Estrogens and development of the mouse and human external genitalia - PubMed (original) (raw)

Review

Estrogens and development of the mouse and human external genitalia

Laurence Baskin et al. Differentiation. 2021 Mar-Apr.

Abstract

The Jost hypothesis states that androgens are necessary for normal development of the male external genitalia. In this review, we explore the complementary hypothesis that estrogens can elicit abnormal development of male external genitalia. Herein, we review available data in both humans and mice on the deleterious effects of estrogen on external genitalia development, especially during the "window of susceptibility" to exogenous estrogens. The male and female developing external genitalia in both the human and mouse express ESR1 and ESR2, along with the androgen receptor (AR). Human clinical data suggests that exogenous estrogens can adversely affect normal penile and urethral development, resulting in hypospadias. Experimental mouse data also strongly supports the idea that exogenous estrogens cause penile and urethral defects. Despite key differences, estrogen-induced hypospadias in the mouse displays certain morphogenetic homologies to human hypospadias, including disruption of urethral fusion and preputial abnormalities. Timing of estrogenic exposure, or the "window of susceptibility," is an important consideration when examining malformations of the external genitalia in both humans and mice. In addition to a review of normal human and mouse external genital development, this article aims to review the present data on the role of estrogens in normal and abnormal development of the mouse and human internal and external genitalia. Based on the current literature for both species, we conclude that estrogen-dependent processes may play a role in abnormal genital development.

Keywords: Estrogen; External genitalia development; Mouse, and human.

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Figures

Fig. 1.

Jost Hypothesis of Sexual Differentiation. T = Testosterone, MIS = Mullerian inhibiting substance.

Fig. 2.

Human Urethral Development from the Indifferent Stage at 8 weeks Gestation to Mature Development at 16 weeks gestation Using Optical Projection Tomography. Note complete formation of the tubular urethra in the penile shaft by fusion of the urethral folds and formation of the glanular urethra extending to the tip of the penis (left column). In contrast, the vestibular groove remains open in females and forms the labia minora (right column) (Overland et al., 2016; Li et al., 2015).

Fig. 3.

Adult CD1 Mouse Penis. Note the penis in the non-erect state is an internal structure housed well within the external preputial space. Position of the tip of the MUMP in the resting state is accurately placed (Cunha et al., 2020c).

Fig. 4.

Mechanism of Mouse Urethral Formation. The proximal portion of the urethra within the glans develops via direct canalization of the urethral plate (A–D), while the distal aspect of the mouse penile urethra and the urethral meatus form via epithelial fusion (E–H). Finer details of these morphogenetic process can be found in Liu et al. (2018) (Liu et al., 2018b).

Fig. 5.

Mechanism of Human Penile Urethral Formation. The distal portion of the human penile urethra (within the glans) develops via direct canalization of the urethral plate (A–D), while the proximal portion of the human penile urethra within the shaft forms via canalization of the urethral plate to form the urethral groove, followed by fusion of the urethral folds (E–H). Finer details of these morphogenetic process can be found in Baskin et al., Shen, et al., Cunha, et al., and Liu et al., 2018 (Baskin et al., 2018a; Liu et al., 2018b; Cunha et al., 2019b; Shen et al., 2016).

Fig. 6.

(A) Gross micrograph, lateral view of an adult CD-1 mouse perineum. Note the female prepuce (white arrowhead) and the overlaid position of the reconstructed mouse clitoris in blue positioned accurately. Representative transverse histologic sections of adult mouse clitoris: (B) proximal, (C) and (D) distal as indicated. (B) The os clitoris is dorsal and completely separate from the urethra. (C) The U-shaped clitoral lamina is separated from the urethral epithelium stroma (small black arrow). (D) The clitoral epithelial lamina defining the clitoral epithelium is tethered to urethral epithelium (Cunha et al., 2020b). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 7.

Child with 5α reductase type 2 deficiency. Note severe hypospadias (arrowhead = pseudo vagina, arrow = ectopic urethral meatus).

Fig. 8.

Human Hypospadias occurs on a spectrum from mild to severe forms (panel A is least severe through panel F, which is most severe), based on the location of the ectopic urethral meatus (white arrows), associated penile curvature, and dorsal hooded asymmetric foreskin.

Fig. 9.

Example of ERα (ESR1) and ERβ (ESR2) expression in the human fetal penis. Note the more prominent expression of ESR2 see text for details.

Fig. 10.

Morphology of the adult penis and clitoris in several strains of mice as indicated in transverse histologic sections. Clitori of estrogen receptor α knock-out (ESR1-KO; panel C) and estrogen receptor nuclear-only (NOER; panel D) females are substantially masculinized, while clitori of aromatase knock-in (AROM+; panel E) female mice are almost completely masculinized and exhibit morphology similar to that of the wild-type penis, with the exception of the ventral tethering (frenulum, red arrow). B = bone, C = cartilage, Ur = Urethra, Wt = wild-type. Note the preputial space (green arrowheads) in all specimens except the wild-type clitoris (A). From Cunha et al. (Rainey et al., 2004) with permission. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 11.

Mild Mouse Hypospadias. (A) Scanning electron micrograph of the penis of an adult prenatally oil-treated mouse with the position of the os penis (green) superimposed. Note that the os penis is positioned proximal to the ventral cleft in the male urogenital mating protuberance (MUMP) ridge (B) and in the corresponding histological section (C). The region of the MUMP ridge containing the open ventral cleft in the diagrams is colored brown. In mild hypospadias in the mouse, the ventral cleft extends more proximally than in the control penis, and thus the os penis and urethral flaps are seen in sections with an open ventral cleft (compare panels B and C to D and E, respectively). Panels C and E are transverse H&E-stained sections taken where indicated by the vertical lines in panels B and D. Note the mild hypospadias in (E). Adapted from Mahawong et al. (Mahawong et al., 2014a). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 12.

Scanning electron micrographs of a normal adult mouse penis. (A) Distal end-on view. The urethral orifice is Y-shaped, with the ventral cleft forming the ventral stem of the “Y”. The male urogenital mating protuberance (MUMP), which is projecting towards the viewer, is dorsally situated and is fused laterally with the MUMP ridge (colorized light green in panel A). The MUMP ridge is demarcated peripherally by the circumferential MUMP ridge groove (small black arrows in A) and labelled MRG in (B). The MUMP corpora cavernosa (MUMP CC) are also seen (C). The MUMP along with the MUMP ridge define the urethral orifice. (B) Scanning electron micrograph of the ventral adult mouse penis. Note the bifid MUMP, the MUMP ridge groove (MRG, small arrows in panel A), and the position of the urethral meatus. Scale bars = 200 µm for (A) and (B), 100 µm for (C). From Blaschko et al. (2013) with permission (Blaschko et al., 2013b). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 13.

Severe Mouse Hypospadias. Wholemount (A) and scanning electron micrographs (B-F) of adult mouse penises. (A–D) Penises of adult CD-1 mice treated daily with diethylstilbestrol (DES; 200 mg/g of body weight) from the day of birth (P0) to postnatal day 10 (P10) (Mahawong et al., 2014b). (A) Adult neonatally DES-treated penis. (B) Side view of an adult neonatally DES-treated penis. (C) End-on view of an adult neonatally DES-treated penis. (D) Ventral view of an adult neonatally DES-treated penis. (A–D) Note truncated male urogenital mating protuberance (MUMP), multiple prominent processes (*) separated by grooves, the prominent groove separating the MUMP from the MUMP ridge (white arrowheads in B), and the ventral tether. (E & F) Adult AROM + penis in end-on view (E) and lateral view (F). From Cunha et al. (2015) with permission (Cunha et al., 2015).

Fig. 14.

Histology of Normal Adult Male Penis. Sections through the glans penis of three adult untreated mice illustrating the clefts (large arrows in A-C) in the male urogenital mating protuberance (MUMP) ridge. In all cases, the prominent ventral cleft is open to the preputial space. Minor clefts penetrate from the penile surface into the incipient urethral lumen in dorsal-lateral positions adjacent to the MUMP, whose core contains cartilage. In all specimens, minor clefts demarcate individual processes on the epithelial surface, which proximally fuse with adjacent structures (the MUMP or other processes) to complete the MUMP ridge. Note minor clefts (black arrows in B and C), which penetrate from the exterior and end blindly within the interior of the MUMP ridge at the 3 to 4 o’clock positions. (D) is a more proximal section in which clefts shown in (A–C) have fused and have become shallow grooves (*). Scale bar = 250 µm for A, B, and D, and 100 µm for C. Adapted from Blaschko et al. (2013) with permission (Blaschko et al., 2013b).

Fig. 15.

Three-dimensional reconstructions (3DR) (A–D, H, J) of the penis of an adult wild-type mouse treated neonatally (P0–P10) with diethylstilbestrol (DES). 3DRs are presented in a variety of orientations with accompanying tissue sections (E–G, K). The patterns of mesenchymal processes across the epithelial surface (p1–p10 at bottom right) denoted in light and dark blue, red, turquoise, magenta, and green are vastly different from that seen in untreated wild-type mice (compare with Fig. 12). From Blaschko et al. (2013) with permission (Blaschko et al., 2013b).

Fig. 16.

DES Affects Penile Growth. Optical projection tomography images of P5 CD-1 external genitalia derived from male mice injected on days 1 and 3 with oil or diethylstilbestrol (DES) stained with anti-E-cadherin. Note the marked reduction in size of all structures, especially that of the preputial lamina, and the truncation of distal structures destined to form the penile urethral meatus. PPG = preputial gland, PPGD = preputial gland duct. From Mahawong et al. (2014) with permission (Mahawong et al., 2014b).

Fig. 17.

Diethylstilbestrol (DES) Affects Penile Growth. Images from transverse serial section sets of P10 CD1 mouse penises treated with oil or DES (Oil, DES P0–P10, and DES E12–P10) assessed at P10. Sections proceed from distal to proximal (left to right). (A–D represent normal prepubertal penile morphology, in which the penis is defined circumferentially by the external preputial lamina and contains a stand-alone urethra (Ur), os penis (B), and male urogenital mating protuberance (MUMP) cartilage (C). Note the smaller penile diameter (defined by the preputial lamina) of the DES P0–P10 and DES E12–P10 groups, as compared to oil control. The green lines emphasize the differences in diameter between the oil group (C–D) versus the DES P0–P10 (K–L) and DES E12–P10 (O–P) groups. Note that the distance from (A) to (D) in the oil treated spans 91 sections (section thickness is 7 µm, for a total of 637 µm), whereas this distance is reduced to 51 sections (for a total of 357 µm) in the DES P0–P10 group and to 78 sections (total of 546 µm) in the DES PE12–P10 group. The lack of developing MUMP cartilage (I, J, M, N) and impaired development of the MUMP corpus cavernosum (K, L, O, P) is visible in the DES P0–P10 and DES E12–P10 groups. All images are at the same magnification. LC = lateral mesenchymal column, VC = ventral mesenchymal column. Adapted from Sinclair et al. (2016) with permission (Sinclair et al., 2016b). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 18.

Penile wholemounts of adult neonatally oil-treated (A) and P0–P10 diethylstilbestrol (DES)-treated (B) CD-1 mice. Note reduction in length of the glans penis and truncation of the male urogenital mating protuberance (MUMP) in the DES-treated specimen (B). Arrows indicate that penile glans length was determined from depth of the preputial space to the tip of the MUMP.

Fig. 19.

Wholemount photos of adult male external prepuce from prenatally and neonatally oil-and diethylstilbestrol (DES)-treated mice. Prenatal DES treatment (upper row) minimally affects the external prepuce, whereas neonatal DES treatment (lower row) elicits substantial malformation of the external prepuce. Note the fine blue suture in G indicating the urethral meatus. From Mahawong et al. (2014) with permission (Mahawong et al., 2014a, 2014b). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 20.

Effects of Diethylstilbestrol (DES) on the Corpora Cavernosa Urethrae. Transverse histologic sections at the level of the corpora cavernosa urethrae (arrows). In oil-treated mice (A1–A2), the capsules of the corpora cavernosa urethrae are well-demarcated (white and black arrows in A1 and A2). In both DES groups, E12–E18 (B1–B2) and P0–P10 (C1–C2), the corpora cavernosa urethrae are indistinct. Ur = urethra. (D) Corpus cavernosa urethrae index results of the oil and DES-treated groups showing a marked effect of timing of DES treatment. * = p ≤ 0.0001. From Sinclair et al. () with permission (Sinclair et al., 2016b).

Fig. 21.

Scanning electron micrographs of adult (P60) mouse penises treated with oil vehicle or diethylstilbestrol (DES) at the ages specified. (A–F) end-on views, (G–L) lateral views, and (M–R) ventral views. Severe truncation of the male urogenital mating protuberance (MUMP) (green arrowheads in I and J), malformation of the urethral meatus with abnormal clefting patterns in the MUMP ridge (C and D), and a ventral tether (purple arrowheads in I, J, O, and P) are observed in the P0–P10 and E12–P10 groups. Other treatment groups show less severe malformations or minimal departure from the oil-treated specimens. IP = internal prepuce, MR = MUMP ridge, MRG = MUMP ridge groove, VC = ventral cleft in the MUMP ridge. (S) Morphometric analysis of MUMP length in perinatally DES-treated mice. Significance symbols: ✚ = p ≤ 0.05, * = p ≤ 0.0001. (A–S) from Sinclair et al. (2016) with permission (Sinclair et al., 2016b). (T) Bone length of the adult mouse penis of neonatally oil- or DES-treated CD-1 and C57BL/6 (C57) male mice assessed at age P60, with DES treatment from P0–P10. Os penis length was significantly reduced by DES treatment in both CD-1 and C57BL/6 mice (p < 0.001 for both). From Mahawong et al. (2014) with permission (Mahawong et al., 2014b). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 22.

Wholemount images of normal female mouse genital tubercles (GTs) at E14, E16, E18, and at P0 (1d) as ventral views. The preputial swellings enlarge and fuse in the midline to form the prepuce, which extends distally to eventually completely cover the GT. Arrowheads outline the distal tip of the P0 GT, which is almost completely covered by the preputial swelling (PS). At E18 and P0, the preputial-urethral groove (PUG) can be seen where the preputial swelling are fusing in the midline (large black arrows). From Cunha et al. (2020) with permission (Cunha et al., 2020a).

Fig. 23.

Diagrammatic representations of development of the prepuce and the preputial lamina (red) from the preputial swellings. Coronal histologic sections depict the process at E14, E16, and P0. Note that as the prepuce grows distally, the preputial lamina is “left in its wake” and therefore grows in length. Black arrows indicate distal growth of the prepuce and dotted lines represent the urethra. At birth, the central core of mesenchyme (green in top row), which is the precursor of the male urogenital mating protuberance (MUMP), penetrates the complex central epithelium. GT = genital tubercle, PPG = preputial gland, PP Lam = preputial lamina, PS = preputial swelling. From Liu et al. (2018) with permission (Liu et al., 2018b). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 24.

Transverse sections of the external genitalia of a normal newborn male CD-1 mouse. Sections are arranged from distal (A) to proximal (I). Distally (A), the right and left preputial swellings are approaching the ventral midline, forming the preputial-urethral groove (PUG). Note the growth and incipient fusion in the midline (large black arrows in C). The prepuce (double-headed red arrows in A, D, and G) consists of a thick wall of loose mesenchyme (PPM). The penis is surrounded by the preputial lamina; A–I) and consists of dense penile stroma. The PUG fuses ventrally (D and F) to complete the prepuce and form the preputial-urethral canal (PUC). A secondary fusion (E) forms a transitory seam, which disappears (F and G) to segregate the urethra and establish the midline mesenchymal confluence. Note that as the urethra is separating from the preputial lamina, a ventral gap in the preputial lamina is observed. Adapted from Sinclair (2016) with permission (Sinclair et al., 2016c). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 25.

Canalization of the external preputial lamina and formation of the penile surface. Transverse sections of the developing CD-1 mouse penis at the postnatal days specified. PS = preputial space, Ur = urethra, MC = MUMP cartilage. From P0–P25, the preputial lamina remains solid. At P30, the preputial lamina is fully canalized, creating the preputial space that houses the penis. From Mahawong et al. (2014) with permission (Mahawong et al., 2014b).

Fig. 26.

Ventral tethering in the mouse penis. Note the continuous stromal channel (large red arrows) linking the connective tissue of the preputial mucosa with the penile stroma. Likewise, note the continuity of epithelium of the preputial mucosa and the penile surface epithelium. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 27.

Immunohistochemistry of developing penises of P10 mice stained for estrogen receptor alpha (ESR1) and estrogen receptor beta (ESR2) as indicated. The male urogenital mating protuberance (MUMP) cartilage (arrows in A–B) expresses both receptors. (C–D) are higher magnifications of the MUMP cartilage in (A–B). Scale bar for (A) and (B) is in the bottom right of (B) and scale bar for (C) and (D) is in the bottom center of (C). From Blaschko et al. (2013) with permission (Blaschko et al., 2013b).

Fig. 28.

Immunohistochemistry of developing penises of P10 mice stained for estrogen receptor alpha (ESR1) and estrogen receptor beta (ESR2) as indicated. Abbreviations: EPL = External preputial lamina, IPL= Internal preputial lamina, MUMPCC = MUMP corpus cavernosum, CCUr = corpus cavernosum urethrae, CCG = corpus cavernosum glandis, Cart = MUMP cartilage. Scale bar for (A) and (C) is in the bottom right of (C) and scale bar for (B) and (D) is in the bottom center of (B). From Blaschko et al. (2013) with permission (Blaschko et al., 2013b).

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Estrogens and development of the mouse and human external genitalia - PubMed (original) (raw)