Structural, ultrastructural and immunohistochemical evidence of testosterone effects and its ablation on the bulbourethal gland of the Artibeus planirostris bat (Chiroptera, Mammalia).

This study evaluated the effects of testosterone in the bulbourethral glands (BG) of the bat, Artibeus planirostris, by performing castration and posterior hormonal supplementation of the animals. The results showed a decrease in testosterone levels in animals 15days after castration, which induced a small reduction in epithelium height, percentage of AR+ cells, and an increase in the amount of basal cells. This reduction became more severe in groups castrated for longer periods (19 and 22days), where there was also an increase in apoptotic cells. Moreover, the hormonal supplementation increased testosterone levels (after 3 and 7days of supplementation), causing a glandular reactivation that increased the epithelium height and AR expression. In conclusion, BG took longer to respond to ablation of testosterone than other reproductive glands, since it showed evident aspects of regression only in animals 22days after castrated.

Ultrastructure of the secretory epithelial cells of the bulbo-urethral glands (Cowper's gland) in the Indian fruit bat, Rousettus leschenaulti (Desmarest) during the reproductive cycle / Ultraestructura de las células epiteliales secretoras de las glándulas bulbouretrales (glándula de Cowper) en el murciélago de la India, Rousettus leschenaulti (Desmarest) durante el ciclo reproductivo

The present paper describes the ultrastructural characteristics of the bulbo-urethral gland (Cowper' glands) of the Indian fruit bat, R. leschenaulti during sexually inactive-breeding cycle. Cyclic histological changes during the seasonal breeding quiescence cycle are not well marked. There are no marked differences. The ultrastructural characteristic of the secretory epithelial cells of Bulbo-Uretrhal Gland gland have been studied during different phases of reproductive cycle. The secretory epithelial cells are characterized by the well-developed rough endoplasmic reticulum, extensive developed complexus golgiensis (Golgi apparatus) and mitochondria. Mitochondria with lamellate cristae are dispersed in the cytoplasm. Three different types of secretory granules can be identified on the basis of electron density. These granules are not of different types but they represent the different stages of granule maturation. The secretory products of bulbo-urethral gland of bat are released into lumen both by apocrine and merocrine modes. The functional significance of the secretions of the bulbo-urethral glands in reproduction is discussed.

El presente trabajo describe las características ultraestructurales de las glándulas bulbouretrales (glándulas de Cowper) del murciélago de la fruta de la India, R. leschenaulti durante el ciclo inactivo de reproducción sexual. Los cambios histológicos cíclicos durante el ciclo de quiescencia estacional de la cría no están bien determinados. No hay diferencias marcadas. La característica ultra estructural de las células epiteliales secretoras de la glándula bulbouretral ha sido estudiada durante diferentes fases del ciclo reproductivo. Las células epiteliales secretoras se caracterizan por un retículo endoplasmático rugoso bien desarrollado, el complexus golgiensis (complejo de Golgi) y mitocondrias desarrollados extensamente. Las mitocondrias con crestas lamelares se encontraron dispersas en el citoplasma. Se pueden identificar tres tipos diferentes de gránulos secretores en base a la densidad de electrones. Estos gránulos no son de tipos diferentes, sino que representan las diferentes etapas de maduración del gránulo. Los productos secretores de las glándulas bulbouretrales de murciélagos son liberados en el lumen tanto por modos apócrinos como merócrinos. Se discute la importancia funcional de las secreciones de la glándula bulbouretral en la reproducción.

Semiquantitative immunohistochemical detection of progesterone receptors in male accessory sex glands as a screening assay for anabolic steroid use in bulls.

We investigated the immunohistochemical expression of progesterone receptors (PRs) in the prostate and bulbourethral glands of thirty-two 10-14-mo-old Charolais bulls following treatment with a low dosage of estrogens. Animals were divided into 2 groups: 16 animals (group T) were treated for 71 d with a therapeutic dose of trenbolone acetate and estradiol by subcutaneous implant, 16 animals (group C) received no treatment. Urine samples were collected both at the beginning of the trial and 9 times during the study. A semiquantitative analysis of immunohistochemistry (IHC) was performed by counting the number of positive cells in 10 randomly selected high-power fields (hpf). Both groups showed no significant histologic lesions. IHC examination showed positive cells in the epithelium of both glands, with different patterns of distribution between groups. In group C, IHC-positive cells per hpf varied from 0 to 40 in the prostate and from 0 to 32 in the bulbourethral gland. In group T, positive cells varied from 0 to 85 per hpf in the prostate and from 0 to 75 in the bulbourethral gland. The treated group showed significantly higher median numbers of positively stained cells in both organs than the controls ( p < 0.001). Chemical analysis of the urine samples confirmed that the experimental treatment mimics continuous, low-dose administration of anabolic steroids. IHC quantification showed good sensitivity with a high predictive power to correctly classify treated animals and could be used as a preliminary screening test in bulls.

[The history of discovery of bulbourethral glands].

The article outlines the chronology of the discovery of the bulbourethral glands. Their first image appeared in 1600 in a collection of anatomical tables by Italian anatomist Hieronymus Fabricius of Acquapendente. In the scientific literature, they were first mentioned in one of the editions of "Journal des scavans" in 1684 as discovered by the French surgeon Jean Mry. The first detailed description of the structure, topography and function of these glands was presented in the work of English physician and anatomist William Cowper in 1699.

Regucalcin Expression as a Diagnostic Tool for the Illicit Use of Steroids in Veal Calves.

It has been previously demonstrated that sex steroid hormone treatment down-regulates regucalcin gene expression in the accessory sex glands and testis of prepubertal and adult male bovines. The aim of this study was to investigate whether low doses of sex steroid hormones combined with other drugs significantly affect regucalcin gene expression in the accessory sex glands and testis of veal calves. The regucalcin expression was down-regulated in the bulbo-urethral glands of estrogen-treated calves, whereas it was up-regulated in the prostate of estrogen-treated calves. Only the testis of androgen-treated calves showed a down-regulation of the regucalcin expression. Thus, the administration of sex steroid hormones, even in low doses and combined with other molecules, could affect regucalcin expression in target organs. Particularly, the specific response in the testis suggests regucalcin expression in this organ as a first molecular biomarker of illicit androgen administration in bovine husbandry.

Structure, histochemistry and seasonal variations of the male reproductive accessory glands in the Pallas's mastiff bat, Molossus molossus (Chiroptera: Molossidae).

Due to their wide geographical distribution, bats suffer considerable influence from abiotic factors on their reproductive strategies, detected through behavioural or functional assessment of the gonads and accessory glands. The present study aimed to characterise anatomically and morphologically the reproductive accessory glands (RAGs) of Molossus molossus (Molossidae) and evaluate their seasonal variations. The RAGs were removed, fixed, sectioned after histological processing and submitted to the following stains: haematoxylin-eosin, periodic acid--Schiff (PAS) and Gömöri's reticulin. Our data demonstrated that the RAGs of M. molossus are composed of a semi-lobed prostatic complex associated with the urethra and a pair of inguinal bulbourethral glands. Histology and three-dimensional reconstruction of the prostatic complex demonstrated the existence of two regions: ventral and dorsal (named according to the position around the urethra). The two regions had distinctive morphological and histological differences, with the ventral region being the most prominent and the dorsal formed by four lobes. Considering the seasonal evaluation, we can infer that the prostatic complex of M. molossus is active throughout the seasons in São Paulo State (Brazil) and, although each of the prostatic regions has inherent characteristics, they synchronise to establish the main reproductive peak in spring (early rainy season).

Effect of long-term testosterone propionate or human chorionic gonadotrophin administration on reproductive glands in adult male rabbits.

The study was aimed to investigate the effect of testosterone propionate (TP) or human chorionic gonadotrophin (hCG) treatment on reproductive glands in sexually mature male rabbits. A total 36 adult male rabbits were randomly distributed to six equal groups. The first control group (CON), the second treated with low-dose TP (TPL), the third treated with high-dose TP (TPH), the fourth treated with low-dose hCG (CGL), the fifth treated with medium-dose hCG (CGM) and sixth treated with high-dose hCG (CGH). At the 16th post-treatment week, the animals were sacrificed, and the testes and accessory sex glands dissected, weighted and stored at -20 °C until assay. Testosterone propionate treatment in both doses resulted in reduction (P < 0.01) in testicular weight and increase (P < 0.01) in weight of vesicular gland, paraprostate and proprostate glands. High-dose TP increased the weight of prostate and bulbouretheral gland (BUG). Testosterone propionate increased total androgen (P < 0.01) with Testosterone (T) predominating in serum, dihydrotestosterone (DHT) predominating in testes and most accessory sex glands. High dose of hCG increased the weight of proprostate and paraprostate glands. Androgen level in serum, testes and accessory sex glands increased (P < 0.01) after hCG treatment.

Structure, histochemistry and ultrastructure of the male reproductive accessory glands in the neotropical flat-faced fruit-eating bat Artibeus planirostris (Chiroptera: Phyllostomidae).

Chiroptera, the second largest mammalian order, presents different reproductive strategies and unique reproductive features. However, there are few reports regarding male reproductive accessory glands (RAGs) in Chiroptera. Thus, the aim of the present study was to characterise the RAGs of the exclusively neotropical bat Artibeus planirostris (Chiroptera: Phyllostomidae) macroscopically, microscopically and ultrastructurally. The RAGs were composed of a prostatic complex with two regions (ventral and dorsal) and paraurethral and bulbourethral glands, but no seminal vesicles. The ventral region had an undefined epithelium, with secretory and basal cells, and its secretions were periodic acid-Schiff (PAS) positive. The dorsal region received both deferens ducts, had a columnar pseudostratified epithelium with secretory and basal cells. There were two types of secretions from the dorsal region: one that was basophilic and another that was mixed PAS positive and PAS negative. The paraurethral glands were dispersed in the connective tissue of the urethra, whereas the bulbourethral glands were located in the penile root. Histological and ultrastructural data confirmed the prostatic nature of the ventral and dorsal regions and the holocrine nature of the ventral region, with the latter finding never having been described previously for the prostate gland. Our findings demonstrate the wide discrepancy of RAGs between A. planirostris and other mammals in terms of their composition, structure and morphology.

Application of absolute qPCR as a screening method to detect illicit 17ß-oestradiol administration in male cattle.

It has been previously demonstrated that the progesterone receptor gene is up-regulated in the sex accessory glands of pre-pubertal and adult male bovines after 17ß-oestradiol treatment. In the present study, a qualitative screening method was optimised to detect 17ß-oestradiol treatment using absolute quantification by qPCR of the progesterone receptor gene to determine the amount of gene expression in bulbo-urethral glands. An external standard curve was generated and developed with TaqMan® technology. Based on two in vivo experiments, the decision limit CCα, sensitivity and specificity of this screening method were established. Trial 1 consisted of 32 Friesian veal calves divided into two groups: group A (n = 12), consisting of animals treated with four doses of 17ß-oestradiol (5 mg week(-1) per animal); and group B (n = 20), consisting of control animals. Trial 2 was performed on 26 Charolaise beef cattle that either received five doses of 17ß-oestradiol (group C; 20 mg week(-1) per animal; n = 6) or remained untreated (group D; n = 20). Further, progesterone receptor gene expression was evaluated in beef and veal calves for human consumption. A specific CCα on 20 Piedmontese control beef cattle was calculated to include these animals in a field investigation. Five out of 190 beef cattle and 26 out of 177 calves tested expressed the progesterone receptor gene above their respective CCα and they were classified as being suspected of 17ß-oestradiol treatment. Additionally, 58% of veal calves that tested suspect via qPCR exhibited histological lesions of the bulbo-urethral gland tissue, which are typical of oestrogen administration and are consistent with hyperplasia and metaplasia of the glandular epithelium.

A histologic study on growth promoter target organs of slaughtered beef in Molise Region (Italy).

A gross pathology and histological investigation was carried out on bovine target organs of anabolic substances in the Molise Region (Italy). One hundred forty-four bovines (12-24 months old, 123 males and 21 females) were included in the survey. An antemortem assessment of their behavior and clinical examination were performed. After slaughter, samples of prostate, Cowper's glands, Bartholin's glands, mammary gland, ovaries, thymus and thyroid were collected, inspected and processed for histopathology, as suggested in the guidelines of the Italian national program for residue surveillance (PNR). Overall, 15.3% of the examined animals were classified as "suspect," 44.4% were classified as "uncertain," and the remaining 40.3% were classified as "negative." The most frequent lesion was a severe thymus atrophy with fat infiltration (15.4% of males and 14.3% of females), strongly suggesting the illegal use of corticosteroids.

Glycosaminoglycans in the accessory sex glands, testes and seminal plasma of alpaca and ram.

The viscous nature of alpaca semen limits its use in cryopreservation and other assisted reproductive technologies. The cause and source of this viscosity is unknown although it has been postulated, but never proven, that glycosaminoglycans (GAGs) secreted by the bulbourethral gland are responsible. The present study investigated the concentration and composition of GAGs in alpaca seminal plasma, testes, bulbourethral gland and prostate gland and compared them to those in the ram to determine the relationship between seminal plasma GAGs and viscosity and to identify the source of seminal plasma GAGs. Alpaca seminal plasma contained more GAGs than ram (P<0.001) and the predominant GAG, keratan sulfate, was correlated with viscosity (P=0.05, R(2)=0.2635). The alpaca bulbourethral gland contained most GAGs compared with prostate or testis (P<0.001). In the ram, the prostate contained most GAGs. These findings suggest that GAGs, particularly keratan sulfate, may be the cause of seminal plasma viscosity in alpacas, and that the seminal plasma GAGs originate from the bulbourethral gland.

Epithelial Bmp (Bone morphogenetic protein) signaling for bulbourethral gland development: a mouse model for congenital cystic dilation.

The bulbourethral gland (BUG) is a male-specific organ, which secretes part of the semen fluid. As the BUG is located in the deep pelvic floor, its developmental process is still unclear. Bone morphogenetic protein (Bmp) signaling plays pivotal roles in various organs. However, the function of Bmp signaling for BUG development is still unclear. The present study aimed to elucidate the role of Bmp signaling in the development of the BUG. We observed the prominent nuclear accumulation of phosphorylated (p) SMAD1/5/8, the downstream molecules of Bmp signaling, during BUG epithelial development. These results suggest that Bmp signaling contributes to BUG development. Bmp receptor1a (Bmpr1a) is known as the major type 1 signal transducer in some organogeneses. To analyze the Bmp signaling function for BUG development, we examined epithelial cell-specific Bmpr1a gene conditional mutant mice utilizing the tamoxifen-inducible Cre recombinase system. We observed cystic dilation and epithelial hyperplasia of the BUG in the Bmpr1a conditional knockout mice. The mutant cystic BUG specimens also showed inflammatory lesions. These BUG abnormalities resembled some of the BUG malformations observed in human congenital syndromes. The current study suggests that Bmp signaling possesses an essential role in BUG development and homeostasis. This would be the first report showing that the mutation of the Bmpr1a gene in the BUG epithelia phenocopied some abnormalities of human congenital syndromes affecting the BUG duct.

Progesterone receptor gene expression in the accessory sex glands of veal calves.

This study investigated progesterone receptor (PR) cDNA expression in the testes, prostate and bulbourethral glands of prepubertal calves treated experimentally with high and low doses of 17beta-oestradiol and with testosterone. Tissue samples were examined histologically and immunohistochemically for PR. Western blot analysis and quantitative PCR against PR was performed on cDNA and protein extracted from the same tissues. Bulbourethral glands from animals treated with low and high dosages of 17beta-oestradiol had 39- and 429-fold increases of PR transcript, respectively, compared with controls. In the prostate there were 7.5- and 16-fold increases, respectively. Animals treated with testosterone showed no increases in PR transcript. The results demonstrate that 17beta-oestradiol specifically induces marked overexpression of the PR gene and protein, particularly in the bulbourethral gland.

Expression and localization of acrosin inhibitor in boar reproductive tract.

Proteinases and proteinase inhibitors play key roles in almost all physiological processes. Proteinase inhibitors are present in all tissues and body fluids. They interfere with the activity of proteinases and thus maintain homeostasis. The main role of proteinase inhibitors in the reproductive tract is the inactivation of prematurely released hydrolytic enzymes from damaged spermatozoa and the protection of reproductive tracts and spermatozoa against proteolytic degradation. In the boar reproductive system, acrosin inhibitors are found in seminal plasma and on spermatozoa. The amino acid sequence of seminal plasma and sperm-associated acrosin inhibitors is 90% identical, and their biochemical properties have been completely resolved. However, their origin and localization have not been fully elucidated. Using rabbit polyclonal antibody, we have studied the expression and localization of the seminal plasma acrosin inhibitor in the boar reproductive tract. The antibody recognizes a 12-kDa band in extracts from the cauda epididymidis, seminal vesicles, prostate, and Cowper's glands, and immunofluorescence has revealed the acrosin inhibitor in the epithelium and lumen of these organs. Gene expression of the acrosin inhibitor has been studied by reverse transcription together with the polymerase chain reaction. Acrosin inhibitor mRNA transcript is detectable in the epididymis, seminal vesicles, prostate, and Cowper's glands. The antibody has localized the acrosin inhibitor on the surface of epididymal and ejaculated spermatozoa in the acrosomal region. In extracts from epididymal and ejaculated spermatozoa, the specific antibody recognizes acrosin inhibitor at 8 kDa and 12 kDa. The presence of acrosin inhibitor on the sperm surface as a protective molecule for receptors mediating the sperm-zona pellucida binding suggests that it is crucial for the reproductive process.

[Age-related changes in glycoprotein composition in human bulbourethral glands].

Bulbourethral gland glycoprotein (GP) composition was studied histochemically in men of different age groups (varying from neonatal period up to 90 years). No less than 5 cases were examined for each age group. In children and juveniles, neutral GP prevailed in the glandular cells, the proportion of neutral GP gradually decreasing with puberty. In youths and in men of mature age, acid GP became the predominant component of glandular secretion. In elderly and senescent men the ratio of acid and neutral GP again was found to change in favor of the latter. In all the age groups, acid GP were represented mainly by sialomucins, both sialidase-labile and sialidase-resistant. In the glands of men aged 17 years and older, sulphomucins were also detected; the intensity of their synthesis increased from the youth to the elderly period and decreased in senescent persons. Functional significance of different types of GP in the secretion of bulbourethral glands is discussed as well as the possible hormonal machanisms of their synthesis.

Lectin histochemistry of the boar bulbourethral glands.

The present study describes, for the first time, the glycosidic content of boar bulbourethral glands using lectin histochemistry. Fourteen horseradish peroxidase- or digoxigenin-labelled lectins with different carbohydrate specificities were used in samples obtained from 3 healthy Landrace boars. The results obtained indicate that endpiece and duct cells synthesize and secrete mainly O-glycoproteins with alpha- and beta-D-N-acetylgalactosamine, beta-D-galactose-beta(1-->3)-D-N-acetylgalactosamine, D-N-acetylglucosamine and neuraminic acid residues. Glycoproteins secreted by bulbourethral glands have a role in the protection and lubrication of the urethra. In addition, they may be also involved in the regulation of the sperm metabolic activity and in the maintenance of the structural integrity of acrosomal and plasma membranes.

[Fertility alpha-2-microglobulin expression in the human bulbourethral glands]. / Ekspressiia al'fa 2-mikroglobulina fertil'nosti v bul'bouretral'nykh zhelezakh cheloveka.

Fertility alpha-2-microglobulin (FAMG) was demonstrated in the human bulbourethral glands by the method of indirect immunoperoxidase staining using monoclonal antibodies against FAMG. The product of the immunohistochemical reaction was primary found in the duct cells. Weak immunopositive reaction was observed in single epitheliocytes of the secretory units. FAMG-positive granularity was typically found in the supranuclear cytoplasmic zone, that corresponds to the localization of secretory granules both in the glandulocytes and in the duct cells. These results suggest, that the bulbourethral glands are probably the additional producers of human seminal plasma FAMG.

Metabolism of oestrone and oestradiol-17beta to conjugated steroids by the accessory sex glands of the male pig.

Oestrogens are secreted in large amounts by boar testes and are known to have a synergistic effect with testosterone on the production of large volumes of seminal plasma. Thus, oestrogens play a role in regulating the large accessory sex glands in the boar. Since testosterone metabolites (e.g. 5alpha-dihydrotestosterone) account for much of its action in target tissues we have looked at the metabolism of oestrogens in the accessory sex glands of the male pig. Tissues from seminal vesicles and bulbourethral glands of 6-week-old castrate and intact males, and 12-week-old castrate animals, were incubated with (3)H-labelled oestrone and oestradiol-17beta. Aliquots of spent culture medium and of methanolic tissue extracts were taken to measure radioactivity, prior to separation of unconjugated and conjugated steroids on Waters C(18) Sep-Pak cartridges. About one-third of the radioactivity appeared as conjugates in the media from both glands with each oestrogen. Subsequently, sulphoconjugated steroids and glucuronidates were recovered in series from C(18) cartridges after solvolysis and enzyme hydrolysis respectively. Furthermore, about one-third of the conjugated fraction in each case remained unhydrolysed after these treatments. In conclusion, it is clear that a study of the actions of oestrogens on these glands must consider the dynamics of metabolism of the oestrogens presented to them by the testes and would include conjugation of steroids by the glands themselves.