Expression of an antimicrobial peptide identified in the male reproductive system of rats

Bin1b is a β-defensins-like molecule originally isolated from the rat epididymis. Owing to its bactericidal activity, Bin1b may have therapeutic properties suitable for the treatment of sexually transmitted diseases. The amino terminus of the mature Bin1b peptide contains a conserved myristoylated Gly residue. We studied the requirement of the terminal myristoylated Gly residue in the bactericidal activity of Bin1b and found that the terminal myristoylated Gly residue is not essential for the bactericidal activity. In addition, we expressed the tandem repeats of Bin1b in Escherichia coli and found that two tandem repeats of Bin1b protein were successfully expressed. The bacterially expressed tandem Bin1b repeats may be used in a diverse array of biochemical and cell biological studies.     

Coccidioidomycosis of the male reproductive tract

Coccidioides immitis infection of the male reproductive tract is a rare entity that can evade diagnosis and pose a dilemma in management. Initially, patients are often evaluated for malignancy or other infections such as tuberculosis. In the past, surgery was the only management option for C. immitis infection of the male reproductive tract, but azole therapy now provides an adjunct or an alternative. We describe two patients who received azole therapy for C. immitis infection of the male reproductive tract. One received fluconazole for prostatic disease, while one received surgery followed by itraconazole for testicular disease. After 12 months of therapy, both remain asymptomatic and have decreased antibody titers against C. immitis.Disclaimer: The Views expressed herein are those of the authors do not reflect the official policy or position of the Department of the Air Force, Department of the Army, Department of Defence, or the U.S. Government.     

Defective postnatal development of the male reproductive tract in LGR4 knockout mice

The final outcome of tube elongation and branching is to maximize the epithelial exchange surfaces in tubular organs. The molecular and cellular basis of these processes is actively studied in model organs such as mammary glands, liver and kidney, but they remain almost unexplored in the male reproductive tract. Here, we report that the orphan G protein-coupled receptor LGR4/GPR48 plays a role in the postnatal tissue remodeling needed for elongation and convolution of the efferent ducts and epididymis. In LGR4 knockout male mice, tube elongation fails, resulting in a hypoplastic and poorly convoluted tract. Cell proliferation is dramatically reduced in KO affected tissues, providing an explanation to the observed phenotype. Detailed analysis showed that LGR4 inactivation manifests differently in the affected organs. In efferent ducts, immune cells infiltrate the epithelium and reach the lumen, blocking the transit of sperm and testicular fluid. In addition, the hypoplasia and low convolution result in a reduction of the epithelial area involved in liquid reabsorption. Both phenomena contribute in tissue swelling upstream the blockade due to liquid and sperm accumulation, with secondary damaging effects on the germinal epithelium. In the epididymis, the thin and highly convoluted duct is replaced by a large cystic tube which is surrounded by a thick condensation of mesenchymal cells. The abnormal organization of the cellular compartments in and around the ducts suggests that LGR4 might play a role in epithelial-mesenchymal interactions. Altogether, our data identify LGR4 as an important signaling molecule implicated in the tube morphogenesis of the male reproductive tract.     

Gossypol modulation of nucleotide metabolizing enzymes in the reproductive tract of male rats

The activities of several pivotal nucleotide metabolizing enzymes from the testis and vasal sperm of rats treated for 7 wk with 0, 20 or 30 mg X kg X day gossypol acetic acid were examined. Total testicular lactate dehydrogenase (LDH) activity increased 40% above control in the highest treatment group examined. However, the specific activity of the testis-specific isozyme of LDH, LDH-C4, decreased to 50 and 20% of control in the 20 and 30 mg X kg X day treatment groups, respectively. Basal soluble adenylate cyclase from a 100,000 X g supernatant of testis homogenate exhibited a 25% decrease in activity only in the 30-mg treatment group. Basal adenylate cyclase activity in the testicular membrane fraction increased 20 to 30% above control in response to gossypol administration. Testis membranes from the 20- and 30-mg treatment group exhibited a 2- and 4-fold greater activation of adenylate cyclase by guanine nucleotides. In vitro dose-response curves showed a half-maximal inhibitory concentration (IC50) for inhibition of soluble testicular adenylate cyclase by gossypol of 400 microM in each treatment group. Caudal epididymal sperm adenylate cyclase activity decreased to 25% of control levels in gossypol-treated animals, and the in vitro sensitivity of the enzyme to the inhibitory effects of gossypol increased 4-fold. IC50 values for gossypol inhibition of sperm adenylate cyclase decreased from 200 microM in control animals to 75 and 50 microM in the 20 and 30 mg X kg X day treatment groups, respectively. Cyclic adenosine 3':5' monophosphate phosphodiesterase activity in caudal sperm increased 6-fold in the 20- and 30-mg treatment groups. These results demonstrate that nucleotide metabolizing enzymes in sperm are major targets for the actions of gossypol and provide a possible mechanism for the inhibition of normal sperm function by this compound.     

Role of Toll-like receptors and defensins in antimicrobial protection of urogenital tract in females

Levels of expression of hBD-1 gene (beta-defensin 1) and Toll-like receptors (TLR1, TLR2, TLR6) in cells of cervical mucosa in healthy nonpregnant and healthy pregnant women as well as in pregnant women with urogenital infection was measured by developed RT-PCR systems. During normal pregnancy compared with nonpregnant women, increase of TLRs genes expression which was correlated with increase of hBD-1 gene expression was observed. During urogenital infection in pregnant women compared with healthy pregnant, 10- fold and 50-fold increase of TLR1 and TLR2 genes expression respectively was associated with 2.5-fold decrease of hBD-1 gene expression in cervical mucosa. In group of women with untrauterine infection more marked increase of TLRs genes expression was observed. Thus significant changes (TLRs, antimicrobial peptides, cytokines etc.) in cells of cervical mucosa can be used as prognostic criteria for development of intrauterine infection.     

Exposure with the environmental estrogen bisphenol A disrupts the male reproductive tract in young mice

Environmental estrogens (endocrine disruptive chemicals) have been shown to affect reproduction in wild life and it has been reported that maternal exposure with those chemicals have adverse effects on the male reproductive tract. However, little is known about the potential effects of prepubertal or pubertal exposure with environmental estrogens on the male reproductive tract. Here we examine plasma hormone levels and histology in the testis of mice following either 4- or 8-week oral administration of bisphenol A. Plasma free testosterone levels were dramatically decreased following 8 weeks of bisphenol A treatment compared with control group and morphologically multinucleated giant cells having greater than three nuclei were found in seminiferous tubules in the testis following the 8-week bisphenol A treatment. No differences in plasma corticosterone and luteinizing hormone levels were seen between bisphenol A and control groups. Thus, exposure with bisphenol A around pubertal period may directly disrupt the male reproductive tract. These facts suggest that more detailed studies will warrant the assessment of the risk to the developing human testis from exposure to bisphenol A and other environmental estrogens in prepubertal and pubertal period.     

Sperm migration and selection in the reproductive tract of female mice is mostly affected by male genotype

The aim of the present study is to assess the influence of male and female genotypes on the transport of sperm to the site of fertilization. We mated B10.BR, B10.BR-Ydel and BALB/c males with B10.BR and BALB/c females and then analyzed the quality and quantity of spermatozoa found five hours post coitus in three successive parts of the female reproductive tract. We found that B10.BR and B10.BR-Ydel spermatozoa are very effectively selected by the uterotubaljunction and other barriers of the female genital tract. On the contrary, severely deformed BALB/c spermatozoa appeared to be able to cross selective barriers and were present both in oviducts and in cumulus oophorus. It cannot be excluded that these morphologically abnormal male gametes take part in fertilization. B10.BR-Ydel spermatozoa were very rarely observed above the uterotubaljunction. This shows that in vivo they migrate with delay and with difficulties pass the border between uterus and oviducts. This finding is in agreement with previous in vitro analyzes, which revealed many irregularities in movement of B10.BR-Ydel spermatozoa. Sperm quality and quantity in the reproductive tracts of B10.BR and BALB/c females were convergent if they were mated with males belonging to one strain, proving that migration and selection of spermatozoa in the female genital tract depend mostly on male genotype.     

The antimicrobial action of histones in the reproductive tract of cow

An infection of any part of female reproductive tract can severely interfere with fertility and reproduction. The fluids and epithelium from the lumen of the female reproductive tract (uterus, oviduct and ovarian follicle) are a known source of antimicrobial action in several species. In this study, we compared the antimicrobial properties of fluids from the reproductive tract of a cow. After removal of small molecules, we demonstrated that there is an antimicrobial activity connected with a fraction of compounds with a molecular mass range between 3500 and 30,000. The most probable candidates responsible for the observed antimicrobial effect were subsequently identified by mass spectroscopy as histones H2A type 2-C, H2B type 1-K, H3.3, and H4. The antimicrobial role of histone H2B was further confirmed by using an antibody against this histone.     

Aquaporin 9 expression along the male reproductive tract

Fluid movement across epithelia lining portions of the male reproductive tract is important for modulating the luminal environment in which sperm mature and reside, and for increasing sperm concentration. Some regions of the male reproductive tract express aquaporin (AQP) 1 and/or AQP2, but these transmembrane water channels are not detectable in the epididymis. Therefore, we used a specific antibody to map the cellular distribution of another AQP, AQP9 (which is permeable to water and to some solutes), in the male reproductive tract. AQP9 is enriched on the apical (but not basolateral) membrane of nonciliated cells in the efferent duct and principal cells of the epididymis (rat and human) and vas deferens, where it could play a role in fluid reabsorption. Western blotting revealed a strong 30-kDa band in brush-border membrane vesicles isolated from the epididymis. AQP9 is also expressed in epithelial cells of the prostate and coagulating gland where fluid transport across the epithelium is important for secretory activity. However, it was undetectable in the seminal vesicle, suggesting that an alternative fluid transport pathway may be present in this tissue. Intracellular vesicles in epithelial cells along the reproductive tract were generally poorly stained for AQP9. Furthermore, the apical membrane distribution of AQP9 was unaffected by microtubule disruption. These data suggest that AQP9 is a constitutively inserted apical membrane protein and that its cell-surface expression is not acutely regulated by vesicular trafficking. AQP9 was detectable in the epididymis and vas deferens of 1-wk postnatal rats, but its expression was comparable with adult rats only after 3--4 wk. AQP9 could provide a route via which apical fluid and solute transport occurs in several regions of the male reproductive tract. The heterogeneous and segment-specific expression of AQP9 and other aquaporins along the male reproductive tract shown in this and in our previous studies suggests that fluid reabsorption and secretion in these tissues could be locally modulated by physiological regulation of AQP expression and/or function.     

The distribution of rat testibumin in the male reproductive tract

A glycoprotein, designated CMB-1, has been identified in media from Sertoli cell-enriched cultures that increases in concentration in response to follicle-stimulating hormone (FSH) and testosterone. Subsequent studies indicated that CMB-1 is immunologically related to albumin and alpha-fetoprotein and is concentrated in the luminal compartment of the testis in adult rats. Thus, CMB-1 was termed testibumin. The goal of the present study was to determine the concentrations of this protein in testes, epididymides, and serum of normal rats between 10 and 180 days of age and to compare them to rat androgen-binding protein (rABP). Testibumin concentration in rat testes increased with age and peaked at Day 60; thereafter, unlike rABP, its concentration declined, reaching a plateau by 150 days of age. Testibumin concentration in the epididymal compartment also increased with age and peaked at Day 90; thereafter, its concentration remained relatively unchanged. Unlike rABP, which accumulates in the caput epididymis, testibumin did not accumulate preferentially in any particular region of the epididymis. In spite of the marked changes of testibumin concentration in the male reproductive tract, the levels in blood remained relatively constant between 10 and 180 days of age. In adult male and female rats, the serum concentrations of testibumin were similar. Following orchiectomy, serum testibumin concentration decreased by 50% with an apparent t1/2 of approximately 8 h. The presence of immunoreactive macromolecules in other species that share epitopes with rat testibumin was also investigated. Material in human sera and extracts of human and monkey testes cross-reacts with rat testibumin. After [35S]methionine was added to the primary Sertoli cell-enriched cultures, anti-testibumin antiserum selectively immunoprecipitated a radiolabeled protein with the same electrophoretic mobility as purified testibumin on sodium dodecyl sulfate (SDS)-polyacrylamide gels. We conclude that 1) rat testibumin is synthesized and secreted by Sertoli cell-enriched cultures; 2) the relative concentrations and distribution of testibumin in testis, epididymis, and serum of the rat as a function of age are strikingly different from those of rABP; 3) rat testibumin shares epitopes with proteins in human serum and testicular extracts of monkey and man.     

Spermiophagy in the male reproductive tract of some passerine birds

In order to elucidate the locus and means of spermiophagy in passerine birds, we examined histologically the entire male reproductive tract of sexually mature birds of three passerine species with different forms of sperm competition, namely, the alpine accentor (Prunella collaris), the redflanked bush robin (Tarsiger cyanurus), and the Bengalese finch (Lonchura striata var. domestica). Spermiophagy occurred consistently and frequently in the epithelial layer of the seminal glomera and ejaculatory duct in each species, which were regularly identified by non-ciliated epithelial cells. The epithelial spermiophagy was occasional or infrequent in other portions of the seminal tract, and spermiophagy by macrophages was uncommon throughout the tract. Quantitative data in the seminal glomera and ejaculatory duct gave no clear answer concerning a possible relationship between the epithelial spermiophagy and different levels of sperm competition among these passerine species. In conclusion, the epithelial lining of the terminal region of the seminal tract is the main site for spermiophagy in the male reproductive tract of these passerine species, which activity serves to maintain the quality of semen by eliminating infertile spermatozoa as well as sperm remaining at the end of the breeding season.     

Expression of cysteine sulfinate decarboxylase (CSD) in male reproductive organs of mice

Cysteine sulfinate decarboxylase (CSD) is the rate-limiting biosynthetic enzyme of taurine, but it is still controversial whether the male reproductive organs have the function to synthesize taurine through CSD pathway. The present study was thus undertaken to detect CSD expression in male mouse reproductive organs by RT-PCR, Western blot and immunohistochemistry. The results show that CSD is expressed both at the mRNA and protein levels in the testis, epididymis and ductus deferens. The relative levels of both CSD mRNA and protein increase from the testis to the epididymis and to the ductus deferens. Immunohistochemical results demonstrate that the main cell types containing CSD are Leydig cells of testis, epithelial cells and some stromal cells throughout the efferent ducts, epididymis and ductus deferens. These results suggest that male genital organs have the function to produce taurine through the CSD pathway, although quantifying the relation of CSD expression to taurine synthesis and the exact functions of taurine in male genital organs still need to be elucidated in future studies.     

Seminal plasma and male factor signalling in the female reproductive tract

In mammals, insemination results in the transmission of seminal factors that act, in the female reproductive tract, to promote sperm survival, to “condition” the female immune response to tolerate the conceptus and to organise molecular and cellular changes in the endometrium to facilitate embryo development and implantation. These events are initiated when signalling agents, including transforming growth factor-β and other cytokines and prostaglandins secreted by seminal vesicle and prostate glands, interact with epithelial cells in the cervix and uterus to activate cytokine synthesis and to induce cellular and molecular changes resembling a classical inflammatory cascade. The consequences are the recruitment and activation of macrophages, granulocytes and dendritic cells, which have immune-regulatory and tissue-remodelling roles that culminate in improved endometrial receptivity to the implanting embryo. Cytokines elicited by seminal activation have embryotrophic properties and also contribute directly to the optimal development of the early embryo. This review summarises our current understanding of the physiology of responses to seminal plasma in the female reproductive tract and considers the evolutionary significance of seminal plasma in influencing female tissues to promote the success of pregnancy.The author acknowledges the support of the NHMRC of Australia Fellowship and Program Grant schemes.     

Testosterone undecanoate in the functional compartments of the male reproductive tract

Background

Assessment of testosterone undecanoate’s (TU) presence in the functional compartments of the male reproductive tract has never been performed despite the evidence that its documented beneficial effect in male infertility might be mediated through an epididymal action and this study was set to examine this possibility.

Materials and methods

In 18 normozoospermic volunteers TU has been administered (40 mg t.i.d.) for 6 days with serum measurements of TU, total testosterone (T), DHT, E2, SHBG, FSH, LH, and PRL before and at the end of medication. Steroid hormones (T, E2, and TU) were also assayed in seminal plasma. In a selected group of 7 men with previously diagnosed non-obstructive azoospermia TU, T, and E2 were assayed in the extracts of testicular biopsy material taken before ICSI and at the end of the same medication.

Results

A marked rise of serum DHT (average 148%, P < 0.001) has been found after treatment, whereas T, E2, FSH, LH, SHBG, and PRL did not significantly change. Measurable amounts of TU were found in the serum of all men but only in 6 cases in seminal plasma (11.1 ± 8.0 ng/mL) and all of them in semen delivered 7–8 h after the last TU capsule was taken. In dilution fluid from testicular tissue extracts, no detectable amounts of TU were found whereas mean values of 92.5 ± 54.3 pg/mL and 43.8 ± 16.3 ng/mL for E2 and T were observed. Positive correlations among TU and E2, T or DHT concentrations were found in serum samples (P < 0.01, 0.02, and 0.002) as well as between E2 and T (P < 0.01), E2 and DHT (P < 0.001), or T and DHT (P < 0.001).

Conclusion

It is concluded that TU was identified and measured for the first time in seminal plasma of a fair percentage (33%) of men on this medication and was associated in all men with a marked rise of DHT concentration, a known epididymal function promoter, in the absence of an effect on pituitary and gonadal activity. On this evidence, it appears that a beneficial effect of TU on epididymal function may be a distinct possibility.     

Estrogen in the adult male reproductive tract: A review

Testosterone and estrogen are no longer considered male only and female only hormones. Both hormones are important in both sexes. It was known as early as the 1930's that developmental exposure to a high dose of estrogen causes malformation of the male reproductive tract, but the early formative years of reproductive biology as a discipline did not recognize the importance of estrogen in regulating the normal function of the adult male reproductive tract. In the adult testis, estrogen is synthesized by Leydig cells and the germ cells, producing a relatively high concentration in rete testis fluid. Estrogen receptors are present in the testis, efferent ductules and epididymis of most species. However, estrogen receptor-α is reported absent in the testis of a few species, including man. Estrogen receptors are abundant in the efferent ductule epithelium, where their primary function is to regulate the expression of proteins involved in fluid reabsorption. Disruption of the α-receptor, either in the knockout (αERKO) or by treatment with a pure antiestrogen, results in dilution of cauda epididymal sperm, disruption of sperm morphology, inhibition of sodium transport and subsequent water reabsorption, increased secretion of Cl^-, and eventual decreased fertility. In addition to this primary regulation of luminal fluid and ion transport, estrogen is also responsible for maintaining a differentiated epithelial morphology. Thus, we conclude that estrogen or its α-receptor is an absolute necessity for fertility in the male.     

Sperm morphological abnormalities appearing in the male rabbit reproductive tract

The role of the excurrent duct system in producing and/or eliminating morphologically abnormal spermatozoa may modify the semen parameters and interfere with sperm fertilizing capacity. To study this process, changes in the morphology of spermatozoa during their transit through the reproductive tract in sexually mature rabbits were investigated. The incidence of head, midpiece and tail abnormalities as well as of multiple defects in a single spermatozoon, and the position of the cytoplasmic droplet along the sperm midpiece were evaluated in samples from the testis, 6 regions of the epididymis and the vas deferens. Spermatozoa were characterized by rapid migration of the cytoplasmic droplet when passing from the proximal to the distal caput of the epididymis, and spermatozoa with no droplet predominated in the distal epididymis and vas deferens. In passing from the testis to the proximal caput of the epididymis, the incidence of spermatozoa with an abnormal midpiece and those with multiple defects decreased significantly. The proportion of spermatozoa with abnormal heads was also lower in the testis, but no statistically significant differences were found, whereas there was no change in the proportion of those with abnormal tails. These results indicate that there must be a mechanism for the disposal of defective spermatozoa. No evidence of spermiophagy by luminal macrophages was observed in the extracts, although a few spermatozoa exhibited signs of degeneration, suggesting, that although intraepithelial phagocytosis has not been clearly demonstrated in the nonexperimental rabbit, sperm cells may undergo a form of autolysis within the lumen of the duct.