Canalization of the Vestibular Plate in the Absence of Urethral Fusion Characterizes Development of the Human Clitoris: The Single Zipper Hypothesis.

PURPOSE: We characterized the early gestation development of the female external genitalia using optical projection tomography to visualize anatomical structures at high resolution. MATERIALS AND METHODS: First and early second trimester human female fetal external genitalia were collected with consent after voluntary termination. Specimens labeled with anti-E-Cadherin antibody underwent analysis with optical projection tomography. Histological sections were immunostained for androgen receptor, 5α-reductase, Ki67 for proliferation and Caspase 3 for apoptosis. RESULTS: Three-dimensional reconstructions demonstrated proximal to distal canalization of the epithelial vestibular plate and formation of a vestibular groove, which remained open. Ki67 was observed throughout with greatest density in the dorsal vestibular plate at the level of the opening groove. Staining for Caspase 3 was minimal in all sections. Androgen receptor staining was seen throughout the mesenchyme and in the apical epithelium of the dorsal vestibular groove. Throughout the epithelium and epidermis 5α-reductase staining was observed. CONCLUSIONS: Early development of the external genitalia in the female is analogous to that in the male, demonstrating a similar opening zipper driving canalization of the vestibular plate with localized epithelial proliferation in the absence of significant apoptosis. Thus we hypothesize that the mechanism underlying the opening zipper must be androgen independent and the absence of androgen driven urethral fusion characterizes the normal development of the human clitoris.

Canalization of the urethral plate precedes fusion of the urethral folds during male penile urethral development: the double zipper hypothesis.

PURPOSE: We describe the "double zipper" mechanism of human male urethral formation, where the distal zipper opens the urethral groove through canalization of the urethral plate, and a second closing zipper follows behind and closes the urethral groove to form the tubular urethra. MATERIALS AND METHODS: Anonymous human fetal genital specimens were acquired and gender was determined by polymerase chain reaction of the Y chromosome. Specimens were processed for optical projection tomography, stained with E-cadherin, Ki67 and caspase 3, and imaged. RESULTS: Eight developing male fetal specimens from 6.5 to 16.5 weeks of gestation were analyzed by optical projection tomography, and an additional 5 specimens by serial sections. Phallus length ranged from 1.3 to 3.7 mm. The urethral plate canalized into a groove with 2 epithelial edges that subsequently fused. Ki67 staining was localized to the dorsal aspect of the urethral plate. In contrast, caspase 3 staining was not observed. The entire process was completed during a 10-week period. CONCLUSIONS: The human male urethra appears to form by 2 mechanisms, an initial "opening zipper" that facilitates distal canalization of the solid urethral plate to form the urethral groove, which involves a high rate of epithelial proliferation (apoptosis not observed), and a "closing zipper" facilitating fusion of the 2 epithelial surfaces of the urethral groove, and thus extending the penile urethra distally. Improved knowledge of the molecular mechanisms of these processes is critical to understanding mechanisms of abnormal urethral development, such as hypospadias.

Do endocrine disruptors cause hypospadias?

INTRODUCTION: Endocrine disruptors or environmental agents, disrupt the endocrine system, leading to various adverse effects in humans and animals. Although the phenomenon has been noted historically in the cases of diethylstilbestrol (DES) and dichlorodiphenyltrichloroethane (DDT), the term "endocrine disruptor" is relatively new. Endocrine disruptors can have a variety of hormonal activities such as estrogenicity or anti-androgenicity. The focus of this review concerns on the induction of hypospadias by exogenous estrogenic endocrine disruptors. This has been a particular clinical concern secondary to reported increased incidence of hypospadias. Herein, the recent literature is reviewed as to whether endocrine disruptors cause hypospadias. METHODS: A literature search was performed for studies involving both humans and animals. Studies within the past 5 years were reviewed and categorized into basic science, clinical science, epidemiologic, or review studies. RESULTS: Forty-three scientific articles were identified. Relevant sentinel articles were also reviewed. Additional pertinent studies were extracted from the reference of the articles that obtained from initial search results. Each article was reviewed and results presented. Overall, there were no studies which definitely stated that endocrine disruptors caused hypospadias. However, there were multiple studies which implicated endocrine disruptors as one component of a multifactorial model for hypospadias. CONCLUSIONS: Endocrine disruption may be one of the many critical steps in aberrant development that manifests as hypospadias.

Peyronie's disease compromises the durability and component-malfunction rates in patients implanted with an inflatable penile prosthesis.

OBJECTIVE: To compare the durability and complication rates of surgery to implant an inflatable penile prosthesis (IPP) between patients with and without Peyronie's disease (PD). PATIENTS AND METHODS: We retrospectively reviewed all patients undergoing IPP surgery at one centre (Memphis) between July 1997 and May 2007. Variables included age at surgery, race, body mass index, presence of PD, brand/type of IPP (two vs three pieces), presence of diabetes mellitus (DM), active tobacco use, and complications. The result were assessed using t-tests, chi-square and regression analysis, with P < 0.05 considered to indicate significant differences. RESULTS: In all, 79 men were analysed (mean age 59.8 years, range 38.1-81.5). Nine (11%) patients had PD and had a IPP implanted, with penile modelling. Overall, 43 (54%) patients had pre-existing DM and 51 (65%) actively used tobacco. At a mean (range) follow-up of 19.6 (0.1-115.3) months, six (8%) patients had component malfunctions. Of these, three had DM and four actively smoked. Of the nine patients with PD, three developed component malfunctions, vs three (4%) who did not have PD (P= 0.002). Both groups had similar infection rates (P= 0.98). The mean (range) time to component malfunction was 4.3 (0.1-9.6) months, which was longer (but not significantly) in the PD group, with a mean (median, range) of 10.9 (6.3, 1.1-9.6) months, than the 3.0 (1.0, 0.2-7.9) months in the group without PD (P= 0.4). Groups were matched for rates of DM (P= 0.1) and tobacco use (P= 0.2). PD was a significant predictor of component malfunction on both univariate (P= 0.001) and multivariate analysis (P= 0.002) when adjusting for age (P= 0.2), body mass index (P= 0.7), DM (P= 0.3) and tobacco use (P= 0.8). CONCLUSION: Patients with PD implanted with a IPP, with penile modelling, had significantly higher component malfunction rates. Further, PD independently predicted component malfunction. These findings might be related to stress on the device at the time of surgery, during use, or both. Further study into this relationship is required.

Formation and mass spectrometric analysis of DNA and nucleoside adducts by S-(1-acetoxymethyl)glutathione and by glutathione S-transferase-mediated activation of dihalomethanes.

The dihalomethane CH(2)Cl(2) is an industrial solvent of potential concern to humans because of its potential genotoxicity and carcinogenicity. To characterize DNA damage by dihalomethanes, a rapid DNA digestion under acidic conditions was developed to identify alkali labile DNA-dihalomethane nucleoside adducts using HPLC-electrospray mass spectrometry. DNA digestion worked best using pH 5.0 sodium acetate buffer, a 30 min incubation with DNase II and phosphodiesterase II, and a 2 h acid phosphatase digest. DNA was modified with S-(1-acetoxymethyl)glutathione (GSCH(2)OAc), a reagent modeling activated dihalomethanes. Adducts to G, A, and T were detected at high ratios of GSCH(2)OAc/DNA following digestion of the DNA with the procedure used here. The relative efficacy of adduct formation was G > T > A >> C. The four DNA nucleosides were also reacted with the dihalomethanes CH(2)Cl(2) and CH(2)Br(2) in the presence of glutathione (GSH) and GSH S-transferases from bacteria (DM11), rat (GST 5-5), and human (GST T1-1) under conditions that produce mutations in bacteria. All enzymes formed adducts to all four nucleosides, with dGuo being the most readily modified nucleoside. Thus, the pattern paralleled the results obtained with the model compounds GSCH(2)OAc and DNA. CH(2)Cl(2) and CH(2)Br(2) yielded similar amounts of adducts under these conditions. The relative efficiency of adduct formation by GSH transferases was rat 5-5 > human T1-1 > bacterial DM11, showing that human GSH transferase T1-1 can form dihalomethane adducts under the conditions used. Although the lability of DNA adducts has precluded more sophisticated experiments and in vivo studies have not yet been possible, the work collectively demonstrates the ability of several GSH transferases to generate DNA adducts from dihalomethanes, with G being the preferred site of adduction in both this and the GSCH(2)OAc model system.