Localization,metabolism, and binding of estrogens in the male rat

Estrogen assimilation by male Wistar rats was examined in these studies in several accessory sex organs (seminal vesicles and anterior, dorsal, lateral, and ventral prostates) as well as in a variety of nonaccessory sex organs. When [³H]estradiol was injected into intact 3- to 4-month-old rats in a pulse dose, no selective accumulation of radioactivity recovered as estradiol was found in the accessory sex glands when compared to other organs. This was due at least in part to the metabolism of estradiol to estrone and to the relatively low concentration of high affinity estrophilic molecules in the accessory sex organs. The order for the rate of formation of estrone from estradiol in tissues obtained from intact animals was ventral prostate > lateral and dorsal prostate > anterior prostate and seminal vesicles. Steroid specificity studies for cytosol estradiol binding by the ventral prostate and seminal vesicles revealed that estrophilic molecules exist in these organs. Based on Scatchard plot analyses in 24-h castrates, the number of available estradiol binding sites was too low in the ventral prostate to quantify accurately, but the seminal vesicles contained distinctly more estrophilic activity than the ventral prostate. The affinity for the seminal vesicle cytosol estradiol-estrophile binding exceeded that quantified for the seminal vesicle dihydrotestosterone-androphile reaction while the number of estradiol binding sites was less than that quantified for dihydrotestosterone. In relation to the accessory sex organs of other species, the rat seminal vesicles have a relatively small amount of cytosol estrophile. The findings that the seminal vesicles catabolize less estradiol and contain significantly more estrophilic activity than the ventral prostate is consistent with and offers insight into the noted estrogenic sensitivity of the seminal vesicles and lack thereof in the rat ventral prostate. With aging of the rat from 3–4 months to 22–26 months, the affinity of the seminal vesicle estradiol-estrophile interaction was unchanged but the number of binding sites increased significantly.     

Antigenic homogeneity of male Müllerian gland (MG) secretory proteins of a caecilian amphibian with secretory proteins of the mammalian prostate gland and seminal vesicles: evidence for role of the caecilian MG as a male accessory reproductive gland

Whereas in all other vertebrates the Müllerian ducts of genetic males are aborted during development, under the influence of Müllerian-inhibiting substance, in the caecilian amphibians they are retained as a pair of functional glands. It has long been speculated that the Müllerian gland might be the male accessory reproductive gland but there has been no direct evidence to this effect. The present study was undertaken to determine whether the caecilian Müllerian gland secretory proteins would bear antigenic similarity to secretory proteins of the prostate gland and/or the seminal vesicles of a mammal. The secretory proteins of the Müllerian gland of Ichthyophis tricolor were evaluated for cross-reactivity with antisera raised against rat ventral prostate and seminal vesicle secretory proteins, adopting SDS-PAGE, two-dimensional electrophoresis and immunoblot techniques. Indeed there was a cross-reaction of five Müllerian gland secretory protein fractions with prostatic protein antiserum and of three with seminal vesicle protein antiserum. A potential homology exists because in mammals the middle group of the prostate primordia is derived from a diverticulum of the Müllerian duct. Thus this study, by providing evidence for expression of prostatic and seminal vesicle proteins in the Müllerian gland, substantiates the point that in caecilians the Müllerian glands are the male accessory reproductive glands.     

Localization and characterization of glutathione peroxidase (GPx) in boar accessory sex glands, seminal plasma, and spermatozoa and activity of GPx in boar semen

Boar ejaculate owes its characteristic large volume mainly to accessory sex gland (ASG) secretions. These are main contributors to the protective functions of seminal plasma, especially against oxidative damage. Numerous antioxidants have been detected in ASG secretions, and, respectively, in seminal plasma. However, as regards one key antioxidant protector -- the Se-dependent enzyme glutathione peroxidase (GPx) -- there is no agreement yet among researchers as to its presence in boar seminal plasma. Nevertheless, the beneficial effect of dietary Se supplementation on male fertility has been widely recognized. The aim of the present study was to investigate the localization and characterization of GPx in boar ASGs, seminal plasma, and spermatozoa, as well as to evaluate GPx activity in boar semen. Immunohistochemical assays demonstrated GPx presence in the epithelial cells, vacuole membranes, and vascular endothelium of boar seminal vesicle, prostate and bulbourethral glands. Western blot analysis demonstrated the presence of a monomer form of GPx with MW 20 kDa in lysates from seminal vesicle, prostate, bulbourethral glands, and spermatozoa, but not in seminal plasma. Surprisingly, peroxidase activity detected in seminal plasma from normal ejaculates was nearly three times as high as in spermatozoa. Our findings confirmed the presence of immunoreactive GPx in the boar reproductive tract, while further investigation is still warranted to uncover the exact protein forms involved and their function.     

Influence of neonatal diethylstilbestrol treatment on androgen and estrogen receptor levels in the mouse anterior prostate, ventral prostate and seminal vesicle

Perinatal exposure to the synthetic estrogen, diethylstilbestrol (DES), affects the structure of both male and female reproductive systems. Changes may also occur in the levels of steroid hormone receptors. Cytosolic and nuclear androgen and estrogen receptor levels (expressed per mg DNA) from the sex accessory glands of male BALB/c mice exposed neonatally to DES were analyzed by exchange assays. Neonatal DES exposure caused significant decreases in: (1) cytosolic androgen and cytosolic and nuclear estrogen receptor levels in the anterior prostate and (2) cytosolic estrogen receptor levels in the ventral prostate. A significant increase was seen in the cytosolic estrogen receptor levels in the seminal vesicle. Significant decreases in cytosolic protein levels occurred in all DES-exposed glands.     

Sonomorphology of the reproductive tract in male and pregnant and non-pregnant female Rothschild's giraffes (Giraffa camelopardalis rotschildi)

The application of real-time-B-mode ultrasonography to wild and zoo animal medicine has been shown to improve the understanding of reproductive physiology in many species. Ultrasound technology is especially helpful for monitoring urogenital health, which in turn has advantages for giraffe breeding and welfare in captivity.This study aimed to ultrasonographically describe the genital organs of reproductively healthy male and female giraffes. Through the use of a restrainer, repeated rectal ultrasound examinations were performed over a 2 year period in 2.6 Rothschild's giraffes. Changes in ovarian activity were monitored throughout four different reproductive stages in the females and included immature, mature-cycling, pregnancy, post-partum-period. In the immature giraffes the ovaries showed multiple follicles of which larger ones luteinized to form pseudo-corpora lutea. By comparison, in the mature giraffes the dominant follicle reached an ovulatory diameter of 18.5 ± 0.89 mm. After ovulation, a single corpus luteum rapidly formed and reached a maximum diameter of 33.0 ± 2.4 mm on average. Pregnancy was detected for the first time by the embryonic vesicle, visualized around 28 days post copulation. Follicular development remained ongoing during early pregnancy. In the males, as in other ruminants, the bulbourethral glands and the seminal vesicles were prominent, whereas the prostate gland was indistinct.Knowledge about the reproductive tract morphology and physiology is necessary for diagnosing medical disorders and abnormalities in giraffes. The aim of this study was to help consolidate the current knowledge on basic reproductive parameters for this species.     

The influence of aging on androgen dynamics in the male rat

Androgen assimilation was investigated in a variety of accessory sex organs (seminal vesicles and anterior, dorsal, lateral, and ventral prostates) and in several nonaccessory sex organs in male Wistar rats. After administration of a pulse dose of [³H]testosterone in vivo to intact young (3–4 months old) rats, [³H]testosterone was the primary radioactive steroid recovered from most organs examined, except for the secondary sex glands where the reduced metabolites, [³H]5α-dihydrotestosterone (DHT) and [³H]5α-androstanediol(s), predominated. At longer postinjection times, [³H]DHT was preferentially retained in the accessory sex glands, presumably reflecting intracellular metabolism of [³H]testosterone to this compound and subsequent specific binding of [³H]DHT to receptor proteins. At the longest postinjection interval investigated, the ventral prostate retained greater concentrations of [³H]DHT than the lateral prostate which in turn had a higher [³H]DHT concentration than the seminal vesicles or anterior or dorsal prostates. The latter three glands retained approximately equal concentrations of [³H]DHT. Scatchard plot analyses of cytosol binding in 24-h castrates indicated that with one exception, the level of high affinity DHT binding sites was generally correlated with the retention of [³H]DHT in vivo in intact rats. Specifically, while the affinity for DHT binding in all accessory sex organs was the same, the number of high affinity binding sites per mg wet tissue weight was on the order of ventral prostate > anterior prostate ≥ seminal vesicles ≥ dorsal prostate > lateral prostate. Studies of the influence of aging to 22–26 months revealed no apparent differences in the affinity of the DHT receptor for its ligand in any of the accessory sex glands from 24-h castrates when the receptors were present in levels sufficiently high to quantify. The concentration of available DHT receptors with advancing age remained constant in the anterior and dorsal prostates, increased in the seminal vesicles, and declined in the ventral and lateral prostates. The decreases observed in the ventral prostate were only partial, but the receptors of the lateral prostate declined to nondetectable levels.     

Basal cells of H-Dunning tumor are myoepithelial cells. A comparative immunohistochemical and ultrastructural study with male accessory sex glands and mammary gland

Recent immunohistochemical studies have shown that basal cells in human prostatic epithelium are not myoepithelial cells. Since in the literature the Dunning tumor, originally described as a rat prostate carcinoma derived from the dorsolateral prostate of a Copenhagen rat, was reported to have myoepithelial cells, a comparative immunohistochemical and ultrastructural study was performed in the H-, HIF- and AT3-lines of the Dunning tumor, the male accessory sex glands (ventral, dorsal, lateral prostate, coagulating gland, bulbourethral gland) and the mammary gland of both Copenhagen and Wistar rats. Mono- and polyclonal antibodies directed against intermediate filament proteins (cytokeratin, desmin, vimentin) and the contractile proteins (alpha-actin, muscle type specific myosin, tropomyosin) were used along with phalloidin decoration of F-actin. As in the human prostate, none of the rat prostate lobes in either strains did contain basal cells expressing cytokeratin along with alpha-actin, myosin and tropomyosin Cells representing fully differentiated myoepithelial cells, however, were present as anticipated in the mammary gland, the bulbourethral gland and the H-tumor line of the Dunning tumor. This finding is difficult to reconcile with the contention of a prostatic origin of the H-Dunning tumor. Further studies are required to classify the epithelial parental tissue in order to define the true origin of the H-Dunning tumor and the tumor lines derived thereof.     

Basal cells of H-Dunning tumor are myoepithelial cells

Summary Recent immunohistochemical studies have shown that basal cells in human prostatic epithelium are not myoepithelial cells. Since in the literature the Dunning tumor, originally described as a rat prostate carcinoma derived from the dorsolateral prostate of a Copenhagen rat, was reported to have myoepithelial cells, a comparative immunohistochemical and ultrastructural study was performed in the H-, HIF- and AT₃-lines of the Dunning tumor, the male accessory sex glands (ventral, dorsal, lateral prostate, coagulating gland, bulbourethral gland) and the mammary gland of both Copenhagen and Wistar rats. Mono- and polyclonal antibodies directed against intermediate filament proteins (cytokeratin, desmin, vimentin) and the contractile proteins (-actin, muscle type specific myosin, tropomyosin) were used along with phalloidin decoration of F-actin. As in the human prostate, none of the rat prostate lobes in either strain did contain basal cells expressing cytokeratin along with -actin, myosin and tropomyosin. Cells representing fully differentiated myoepithelial cells, however, were present as anticipated in the mammary gland, the bulbourethral gland and the H-tumor line of the Dunning tumor. This finding is difficult to reconcile with the contention of a prostatic origin of the H-Dunning tumor. Further studies are required to classify the epithelial parental tissue in order to define the true origin of the H-Dunning tumor and the tumor lines derived thereof.     

Anatomic,Histologic and Certain Enzymatic Studies on the Male Genital Organs of Hemiechinus auritus collaris Gray,the Indian Long Eared Hedgehog

Anatomically the male reproductive organs of Hemiechinus auritus collaris are of considerable interest and present a unique pattern of arrangement of different glands. The testes are inguinal and a true scrotum is absent. A pair of accessory glands, the seminal vesicles, are situated dorsal to the bladder. A pair of obviously lobulated glands, ventral to the bladder, represent the internal prostate and a pair of compact glands situated outside the pelvis in para-anal position the external prostate. The Cowper's gland and the gland of ampulla are absent. The present studies also concern enzymes of the phosphatase group, including both specific and non-specific phosphatases, in the testes and the sex accessory glands during both active and inactive periods in this insectivore.     

Expression of transforming growth factor-β isoforms in the rat male accessory sex organs and epididymis

We studied the expression and distribution of transforming growth factor-β (TGF-β) isoforms in the rat male accessory sex glands and the epididymis. Our data demonstrate the expression of both TGF-β1 and -β3 isoforms in ventral prostate (VP), seminal vesicle (SV), coagulating gland (CG), and epididymis (E) by Northern blot analysis. In addition, there was differential expression of TGF-β3 in the three regions of epididymis, the corpus region being the highest. Immunostaining data showed intense staining for latent TGF-β1 in all the male accessory glands. In contrast, no staining using antibodies specific for active TGF-β1 was observed. No expression of TGF-β2 was evident either by immunohistochemistry or Northern blot analysis. The presence of mature TGF-β3 protein was observed in the secretory epithelium of VP, CG, and corpus E. There was no detectable staining of TGF-β3 in the seminal vesicle and caput and cauda regions of epididymis. These data suggest possible differential regulation of TGF-β isoform expression in the male reproductive system and predict unique roles for individual TGF-β isoforms in sperm maturation and maintenance.     

Lactate dehydrogenase C4 in male sex accessory glands of normal mice and in testes of sex-reversed mice.

Lactate dehydrogenase (LDH) C4 activity was observed in testis extracts of sex-reversed mice (Sxr) and in male sex accessory gland (seminal vesicle and prostate) extracts from C3H/He and C57BL/Go mice. These results reflect either (1) the presence of low concentrations of germinal cells in these tissues; or (2) the synthesis of LDH-C4 by somatic cells in Sxr testes and normal male sex accessory glands.     

A secretory protease inhibitor requires androgens for its expression in male sex accessory tissues but is expressed constitutively in pancreas.

A full length cDNA clone encoding a mouse prostatic secretory glycoprotein (p12) whose synthesis is dependent upon testicular androgens has been cloned and characterized. The predicted amino acid sequence of p12 shares extensive homology with several members of the Kazal family of secretory protease inhibitors, in particular the pancreatic secretory trypsin inhibitors. In agreement with sequence data, prostatic secretory p12, purified from mouse ventral prostate secretion, exhibits anti-trypsin activity. Steady-state levels of protease inhibitor mRNA in ventral prostate are reduced from approximately 0.06% in normal mice to undetectable after androgen withdrawal but are inducible within 4 h by re-administration of testosterone. Androgen-dependent expression of the secretory protease inhibitor mRNA was also observed in coagulating gland and seminal vesicle. In seminal vesicle, a tissue of different embryonic origin to the prostate, the kinetics of secretory protease inhibitor mRNA loss after castration are not as rapid as in the ventral prostate and coagulating gland. Low-level androgen independent expression was also observed in the pancreas. There appears to be a single gene for this secretory protease inhibitor and yet expression is markedly stimulated by testosterone in the sex accessory tissues and unaffected by this hormone in the pancreas.     

Effects of male accessory sex gland secretions on early embryonic development in the golden hamster

The ventral prostates, dorsolateral prostates, coagulating glands, seminal vesicles and/or ampullary glands were bilaterally removed from adult male hamsters. Removal of these glands did not affect the fertilization rate and cleavage of the embryos at 48 h post coitum (p.c.). Air-dried preparations of the embryos showed a delay in cleavage at 72 h p.c. and a significant number of degenerated embryos was also found in females mated with males from which all the male accessory sex glands had been removed. A significant implantation loss was also observed at 122 h p.c. The results suggest that, in the golden hamster, removal of the male accessory sex gland causes a slower cleavage rate in embryonic development and a significant embryonic loss during pregnancy.     

Morphological analysis of the male reproductive accessory glands of the bat Artibeus lituratus (Phyllostomidae: Chiroptera)

Bats are distributed worldwide from tropical to temperate regions. Despite their wide geographical radiation and advances in studies using evolutionary approaches, aspects related to the reproduction of these animals remain poorly explored, especially those related to the male reproductive accessory glands (RAGs). Thus, the aim of this study was to analyze the morphophysiology of the male RAGs in the bat Artibeus lituratus. The RAGs in A. lituratus are composed of a compact intra‐abdominal glandular complex, consisting of the prostate with two prostatic regions (ventral and dorsal), plus Littre glands and a pair of extra‐abdominal bulbourethral glands. The ventral region of the prostate has an epithelium with variable morphology, due to its holocrine type of secretion. In contrast, the dorsal region has a typical cubic‐to‐columnar pseudostratified epithelium. Both regions contain two cell types, basal and secretory cells. Similar to the epithelial morphology, the secretion also varies, with the ventral region containing numerous PAS‐positive globular vesicles, whereas the dorsal region has a more fluid, hyaline and PAS‐negative secretion. Littre glands are dispersed in the connective tissue of the urethra, while the bulbourethral glands are located in the penile root, both glands with cubic‐to‐columnar pseudostratified epithelium and globular PAS‐positive secretion. The results demonstrate that the RAGs of A. lituratus are composed of two prostatic regions, ventral and dorsal, and urethral and bulbourethral glands, with no seminal vesicles. Each prostatic region has unique and distinctive characteristics, with the ventral region presenting an exclusive holocrine nature and the dorsal region having similarities to the ventral prostate of rodents.     

Effects of CdCl2 on the secretory functions of sex accessory glands of rats

The effects of cadmium chloride (Cd) alone (1 mg given as a single injection) or in combination with ascorbic acid (AA; 100 mg/day for 10 days) on the secretory functions of sex accessory glands of rats were studied in healthy male albino rats. Animals were sacrificed after 10 days treatment and the seminal vesicles (SVs), dorsolateral prostate (DLP), ventral prostate (VP), bulbourethral glands (BU), and coagulating glands (CO) were excised and weighed. Weight of all accessory glands were decreased by 10 days treatment with Cd. Cd + AA gave similar results. AA concentration increased in all glands and was significantly increases in CO (p less than .01). Levels of ascorbogin increased in all glands except CO and BU and in the latter a significant (p less than .001) increase was found. The rate of AA utilization increased significantly (p less that .001) in the accessories. A significant (p less than .001) reduction in the activity of succinate dehydrogenase was observed in Cd-treated rats with a further reduction with combined treatment. Alkaline phosphatase decreased (p less than .001) after Cd treatment but AA in combination restored it to control levels. Cd increased acid phosphatase (p less than .001) and was further activated by Cd + AA. Phosphorylase activity was elevated with Cd (p less than .001) but recovery occurred in SV and BU with Cd + AA. Glycogen increased (p less than .001; .01) with both treatments as did citric acid. Protein results were inconsistent with Cd but activation was found in most glands under combined treatment. The results reveal that most androgen-dependent biochemical constituents and organ weights were affected significantly by a single injection of Cd. AA had a protective and beneficial influence on the restoration of structural integrity and metabolism in sex accessory glands.     

Immunocytochemical localization of seminal proteins in salivary and lacrimal glands of the rat

Antibodies against 10 different secretory proteins from the accessory sex glands of the male rat were used for immunohistochemical studies of salivary and lacrimal glands from intact and castrated rats, at the light- and electron-microscopic levels. In the parotid gland, secretory acinar cells showed immunoreactivity with antibodies against prostatic binding protein, cystatin-related peptide and acid phosphatase (isoenzyme pI 8.0; 5.6) typical of ventral prostate, and seminal vesicle secretion VI. Western blotting analysis indicated that immunoreactivity against prostatic binding protein was attributable to a subunit, presumably C3. Acid phosphatase pI 5.6 showed a molecular weight of 66 kDa, which is at variance with the prostatic form. Immunoreactivity for secretory transglutaminase, derived from the coagulating gland, was restricted to myoepithelial and stromal cells. In castrated animals, the immunoreactivity of acinar cells was reduced to the background level, whereas stromal transglutaminase immunoreactivity was unaltered. The distribution pattern of immunoreactivity for the proteins mentioned was almost identical in the lacrimal gland. Significant differences were however observed in the immunoreactivity of the inframandibular gland, where serous glandular cells were non-immunoreactive for seminal proteins, with the exception of acid phosphatase isoenzyme pI 8.0. Granules present in the convoluted granular ducts were immunoreactive particularly for acid phosphatase (isoenzyme pI 5.6)but much less for cystatin-related peptide; immunoreactivity was reduced after castration. The straight portion of the inframandibular duct system was immunoreactive for transglutaminase, but no influence of castration was visible. The distribution of immunoreactivity for seminal proteins present in the salivary and lacrimal glands and the pronounced androgen-dependence of their expression point to functional relationships of the respective proteins at both glandular sites.     

Comparative analysis of the male reproductive accessory glands of bats Noctilio albiventris (Noctilionidae) and Rhynchonycteris naso (Emballonuridae)

In eutherian mammals, the male reproductive accessory glands (RAGs) comprise the prostate, bulbourethral glands, ampullary glands, and the seminal vesicles. Their composition, anatomy and function vary widely between species. This study aimed to characterize histologically and compare the RAGs of bats. The RAGs of Noctilio albiventris (Noctilionidae) and Rhynchonycteris naso (Emballonuridae) were studied using anatomical and histological methods, and were reconstructed three dimensionally. The RAGs of N. albiventris and R. naso are composed of a compact glandular complex that surrounds the urethra and a pair of bulbourethral glands, which are extra‐abdominally located in the inguinal region. In both species, the glandular complex is composed of two well‐defined prostatic regions (ventral and dorsal). The ventral region showed an atypical epithelium (holocrine), where no obvious cellular limits were observed, and PAS‐positive secretion. The dorsal region had a pseudostratified cuboidal epithelium, with basal and secretory cells, and PAS‐negative secretion. Noctilio albiventris also had urethral glands (Littre glands) surrounding the urethra, however, R. naso had only muscles. Both species had bulbourethral glands, with simple columnar epithelium and PAS‐positive secretion. In conclusion, the RAGs of N. albiventris and R. naso comprised a pair of bulbourethral glands and an intra‐abdominal complex, composed of a prostate with two different regions (ventral and dorsal), while the ampullary glands and seminal vesicles were missing in both species. This morphology was more closely related between N. albiventris and R. naso, and to species of the family Phyllostomidae than to families Molossidae and Vespertilionidae. J. Morphol. 277:1459–1468, 2016. © 2016 Wiley Periodicals, Inc.     

Selenoprotein P in seminal fluid is a novel biomarker of sperm quality

Hepatically-derived selenoprotein P (SePP) transports selenium (Se) via blood to other tissues including the testes. Male Sepp-knockout mice are infertile. SePP-mediated Se transport to Sertoli cells is needed for supporting biosynthesis of the selenoenzyme glutathione peroxidase-4 (GPX4) in spermatozoa. GPX4 becomes a structural component of sperm midpiece during sperm maturation, and its expression correlates to semen quality. We tested whether SePP is also present in seminal plasma, potentially correlating to fertility parameters. Semen quality was assessed by sperm density, morphology and motility. SePP was measured by an immunoluminometric assay, and trace elements were determined by X-ray fluorescence spectroscopy. SePP levels were considerably lower in seminal plasma as compared to serum (0.4 ± 0.1 mg/l vs. 3.5 ± 1.0 mg/l); Se concentrations showed a similar but less pronounced difference (48.9 ± 20.7 μg/l vs. 106.7 ± 17.3 μg/l). Se and Zn correlated positively in seminal fluid but not in serum. Seminal plasma SePP concentrations were independent of serum SePP concentrations, but correlated positively to sperm density and fraction of vital sperm. SePP concentrations in seminal plasma of vasectomized men were similar to controls indicating that accessory sex glands are a testes-independent source of SePP. This notion was corroborated by histochemical analyses localizing SePP in epithelial cells of seminal vesicles. We conclude that SePP is not only involved in Se transport to testes supporting GPX4 biosynthesis but it also becomes secreted into seminal plasma, likely important to protect sperm during storage, genital tract passage and final journey.