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

Abstract

We recently described a two-step process of urethral plate canalization and urethral fold fusion to form the human penile urethra. Canalization (“opening zipper”) opens the solid urethral plate into a groove, and fusion (“closing zipper”) closes the urethral groove to form the penile urethra. We hypothesize that failure of canalization and/or fusion during human urethral formation can lead to hypospadias. Herein, we use scanning electron microscopy (SEM) and analysis of transverse serial sections to better characterize development of the human fetal penile urethra as contrasted to the development of the human fetal clitoris. Eighteen 7-13 week human fetal external genitalia specimens were analyzed by SEM, and fifteen additional human fetal specimens were sectioned for histologic analysis. SEM images demonstrate canalization of the urethral/vestibular plate in the developing male and female external genitalia, respectively, followed by proximal to distal fusion of the urethral folds in males only. The fusion process during penile development occurs sequentially in multiple layers and through the interlacing of epidermal “cords”. Complex epithelial organization is also noted at the site of active canalization. The demarcation between the epidermis of the shaft and the glans becomes distinct during development, and the epithelial tag at the distal tip of the penile and clitoral glans regresses as development progresses. In summary, SEM analysis of human fetal specimens supports the two-zipper hypothesis of formation of the penile urethra. The opening zipper progresses from proximal to distal along the shaft of the penis and clitoris into the glans in identical fashion in both sexes. The closing zipper mechanism is active only in males and is not a single process but rather a series of layered fusion events, uniquely different from the simple fusion of two epithelial surfaces as occurs in formation of the palate and neural tube.

Imaging the developing human external and internal urogenital organs with light sheet fluorescence microscopy

“…Recent work from our group using optical projection tomography (OPT) challenged prior dogma regarding mechanisms of development of the human penile urethra and analogous structures in the clitoris. We demonstrated that two sequential distal canalization and proximal fusion processes are involved rather than a single fusion process of paired urethral folds (Li et al, 2015;Shen et al, 2016). Prior to our use of 3D reconstruction, this Despite these advances, differential protein localization within developing human urogenital organs remains poorly characterized in three dimensions.…”

“…3). This is followed by the subsequent fusion of multiple interlacing ventral epithelial "cords" to form the mature tubular penile urethra (Li et al, 2015;Shen et al, 2016). As described prior, SPIM has since supplanted the use of OPT in our laboratory (Isaacson et al, 2018a).…”

“…Recent investigations by our group employing whole-mount, 3D imaging techniques have elucidated the mechanisms by which the human external genitalia form during the late embryonic and early fetal periods (Li et al, 2015;Shen et al, 2016;Isaacson et al, 2018a). We recently reviewed the workings and applications of these techniques (Isaacson et al, 2018a).…”

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

“…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]. This results in formation of the tubular penile urethra and superficial midline penile raphe.…”

Flutamide-induced hypospadias in rats: A critical assessment

“…However, in mice and rats the urethral plate does not extend into the distal aspect of the developing penis (Hynes and Fraher, 2004a, b;Sinclair et al, 2016bSinclair et al, , 2016aKluth et al, 2011), and thus a different morphogenetic mechanism is utilized to generate the distal aspect of the penile urethra in mice. In the developing human penis, the urethral plate canalizes to form a widely open urethral groove, whose edges (urethral folds) fuse in the ventral midline to form the tubular penile urethra (Li et al, 2015b;Shen et al, 2016). In mice a ventral groove (the preputial-urethral groove) forms in the distal aspect of the developing mouse penis independent of and distal to the urethral plate.…”