toward the shorter or longer AR allele. The length of the AR CAG repeat polymorphism was positively related to traits of hypogonadism. XX males are distinctly different from Klinefelter patients in terms of clinical and epigenetic features. Nonrandom X chromosome inactivation ratios are common in XX males, possibly due to the translocated SRY gene. The existence of a Y-chromosomal, growth-related gene is discussed.

Identification and characterization of an Xp22.33;Yp11.2 translocation causing a triplication of several genes of the pseudoautosomal region 1 in an XX male patient with severe systemic lupus erythematosus

The X;Y translocation break point sequence in an XX male patient with prepubertal systemic lupus erythematosus (SLE) was characterized with the intention of identifying a predisposing gene(s) for SLE. Spectral karyotyping of the patient's metaphase chromosomes showed normal autosomes and 2 X chromosomes, one of which displayed a small portion of the Y chromosome. Using a Y chromosome polymerase chain reaction (PCR) walking strategy and inverse PCR, we found that the abnormal recombination occurred between retroviral long terminal repeats located at Xp22.33 (position 0.95 Mb; inside the pseudoautosomal regions) and Yp11.2 (4.20 Mb) downstream of the sex-determining region Y (SRY) gene. The complete DNA sequence of the break point was determined, revealing a partial duplication of the pseudoautosomal region 1 (PAR1) in the derivative X chromosome and causing a partial trisomy of the 12 known genes located between the interleukin-3 receptor alpha (IL3RA; position 1.1 Mb on the X and Y chromosomes) and CD99 (position 2.2 Mb) genes inclusively. All other X chromosome genes were present as 2 copies. Real-time quantitative PCR confirmed the presence of 3 copies of each of the 12 genes in the patient's genomic DNA. We also found that RNA for 1 of the candidate genes was indeed overexpressed in the patient's blood as compared with normal subjects. Taken together, the uniqueness of the translocation, the rarity of severe prepubertal SLE in males, and the presence of SLE in some patients with Klinefelter's syndrome (who have a triplication of the 2 PAR regions) point to a possible relationship between the partial triplication of the PAR1 region and the development of SLE.

Abnormal Bone Marrow-Derived Mesenchymal Stem Cell: Another Clone in Hematological Malignancy?.

The growth, differentiation, and survival of normal hematopoietic stem cells as well as bone marrow (BM) cancer cells are regulated closely by the BM microenvironment. In case of lymphoma, recent study has demonstrated that the endothelial cells (not lymphoma cells) in the lymphoma microenvironment have the same chromosomal abnormality to that of lymphoma cells themselves. Besides, in our prior study, the mesenchymal stem cells (MSCs) isolated from BM of patients with various hematological malignancies may have different surface antigens expression as comparing to normal control (Leukemia, 2005, doi:10.1038/sj.leu.2403795, published online 19 May 2005). We therefore hypothesize that the MSCs isolated from BM of patients with hematological malignancies are “different” to those of normal and may probably has the same chromosomal abnormality as to that of the hematological cancer cells. In this study, cytogenetic studies were performed on both BM-blood cells and BM-derived MSCs at the same time by using method of conventional G-banding. At submission of this abstract, we have just examined 4 specimens obtained from patients of various diseases as well as normal adult (see table). Surprisingly, we identified a new chromosomal translocation, t(3;9)(q25;q31), on the BM-derived MSCs of a patient (Case 1 of the table, Figure 1) with acute promyelocytic leukemia harboring t(15;17) on the BM blood cells. Besides, a complex chromosomal abnormalities was found in a male patient of AML (Case 2 of the table, Figure 2). Though he has received a successful allotransplant 2 years ago and the BM blood cells had completely converted to donor type (normal 46 XX), the BM-derived MSCs were still recipient in origin and having a chromosomal abnormality that was different to the BM blood cells (45, -Y) at initial diagnosis of AML. BM-derived MSCs from a patient of CML and normal adult were normal. It is still too early to making a solid conclusion at present and we are now analyzing more specimens to look into the phenomenon we found. However, this is a new field in leukemic research that deserved further investigation and our novel finding indicates that abnormal cells (another clone?) may present in the BM stroma of patients with hematological malignancies.Chromosomes of BM-blood cells and MSCsCase No.DiseasesChromosomes of BM-blood cellsChromosomes of BM-MSCsCase 1APL, at diagnosis46 XX, t(15;17)(q21;q11)46 XX, t(3;9)(q25;q31). (see figure 1)Case 2AML (45, -Y at diagnosis), after allotransplantNormal 46 XX (male patient, female donor)46 XY, t(7;22)(p22;q11); del(13)(q12q22); del(15)(q15?). (see figure 2)Case 3CML, at diagnosis46 XY, t(9;22)(q34;q11)Normal 46 XYCase 4Normal adultNormal 46 XYNormal 46 XY [Display omitted]

46,XX Male Patients—Clinical and Genetic Findings in Six Patients

We describe the clinical and molecular genetic findings of six 46,XX male patients evaluated at our institution. 46,XX male syndrome is described elsewhere in the literature as a true intersex condition with patients demonstrating considerable variability in physical exam findings, endocrinologic evaluation, and molecular genetics. Proper diagnosis of this condition is important with regard to counseling patients as to their future fertility. Chart Review - Retrospective We retrospectively reviewed the database of infertile male patients evaluated at our institution over the past 11 years. Six patients with 46,XX male syndrome were identified. Physical exam data, endocrinologic evaluation, and molecular genetic analysis were recorded for all patients. Testes biopsy was performed on two of six patients. All patients were found to have positive probes for the SRY sex-determining region with fluorescence in situ hybridization. Four of six patients were found to have translocation of SRY to the short arm of chromosome X. Two patients had cytogenetically evident translocation of a larger portion of chromosome Y including SRY (der(X)t(X;Y)(p22.3;p11.3)). Endocrinologic evaluation demonstrated elevated FSH (range 18.0-62.9 iU/L) and LH (range 7.4-22.5 iU/L) levels in all patients. Total testosterone levels were in the low-normal to normal range in all patients (range 226-625 ng/dL). In two patients undergoing testis tissue biopsy, no sperm were found on wet-prep examination. Final pathology demonstrated no evidence of spermatogenesis in either specimen. 46,XX male patients represent a small proportion of male patients evaluated for infertility; correct diagnosis of this condition and a true understanding of its implications are essential. This condition does not represent a variant of Klinefelter’s syndrome, as it has been previously termed elsewhere. In the 46,XX male syndrome, the presence of the SRY-determining region on the X choromosome of these patients accounts for their male phenotype. Without the remainder of the Ychromosome, however (specifically the AZF-a, AZF-b, and AZF-c regions), these patients cannot be expected to demonstrate any normal spermatogenesis. Others have described 46,XX males with absence of the SRY determining region, in addition to patients such as the ones above with a cytogenetically evident translocation der(X)t(X;Y)(p22.3;p11.3). Given this great variability in the molecular genetics of these patients, all male patients with a 46,XX karyotype should undergo FISH molecular analysis with SRY probe.

Three New 46,XX Male Patients: A Clinical, Cytogenetic and Molecular Analysis

XX males range phenotypically from completely masculinised individuals to true hermaphrodites and include a subset of SRY negative patients. The correlation between genotype (SRY+/-) and phenotype is still unclear. To report three new patients with this rare condition, one of whom was diagnosed prenatally and another was SRY negative, and to verify in our patients whether the presence of SRY results in a more masculinised phenotype. We present two phenotypically normal XX male patients (10 and 13.5 years) and one 3.1 years old XX male with ambiguous external male genitalia Prader IV. The patients were diagnosed by clinical, hormonal, sonographic, genetic and histological criteria. Basal hormonal status was normal for phenotype but an excessive response to GnRH testing was noticed in the second patient together with insufficient hCG stimulation in all three patients. Pelvic ultrasound displayed male structures without Müllerian ducts; testicular biopsy, performed only in the intersex patient, showed Sertoli and Leydig cell hypoplasia. Chromosome analysis confirmed 46,XX karyotype. FISH analysis and molecular analysis by PCR were positive for Yp fragments/SRY gene on Xp in two patients and negative in the patient with ambiguous external genitalia. In our observation Y chromosome-specific material containing the SRY gene translocated to the X chromosome results in a completely masculinised phenotype. In the intersex patient, incomplete masculinisation without SRY suggests a mutation of one or more downstream non-Y testis-determining genes.

Mutations in the SRY, DAX1, SF1 and WNT4 genes in Brazilian sex-reversed patients.

In most mammals, male development is triggered by the transient expression of the SRY gene, which initiates a cascade of gene interactions ultimately leading to the formation of a testis from the indifferent fetal gonad. Mutation studies have identified several genes essential for early gonadal development. We report here a molecular study of the SRY, DAX1, SF1 and WNT4 genes, mainly involved in sexual determination, in Brazilian 46,XX and 46,XY sex-reversed patients. The group of 46,XX sex-reversed patients consisted of thirteen 46,XX true hermaphrodites and four 46,XX males, and was examined for the presence of the SRY gene and for the loss of function (inactivating mutations and deletions) of DAX1 and WNT4 genes. In the second group consisting of thirty-three 46,XY sex-reversed patients we investigated the presence of inactivating mutations in the SRY and SF1 genes as well as the overexpression (duplication) of the DAX1 and WNT4 genes. The SRY gene was present in two 46,XX male patients and in none of the true hermaphrodites. Only one mutation, located outside homeobox domain of the 5' region of the HMG box of SRY (S18N), was identified in a patient with 46,XY sex reversal. A novel 8-bp microdeletion of the SF1 gene was identified in a 46,XY sex-reversed patient without adrenal insufficiency. The dosage of DAX1 and WNT4 was normal in the sex-reversed patients studied. We conclude that these genes are rarely involved in the etiology of male gonadal development in sex-reversed patients, a fact suggesting the presence of other genes in the sex determination cascade.

Quantitation of X-Y homologous genes in patients with schizophrenia by multiplex polymerase chain reaction.

The genetic basis of schizophrenia is obscure. In an XX male patient with schizophrenia we previously showed that one X;Y translocation breakpoint was in pseudoautosomal region 1 (PAR1) with the effect that the proximal segment of PAR1 from the PAR1 boundary to acetylserotonin N-methyl transferase (ASMT) distally was triplicated in this patient. This study determined whether dosage imbalances of X-Y homologous regions in general are associated with schizophrenia. A multiplex semi-quantitative polymerase chain reaction assay was developed to quantify MIC2 gene as a representative of PAR1 and compare it with the SYBL1 gene which maps in pseudoautosomal region 2 (PAR2) and protocadherin XY (PCDHXY), located at Xq21.3. Each of these three loci was co-amplified with the autosomal gene MSX2 using Cy5-labelled primers and the products separated by electrophoresis in polyacrylamide gels. Results were expressed as ratios of peak area of the target gene to MSX2 which served as an internal dosage control. Using genomes with sex chromosome aneuploidies, the method was found sensitive enough to detect a two-fold difference in gene copy number. We confirmed the MIC2 triplication in the XX male patient but found no significant difference in gene dosage of MIC2, PCDHXY and SYBL1 in a panel of 17 patients with schizophrenia compared to controls. No evidence was obtained for gene dosage imbalances in MIC2, PCDHXY and SYBL1 in patients with schizophrenia.

Prevalence and Molecular Analysis of Two Hot Spots for Ectopic Recombination Leading to XX Maleness

Two hot spots of ectopic Xp-Yp recombination have previously been shown to be at the origin of XX maleness (Well et al., 1994, Nature Genet. 7:414-419). To get more insight into the molecular basis of the abnormal interchange, 25 Y(+) class 3 XX male patients were studied. The two hot spots were found to account for the aberrant exchange in more than 50% of the cases. In addition, there is a correlation between the prevalence of each hot spot and the degree of X-Y homology between the corresponding fragments. Sequencing of the X-Y junctions in six patiens, who carried a breakpoint mapping in either of these two hot spot fragments, showed that their precise lacations were different from one individual to the other. In particular, the results obtained here in four new patients exclude the possibility that the repeated elements, present in these X-Y homologous fragments, are responsible for the high incidence of X-Y interchanges observed. Moreover, the breakpoints of all 25 class 3 XX males were found to be arranged in the same order on the X and Y chromosomes. This suggests that most ectopic recombinations leading to class 3 XX maleness involve X-Y homologous sequences persisting from an ancestral larger block of homology on the short arms of both sex chromosomes.

SRY alone can induce normal male sexual differentiation.

Most individuals with the rare 46,XX male "syndrome" arise due to an unequal interchange between Xp and Yp termini during paternal meiosis. The pattern of Y-sequences in these patients varies considerably, but very few cases have been reported showing only SRY. The phenotype in these patients is also variable ranging from severe impairment of the external genitalia through hypospadias and/or cryptorchidism to occasional normal male phenotype. We report a Mexican 46,XX male patient without genital ambiguities in whom DNA analysis showed the presence of SRY and the absence of ZFY. We conclude that in this case SRY alone was enough for complete male sexual differentiation.

Detection of the Testis Determining Factor in an XX Man

An XX male patient was examined for the presence of 25 loci on the Y chromosome. Only 2 loci, the proximal border of the pseudoautosomal region Y and the sex determining region Y, were detected in this patient. The other 23 loci, including the zinc finger protein Y, were absent. We presume that a crossing over between the X and Y chromosomes occurred at the region proximal to the sex determining region Y but distal to the zinc finger protein Y during meiosis of the father.

Deoxyribonucleic Acid Study On 2 Xx Male Patients

Two XX male patients were examined for the presence of 25 Y-specific loci using Southern blot analysis and polymerase chain reaction techniques. They were phenotypically normal male patients, although 1 had bilateral undescended testes and 1 had hypoplastic testes. The former patient had 10 of 11 loci on the short arm, while the latter lacked any of the 25 Y-specific loci. These findings may suggest the existence of a nonY gene that is controlled by the testis-determining factor on the short arm of the Y chromosome and has a role in the testis formation in man.