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

Abstract

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.

Complex epithelial remodeling underlie the fusion event in early fetal development of the human penile urethra

“…In agreement with our previous work using optical projection tomography to visualize epithelial structures in developing human genitalia, 1,2 high resolution SEM imaging of human fetal specimens demonstrates proximal to distal “unzipping” or canalization of the urethral plate in both the developing urethral plate of the penis and the vestibular plate in the clitoris. In contrast, a subsequent proximal to distal fusion of the urethral folds is seen only in the male with no evidence of the analogous process in the female.…”

“…The “opening zipper” refers to the proximal to distal canalization of the urethral plate to form the urethral groove in males and the opening of the vestibular plate into the vestibular groove in females, and is likely to be an androgen-independent event. 1,2 The “closing zipper” refers to the proximal to distal closure of the urethral groove in males to form the penile urethra, and thus is presumed to be androgen dependent. 1 This fusion of the urethral folds is the key event that distinguishes the development of male from female human external genitalia in that the closing zipper mechanism is not evident during normal female development.…”

“…1 This fusion of the urethral folds is the key event that distinguishes the development of male from female human external genitalia in that the closing zipper mechanism is not evident during normal female development. 2 …”

“…Using optical projection tomography, we were previously able to accurately visualize the internal epithelial structures in developing human male and female external genitalia. 1,2 However, the detail in these 3D reconstructions is limited by the ability of labeled antibodies to penetrate fixed human fetal tissue and by the physical resolution limits of light microscopy. Based on immunostained sections from developing human fetal genitalia, we recognized that the mechanistic details of urethral plate canalization and urethral fold fusion were more intricate than we were able to visualize using wholemount fluorescence microscopy.…”

Renal Subcapsular xenografing of human fetal external genital tissue – A new model for investigating urethral development

“…Recent studies by our group have elucidated mechanisms of human penile urethral and clitoral development [62, 63]. The urethra in the penile shaft forms via distal canalization of a solid urethral plate to form a wide-open urethral groove, followed by proximal to distal fusion of the urethral folds, a process far more complicated than originally envisioned [64].…”