Male Genital Development Research Articles

A pilot study of the association between genetic polymorphisms involved in estrogen signaling and infant male genital phenotypes

Single nucleotide polymorphisms (SNPs) in genes that influence development of the male reproductive tract have been associated with male genitourinary abnormalities. However, no studies have tested the relationship between SNPs and intermediate phenotypes such as anogenital distance (AGD), anoscrotal distance (ASD) and penile width (PW). We tested whether 24 common SNPs in eight genes that influence male genital development were associated with intermediate phenotypes in 106 healthy male infants from the Study for Future Families. We used DNA from buccal smears and linear regression models to assess the relationship between anogenital measurements and SNP genotypes with adjustment for covariates. We found that the rs2077647 G allele, located in the coding region of estrogen receptor alpha (ESR1), was associated with a shorter AGD (P=0.02; -7.3 mm, 95% confidence interval (CI): -11.6 to -3.1), and the rs10475 T allele, located in the 3' untranslated region of activating transcription factor 3 (ATF3), was associated with a shorter ASD (-4.3 mm, 95% CI: -7.2 to -1.4), although this result was not significant (P=0.07) after controlling for multiple comparisons. We observed no association between PW and the SNPs tested. Minor alleles for two SNPs in genes that regulate estrogen signaling during male genital development were associated with AGD and ASD, although the significance of the association was marginal. Our findings suggest that AGD and ASD are influenced by heritable factors in genes known to be associated with frank male genital abnormalities such as hypospadias and cryptorchidism.

A Cell Model for Conditional Profiling of Androgen-Receptor-Interacting Proteins

Partial androgen insensitivity syndrome (PAIS) is associated with impaired male genital development and can be transmitted through mutations in the androgen receptor (AR). The aim of this study is to develop a cell model suitable for studying the impact AR mutations might have on AR interacting proteins. For this purpose, male genital development relevant mouse cell lines were genetically modified to express a tagged version of wild-type AR, allowing copurification of multiprotein complexes under native conditions followed by mass spectrometry. We report 57 known wild-type AR-interacting proteins identified in cells grown under proliferating and 65 under nonproliferating conditions. Of those, 47 were common to both samples suggesting different AR protein complex components in proliferating and proliferation-inhibited cells from the mouse proximal caput epididymus. These preliminary results now allow future studies to focus on replacing wild-type AR with mutant AR to uncover differences in protein interactions caused by AR mutations involved in PAIS.

URINARY MARKERS OF FETAL EXPOSURES TO PHTHALATES AND PHENOLS AND RISK OF MALE GENITAL ANOMALIES, IN EDEN AND PELAGIE MOTHER-CHILD COHORTS

Background and Aims: Exposure during pregnancy to chemicals suspected to disrupt hormonal signalling may alter male reproductive organogenesis, which is controlled by sex hormones. Our aim was to assess the association between risk of hypospadias and undescended testis at birth and fetal exposure to phthalates and phenols. Methods: We conducted a case-control study nested in two French mother-child cohorts (EDEN, PELAGIE) that enrolled pregnant women between 2002 and 2006. Cases were all boys members of the cohorts with hypospadias (n=21) or undescended testis (n=50) at birth. Three matched controls per case were selected among male singleton livebirths. Eleven phthalate metabolites and 9 phenols (EDEN cohort only) concentrations were measured in the pregnant mothers’ urine collected between the 3rd and the 28th gestational week. Conditional logistic analyses adjusted for maternal age, parity, educational level, gestational duration and urinary creatinine were carried out. Results: Prevalence ranged from 0.3% to 1.1% for hypospadias and from 0.7% to 5.1% for undescended testis according to residence area. Decreased risk of hypospadias, but not statistically significant, was observed with urinary concentrations of both low- and high-molecular-weight phthalate metabolites (p_trend, 0.13 and 0.10, respectively). No evidence of association (p_trend>0.10) with urinary concentrations of phthalates metabolites was observed for undescended testis risk. Risk of undescended testis increased with urinary concentrations of ethylparaben (OR_1st_tertile=reference; OR_2nd_tertile=1.5, 0.4-5.3; OR_3rd_tertile=2.9, 0.9-9.9; p_trend=0.08). Excluding preterm births led to similar conclusions. Conclusions: Our study did not support an impact of fetal exposure to phthalates or bisphenol A on male genital tract development. It however pointed out a possible role of ethylparaben on testicular descent which needs to be replicated by further studies since it is not supported by animal data. Our study was limited by a modest sample size but provided prenatal exposure assessment (that covers the gestational window of male reproductive organogenesis).

A Novel Entity of Clinically Isolated Adrenal Insufficiency Caused by a Partially Inactivating Mutation of the Gene Encoding for P450 Side Chain Cleavage Enzyme (CYP11A1)

Cytochrome P450 side-chain cleavage enzyme (CYP11A1) facilitates the first and rate-limiting step of steroidogenesis. Only nine patients with CYP11A1 deficiency have been described. All patients presented with adrenal insufficiency (AI) and disorder of sex development in 46,XY individuals. Our objective was to define the pathogenic consequences of a novel CYP11A1 mutation (p.R451W) found in two brothers with isolated adrenal insufficiency. The two brothers (46,XY) presented with AI and normal male genital development. The older boy first presented with signs and symptoms suggestive of AI at the age of 2.8 yr but was only diagnosed at the age of 4.1 yr during an adrenal crisis. The younger brother was diagnosed with AI at the age of 2.5 yr while being clinically asymptomatic. Both boys had entirely normal appearance of their external genitalia. The novel p.R451W mutation and five published missense CYP11A1 mutations were characterized employing two in vitro approaches using the natural substrate cholesterol and the intermediate 22R-hydroxycholesterol, respectively. Pregnenolone generation was measured by highly specific liquid chromatography tandem mass spectrometry. p.R451W had 30% of wild-type activity consistent with the clinical phenotype in our patients. Two previously published mutations (p.L222P and p.A359V) had 2- to 3-fold higher in vitro activities than originally reported, correlating better with the associated phenotypes. We provide the first evidence that partial CYP11A1 deficiency has to be considered as a differential diagnosis in clinically isolated adrenal insufficiency. Our assays demonstrate a tighter genotype-phenotype correlation in CYP11A1 deficiency than previous in vitro studies.

In utero exposure to bisphenol-A and anogenital distance of male offspring

Bisphenol-A (BPA) is an endocrine disruptor with widespread human exposure. The effect of in utero BPA exposure on human offspring remains largely unknown. Anogenital distance (AGD) of sons of workers who did or did not have occupational BPA exposure during pregnancy were compared in an occupational cohort study. Parental BPA exposure level during the index pregnancy was estimated through a job-exposure matrix based on personal air sampling measurement. Maternal exposure was considered direct in utero exposure to the fetus, whereas paternal exposure was considered indirect in utero exposure. A total of 153 boys were included in the final analysis, among them 56 with parental occupational exposure during pregnancy and 97 without. After controlling for the boys' ages and weights using linear regression, parental occupational exposure to BPA during pregnancy was associated with shortened AGD in male offspring. The association was stronger for maternal exposure (p < 0.01). There was also a dose-response relationship with increased BPA exposure levels in pregnancy associated with greater magnitude of shortened AGD in male offspring, with a statistically significant trend for the association (p = 0.008). Our findings provide the first epidemiologic evidence that in utero BPA exposure may adversely affect male genital development.

Why Boys Will Be Boys: Two Pathways of Fetal Testicular Androgen Biosynthesis Are Needed for Male Sexual Differentiation

Human sexual determination is initiated by a cascade of genes that lead to the development of the fetal gonad. Whereas development of the female external genitalia does not require fetal ovarian hormones, male genital development requires the action of testicular testosterone and its more potent derivative dihydrotestosterone (DHT). The "classic" biosynthetic pathway from cholesterol to testosterone in the testis and the subsequent conversion of testosterone to DHT in genital skin is well established. Recently, an alternative pathway leading to DHT has been described in marsupials, but its potential importance to human development is unclear. AKR1C2 is an enzyme that participates in the alternative but not the classic pathway. Using a candidate gene approach, we identified AKR1C2 mutations with sex-limited recessive inheritance in four 46,XY individuals with disordered sexual development (DSD). Analysis of the inheritance of microsatellite markers excluded other candidate loci. Affected individuals had moderate to severe undervirilization at birth; when recreated by site-directed mutagenesis and expressed in bacteria, the mutant AKR1C2 had diminished but not absent catalytic activities. The 46,XY DSD individuals also carry a mutation causing aberrant splicing in AKR1C4, which encodes an enzyme with similar activity. This suggests a mode of inheritance where the severity of the developmental defect depends on the number of mutations in the two genes. An unrelated 46,XY DSD patient carried AKR1C2 mutations on both alleles, confirming the essential role of AKR1C2 and corroborating the hypothesis that both the classic and alternative pathways of testicular androgen biosynthesis are needed for normal human male sexual differentiation.

Foetal exposure to phthalate esters and anogenital distance in male newborns

Phthalate esters, commonly used as plasticizers, show anti-androgenic activity and cause male reproductive malformation in experimental animals. However, the effects of prenatal exposure to phthalate esters in humans have not been extensively studied. The purpose of this study was to examine the relationship between prenatal exposure to phthalate esters and the anogenital distance (AGD) as a reproductive endpoint in human male newborns. Spot urine samples were collected from 111 Japanese pregnant women after obtaining their informed consent. Seven urinary phthalate ester metabolites were determined by high performance liquid chromatography-tandem mass spectrometry. Urinary isoflavones concentrations were measured as possible covariates because their oestrogenicities and high exposure levels among Japanese have the potential to affect male genital development. Birth outcomes and AGD, the distance from the centre of the anus to external genitalia, were measured for their male newborns. In a multiple regression model, the log-transformed mono-2-ethylhexyl phthalate concentration (specific gravity-corrected) was negatively significant, and maternal smoking status was positively significant, in explaining anogenital index (AGI) when potential covariates were controlled for. Urinary isoflavones did not significantly contribute to AGI in any models. Our results suggest that prenatal exposure to di(2-ethylhexyl) phthalate affects reproductive development in human males.

Prevalence of hypospadias in grandsons of women exposed to diethylstilbestrol during pregnancy: a multigenerational national cohort study

Prenatal diethylstilbestrol (DES)-exposed mice have raised the suspicion of a transgenerational effect in the occurrence of genital malformation in males. This nationwide cohort study in collaboration with a French association of DES-exposed women studied 529 families and showed that a significant proportion of boys born to DES daughters exhibited hypospadias with no other molecular defects identified.

Immunohistochemical Characterization of Sonic Hedgehog and its Downstream Signaling Molecules During Human Penile Development

Hypospadias is a common congenital anomalies, yet its molecular basis remains unknown. Recent studies have linked perturbations in the Hedgehog signaling pathway to hypospadias. However, the expression of Sonic hedgehog (Shh) has not been reported during genital development. Immunohistochemical staining for Shh and its receptors was applied to 10 human fetal penises ranging from 12 to 29 weeks gestation. The intensity of Shh staining was greatest in the urethral epithelium at 14 weeks gestation, correlating with the time of urethral tubularization. Results suggest a role for Shh in human male genital development.

Second to fourth digit ratios, male genital development and reproductive health: a clinical study among fertile men and testis cancer patients

Second- to fourth-digit length ratio, 2D:4D, is a marker of testosterone level during foetal life that was found associated with sperm concentration or testosterone levels in some studies, but not in others, a difference possibly related to the way the ratio is assessed. In this study, 2D:4D was assessed in 122 men partners of pregnant women and in 71 testicular cancer patients using a new method based on direct measurements of finger lengths. In addition, we investigated the association between 2D:4D, birth weight, testicular volume, semen quality and time to pregnancy. A validation study of the method demonstrated high reliability and reproducibility. Neither digit lengths nor 2D:4D significantly differed in both groups of men. We found a significant negative association between 2D:4D and birth weight in testicular cancer patients. In fertile men, 2D:4D was associated with testicular volume (r=-0.36, p<0.001), total sperm number (r=-0.18, p=0.04) and time to pregnancy (r=0.24, p<0.02). In addition, participants with a history of epididymal cyst had a significantly higher 2D:4D than those without cysts. In conclusion, all significant findings indicate that the human male reproductive function is negatively related to 2D:4D. However, 2D:4D for testicular cancer patients does not point to a hormonal imbalance during foetal life as the common cause for developing germ-cell cancer. Such results obtained, thanks to an easy, direct and reliable method for measuring finger lengths, suggest the usefulness of this new tool in fertility studies as well as for studying men with developmental disorders of the reproductive tract.

Endocrine disrupters and male infertility: is there a causative link?

Event Abstract Back to Event Endocrine disrupters and male infertility: is there a causative link? Dimitrios G. Goulis1* 1 Aristotle University of Thessaloniki, First Department of Obstetrics and Gynecology, Greece Cryptorchidism is the most frequent congenital birth defect in boys, with a prevalence of 3%. Although it is considered a mild malformation, it represents a risk factor for reduced fertility and testicular cancer. Its etiology remains unknown. Major regulators of testicular descent from intra-abdominal location into the bottom of the scrotum are testosterone and insulin-like factor 3. Mutations in the gene for insulin-like factor 3 and its receptor as well as in the androgen receptor gene have been recognized as causes of cryptorchidism. Environmental factors acting as endocrine disruptors of testicular descent might also contribute to its etiology and increased incidence in recent years. Furthermore, polymorphisms in different genes have recently been investigated as contributing risk factors for cryptorchidism, alone or by influencing susceptibility to endocrine disruptors. Obviously, the interaction of environmental and genetic factors is fundamental. Hypospadias is another genital malformation in the male newborn and results from an abnormal penile and urethral development. This malformation appears to be at the crossroads of various mechanisms implicating genetic and environmental factors. The genes of penile development (HOX, FGF, Shh) and testicular determination (WT1, SRY) and those regulating the synthesis (LH receptor) and action of androgen (5-alpha reductase, androgen receptor) can cause hypospadias, if altered. Besides these genomic and hormonal factors, multiple substances found in the environment can also potentially interfere with male genital development due to their similarity to hormones. An interaction between genetic background and environment is likely. The testicular dysgenesis syndrome (TDS) hypothesis proposes that a proportion of the male reproductive disorders, such as cryptorchidism, hypospadias, infertility and testicular cancer, may be symptoms of one underlying developmental disease, TDS, which is most likely a result of disturbed gonadal development in the embryo. TDS may be caused by genetic factors, environmental / life-style factors, or a combination of both. Among the environmental and life-style factors that are suspected to influence the hormonal milieu of the developing gonad are the endocrine disrupters. A prenatal exposure to commonly used chemicals, such as phthalates, may result in a TDS-like phenotype in rats. In humans, the situation is more complex and TDS exists in a wide range of phenotypes: from the mildest and most common form, in which impaired spermatogenesis is the only symptom, to the most severe cases, in which the patient may develop testicular cancer. It is of great importance that clinicians in different specialties treating patients with TDS are aware of the association between the different symptoms. Keywords: Endocrine disrupters, male infertility, Reproduction Conference: 8th Southeast European Congress on Xenobiotic Metabolism and Toxicity - XEMET 2010, Thessaloniki, Greece, 1 Oct - 5 Oct, 2010. Presentation Type: Invited speaker Topic: Xenobiotics and reproduction Citation: Goulis DG (2010). Endocrine disrupters and male infertility: is there a causative link?. Front. Pharmacol. Conference Abstract: 8th Southeast European Congress on Xenobiotic Metabolism and Toxicity - XEMET 2010. doi: 10.3389/conf.fphar.2010.60.00126 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 28 Oct 2010; Published Online: 04 Nov 2010. * Correspondence: Dr. Dimitrios G Goulis, Aristotle University of Thessaloniki, First Department of Obstetrics and Gynecology, Thessaloniki, Greece, dimitrios.goulis@otenet.gr Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Dimitrios G Goulis Google Dimitrios G Goulis Google Scholar Dimitrios G Goulis PubMed Dimitrios G Goulis Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Parental occupational exposure to potential endocrine disrupting chemicals and risk of hypospadias in infants

ObjectivesHypospadias is a urogenital birth defect affecting infant boys. Periconceptual parental occupational exposure to endocrine disrupting chemicals (EDCs) with oestrogenic or anti-androgenic properties may adversely affect male genital development in...

Frasier syndrome, a potential cause of end-stage renal failure in childhood

The diagnosis of Frasier syndrome is based on the association of male pseudohermaphroditism (as a result of gonadal dysgenesis), with steroid-resistant nephrotic syndrome due to focal and segmental glomerular sclerosis (FSGS), which progresses to end-stage renal failure (ESRF) during adolescence or adulthood. Frasier syndrome results from mutations in the Wilms' tumour suppressor gene WT1, which is responsible for alterations in male genital development and podocyte dysfunction. We describe the case of a 7-year-old girl who was referred to the paediatric emergency department with ESRF. Haemodialysis was started immediately because of severe hypertension and hyperkalaemia. In view of the fact that our patient had a past medical history of pseudohermaphroditism, we suspected that the acute presentation in ESRF may be related to a new diagnosis of Frasier syndrome. Our hypothesis was confirmed on examination of the medical records. There had been no medical follow-up for several years and, in particular, no renal imaging or functional assessment had ever been performed. This lack of surveillance explains why our patient presented with ESRF much earlier in this disease than expected and subsequently had to undergo kidney transplantation at a very young age.

Estrogen Effects on Fetal Penile and Urethral Development in Organotypic Mouse Genital Tubercle Culture

We developed an organotypic genital tubercle culture system in vitro and used it to investigate the direct effects of the hyperestrogenic state on fetal mouse penile and urethral development. Genital tubercles were dissected from embryonic day 14.5 C57B/L6 male mouse fetuses and cultured using an air-liquid interface on a microporous membrane support soaked in synthetic medium. Cultures were separated into 4 groups. Groups 1 to 3 were supplied with 10 nM dihydrotestosterone, estradiol and 10 nM dihydrotestosterone plus estradiol, respectively. Group 4 was cultured in hormone-free medium. After 36 to 72-hour culture morphological, histological, proliferation, apoptosis, androgen signaling and activating transcription factor 3 analyses were done. The physiological concentration of 10 nM dihydrotestosterone was essential for genital tubercle growth in vitro. Androgen induced growth and urethral development were significantly suppressed by high dose estrogen. Concurrently we observed increased apoptosis and decreased proliferation in the mesenchyma. Androgen signaling was disrupted and activating transcription factor 3, a factor related to hypospadias genesis, was up-regulated. High dose estrogen suppressed male genital tubercle development in vitro. The organotypic genital tubercle culture system in vitro consisting of urethral epithelial and mesenchymal cells can recapitulate the hormonal sensitivity of fetal penile and urethral development. This method is potentially useful for studying the effects of various factors, particularly endocrine disruptors.

Is hypospadias a genetic, endocrine or environmental disease, or still an unexplained malformation?

Hypospadias is one of the most frequent genital malformations in the male newborn and results from an abnormal penile and urethral development. This process requires a correct genetic programme, time- and space-adapted cellular differentiation, complex tissue interactions, and hormonal mediation through enzymatic activities and hormonal transduction signals. Any disturbance in these regulations may induce a defect in the virilization of the external genitalia and hypospadias. This malformation thus appears to be at the crossroads of various mechanisms implicating genetic and environmental factors. The genes of penile development (HOX, FGF, Shh) and testicular determination (WT1, SRY) and those regulating the synthesis [luteinizing hormone (LH) receptor] and action of androgen (5alpha reductase, androgen receptor) can cause hypospadias if altered. Several chromosomal abnormalities and malformative syndromes include hypospadias, from anterior to penoscrotal forms. More recently, CXorf6 and ATF3 have been reported to be involved. Besides these genomic and hormonal factors, multiple substances found in the environment can also potentially interfere with male genital development because of their similarity to hormones. The proportion of hypospadias cases for which an aetiology is detected varies with the authors but it nevertheless remains low, especially for less severe cases. An interaction between genetic background and environment is likely.

Prenatal Equol Treatment: Does Not Alter Newborn Genital Development but Alters Depressive‐like Behaviors in Pre‐pubertal Offspring.

In this study we examined equol, the intestinal metabolite of daidzein, and its effects on genital morphology and subsequent postnatal behavior. Pregnant rats were divided into six treatment groups before subcutaneous injection treatments: 1) Non‐injected Controls, 2) dimethyl sulfoxide (DMSO)‐injected controls, 3) Equol at 10.5 mg/Kg (in DMSO), 4) Equol at 21.0 mg/Kg (in DMSO), 5) Equol at 63.0 mg/Kg (in DMSO), and 6) Flutamide (90 mg/Kg, androgen‐receptor blocker). The mothers received daily treatment injections from gestational days 14‐20. After birth, body weight and anogenital distance (AGD) were measured and sex determined. Some of the pups and mothers (a subset at birth) were sacrificed and trunk blood collected for determining isoflavone and 5alpha‐dihydrotestosterone (5alpha‐DHT) levels. The remainder of the pups developed postnatally until day 29 where animals underwent analysis via the Porsolt forced swim test to determine depressive‐like behaviors. In summary, equol exposure during late pregnancy 1) does not alter 5alpha‐DHT levels in the males or females or alter male external genital development, 2) the high equol dosed mothers weighed the least before and after birth and their offspring also weighed the least at birth, and 3) after birth the high dosed equol treated animals that gained the most weight postnatally displayed the least depressive‐like behaviors. (USDA 2004‐01811)

Identification of potential mechanisms of toxicity after di-(2-ethylhexyl)-phthalate (DEHP) adult exposure in the liver using a systems biology approach

Phthalates are industrial additives widely used as plasticizers. In addition to deleterious effects on male genital development, population studies have documented correlations between phthalates exposure and impacts on reproductive tract development and on the metabolic syndrome in male adults. In this work we investigated potential mechanisms underlying the impact of DEHP on adult mouse liver in vivo. A parallel analysis of hepatic transcript and metabolic profiles from adult mice exposed to varying DEHP doses was performed. Hepatic genes modulated by DEHP are predominantly PPARα targets. However, the induction of prototypic cytochrome P450 genes strongly supports the activation of additional NR pathways, including Constitutive Androstane Receptor (CAR). Integration of transcriptomic and metabonomic profiles revealed a correlation between the impacts of DEHP on genes and metabolites related to heme synthesis and to the Rev-erbα pathway that senses endogenous heme level. We further confirmed the combined impact of DEHP on the hepatic expression of Alas1, a critical enzyme in heme synthesis and on the expression of Rev-erbα target genes involved in the cellular clock and in energy metabolism. This work shows that DEHP interferes with hepatic CAR and Rev-erbα pathways which are both involved in the control of metabolism. The identification of these new hepatic pathways targeted by DEHP could contribute to metabolic and endocrine disruption associated with phthalate exposure. Gene expression profiles performed on microdissected testis territories displayed a differential responsiveness to DEHP. Altogether, this suggests that impacts of DEHP on adult organs, including testis, could be documented and deserve further investigations.

EHP Papers of the Year, 2008

<i>EHP</i> Papers of the Year, 2008

Reliability of pubertal self‐assessment in Hong Kong Chinese children

To validate a Tanner stages self-assessment questionnaire using gender-specific line drawings and brief explanatory text in Chinese. A cross sectional study design. One primary and two secondary schools. 172 boys and 182 girls aged between 8 and 18 years. Students' self-assessments of pubertal maturation were compared with assessments made by a same gender rater using visual depiction physical examination. Raters' physical examinations were performed after the children had answered the self-assessment questionnaire individually and in private. Raters were blinded to the self-assessment results. Accuracy rates and weighted kappa statistic were used to evaluate the degree of agreement between children and raters. Substantial to almost perfect agreement was found between self- and rater's assessments of breast development and pubic hair growth in girls [weighted kappa 0.72 (P < 0.0001, 95% CI 0.66, 0.79) and 0.83 (P < 0.0001, 95% CI 0.78, 0.87) respectively]. Moderate to substantial agreement was found between self- and rater's assessments of male genital development and pubic hair growth [weighted kappa 0.58 (P < 0.0001, 95% CI 0.48, 0.68) and 0.80 (P < 0.0001, 95% CI 0.74, 0.86) respectively]. Most agreements between self- and rater's assessments differed by only one Tanner stage. Agreement was higher for girls than boys. Girls tended to overestimate their breast stages and boys tended to underestimate their genitalia development. This study confirms that a Tanner pubertal self-assessment questionnaire with line drawings and explanatory Chinese text can reliably estimate sexual maturation status in Hong Kong Chinese children.

Activating transcription factor 3: a hormone responsive gene in the etiology of hypospadias

Hypospadias is a common inborn error of the genital development, whose complex etiology remains elusive. Defects of the androgen metabolism and activity have been found in a subset of boys with hypospadias. Moreover, the balance between androgens and estrogens seems to be important to the proper male genital development. Activating transcription factor 3 (ATF3), an estrogen responsive gene, has been reported to be expressed during sexual development and up-regulated in hypospadic genital skin. We investigated ATF3 as a candidate gene for hypospadias. Genotyping of eight-tagged single nucleotide polymorphisms (SNP)s was performed in 330 boys with hypospadias and in 380 healthy controls. Screening for mutations in ATF3 was conducted in a subset of boys with hypospadias. ATF3 expression was evaluated in the foreskin of boys with hypospadias and in healthy controls and in the human fetal genitalia by immunohistochemistry. Three common SNPs, spanning a region of about 16 kb in intron 1 of ATF3, are associated with hypospadias. These SNPs are not linked and their effects are independent. The combination of the three risk SNPs yields the highest significance. Mutation screening identified the gene variant c536A>G in one patient and c817C>T in the 3'-UTR in two other patients. ATF3 expression was evidenced in the developing male urethra. ATF3 gene variants influence the risk of hypospadias. Its hormonal responsiveness may underlie this risk effect. But also other ATF3-dependent biological aspects, such as cell survival and death, response to stress stimuli, or the control of epithelial-mesenchymal interactions, may be of importance.