The fine structure and development of the seminal vesicles and prostate in the fetal rat

Summary The development of the seminal vesicle from the Wolffian duct and of the prostate from the urogenital sinus has been studied in rat fetuses from day 14 of gestation to birth with the use of the electron microscope. Prior to the onset of androgen secretion, the cells of the urogenital sinus and the caudal part of the Wolffian duct have a simple undifferentiated appearance. After the onset of androgen secretion by the fetal testes at day 15, intracytoplasmic confronting cisternae of the granular reticulum appear in both urogenital sinus and Wolffian duct. Portions of the granular endoplasmic reticulum of the urogenital sinus become distended with a finely granular, moderately dense material. In the urogenital sinus, many hemidesmosomes are formed at the basal surface of the epithelium. Specializations of the extracellular materials are present opposite the hemidesmosomes. The formation of the seminal vesicles and the prostate begins at day 18–19 of gestation. The cells of the seminal vesicle are taller than the Wolffian duct cells from which they arise, the granular endoplasmic reticulum increases moderately in amount, and a patent lumen is formed. The cells of the fetal prostate do not differ greatly from those of the urogenital sinus from which they arise except that the prostatic cells initially lack hemidesmosomes. The fine structural changes are discussed in relation to the onset of fetal androgen secretion, the formation of the organs, and the functions of the cells in adult life.This study was supported by Contract No. 69-2104, Program Project HD-02282, Health Sciences Advancement Award FR-02084 and a Research Career Development Award (1-K3-GM-28, 214-01) from the National Institutes of Health. The author wishes to acknowledge the technical assistance of Mrs. Stephanie Krah.     

Sexual selection affects the sizes of the mammalian prostate gland and seminal vesicles

The accessory reproductive glands of male mammals contribute the bulk of the secretions in which spermatozoa are transported to the female tract during copulation. Despite their morphological diversity,and the chemical complexity of their products,little is known about the possible effects of sexual selection upon these glands in mammals. Here we consider the seminal vesicles and prostate glands in a sample of 89 species and 60 genera representing 8 Orders of mammals. The sizes of the accessory glands are analysed in relation to body weight and testes weight. Both the seminal vesicles size and prostate size (corrected for body weight) correlate positively with relative testes size in this sample; this finding remains highly significant after application of procedures to correct for possible phylogenetic biases in the data set. The accessory reproductive glands are also significantly larger in those mammals which have large relative testes sizes,and in which the likelihood of sperm competition is greatest. These results support the hypothesis that sexual selection has played an important role in the evolution of the mammalian prostate gland and seminal vesicles.     

Characterization of the major proteins of bovine seminal fluid by two-dimensional polyacrylamide gel electrophoresis

Recently, we demonstrated that the major proteins from bovine seminal plasma BSP-A1, -A2, -A3 and -30-kDa (collectively called BSP proteins) specifically interact with choline phospholipids. These proteins coat the surface of the spermatozoa after ejaculation and are believed to play an important role in membrane modifications occurring during capacitation. In this study we determined the isoelectric point (pl) and analysed the molecular heterogeneity of BSP proteins. Total protein from bovine seminal plasma (CBSP) and purified BSP proteins were iodinated using chloramine T. Samples were reduced, denatured, separated by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), and visualized by autoradiography. Analysis of CBSP proteins demonstrated the presence of polypeptides migrating in the pH range of 3.5–7.8 and at molecular weights (Mr) between 6 and 100 kDa. Many isoforms of each BSP protein were found when purified iodinated proteins were analysed by 2D-PAGE. BSP-A1 was found at a Mr of 16.5 kDa and in the range of pl of 4.7–5.0; BSP-A2 at 16 kDa and at a pl of 4.9–5.2; BSP-A3 at 15 kDa and at a pl of 4.8–5.2, and BSP-30-kDa at 28 kDa and at a pl of 3.9–4.6. Similar results were obtained with immunolocalization of BSP proteins after Western blot using specific antibodies. The treatment of purified iodinated BSP proteins with neuraminidase increased the pl of BSP-30-kDa to 4.8–5.0 and decreased its Mr to 25 kDa, but no change was observed for BSP-A1, -A2 and -A3. The treatment of BSP proteins with sulfatase or acid phosphatase modified neither their Mr nor their pl. Furthermore, when CBSP proteins were separated in 2D-PAGE and the gels stained for glycoproteins with dansyl hydrazine, BSP proteins were among the major glycoproteins found in the bovine seminal plasma. In conclusion, BSP proteins are acidic and have several isoforms. Furthermore, the heterogeneity of BSP-30-kDa is mainly due to its sialic acid content. © 1994 Wiley-Liss, Inc.     

Continuous culture of Humicola lutea 120-5 for acid proteinase production

Acid proteinase production by the fungus Humicola lutea 120-5 in continuous culture was studied. The maximum activity of the culture broth reached 2200 U/ml at a dilution rate (D) of 0.05/h. The continuous process was carried out for 1 month without any bacterial contamination, due to low pH (3.0–3.5) during the cultivation.     

Partial amino acid sequence of a human seminal plasma peptide with inhibin-like activity

An extract of human seminal plasma was found to have inhibin-like activity. The active factor was purified to homogeneity by ion exchange chromatography, molecular sieving and high performance liquid chromatography. The purified material has a mass of approximately 5 kDa and is very basic. Amino acid analysis showed the presence of approximately 35 residues while the sequencing data allowed the determination of the N-terminal 31 amino acids. There is a possibility of an additional 2–4 residues at the C-terminus, which could not be determined.     

Binding of a major secretory protein from bull seminal vesicles to bovine spermatozoa

Summary The seminal vesicles synthesize in an androgen-dependent manner a neutral protein of 13.5 kDa molecular weight that makes up about 40% of their secretion (major protein). An antiserum against this protein raised in rabbits was used to localize the antigen within the seminal vesicles. In addition to intraluminal secretion of the seminal vesicles and the ampulla of the vas deferens, ejaculated and ampullary spermatozoa revealed an intense immunoreaction, which was restricted to the neck region of the sperm head and the middle piece, while the principal piece of the tail as well as the sperm head were devoid of immunoreactive material. Comparison of spermatozoa taken from the tail of the epididymis with ampullary spermatozoa showed that about 90% of the latter, but only 10–20% of the former presented this distributional pattern of immunoreactive sites. Epididymal epithelium as well as calf seminal vesicle epithelium showed no immunoreactivity with major protein antiserum. Using a pre-embedding staining technique with gold-labeled primary or secondary antibodies, respectively, no immunostaining could be achieved at the ultrastructural level. Incubation experiments of epididymal spermatozoa in EGTA-containing solutions in the absence of calcium resulted in a gradual labilization and eventual loss of the plasma membrane of the sperm middle piece. After removal of (at least part of) the plasma membrane, bound major protein could be visualized immunohistochemically close to the mitochondria of the middle piece using a gold-labeled primary or secondary antibody. The acceptor site for major protein therefore seems to reside inside the plasma membrane of the sperm middle piece. Incubation of epididymal spermatozoa in phospholipase-containing solutions removed the acceptor site from the spermatozoa. Separation by polyacrylamide treatment of proteins from epididymal sperm cells extracted by sodium hydroxide or phospholipase treatment, subsequently transblotted on nitrocellulose sheets and directly labeled with gold-tagged major protein, demonstrated a protein duplet with a molecular weight of 65 and 67 kDa, respectively, which appears to represent the specific binder of major protein underneath the sperm surface. Binding of major protein to this 66 kDa acceptor site is regarded as a physiological event that may be related to the onset of hyperactivated sperm motility.Dedicated to Professor Dr. Th.H. Schiebler on the occasion of his 65^th birthdayThis study was supported by the Deutsche Forschungsgemeinschaft (grant Au 48/7-8)     

Subcellular localization of acid proteinase in barley mesophyll protoplasts

Chloroplasts prepared from lysed protoplasts of barley mesophyll contain 2–8% of the total acid proteinase activity. This residual activity is not associated with intact chloroplasts isolated by means of density gradient centrifugation. Vacuoles isolated from lysed protoplasts contain 80–85% of the total acid proteinase activity, indicating that the enzyme(s) which is presumably responsible for the degradation of chloroplastic proteins is located largely in the central vacuoles of mesophyll cells.     

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.     

Transfer and fate of seminal fluid molecules in the beetle, Diaprepes abbreviatus: implications for the reproductive biology of a pest species

Molecules transferred from males to females via seminal fluids are important to the study of insect reproduction because they affect female physiology, reproductive behavior, and longevity. These molecules (seminal fluid molecules or SFMs) interest applied entomologists because of their potential use in insect control. SFMs are also interesting because of their relatively rapid evolution and important role in post-mating sexual selection. We studied SFMs in Diaprepes abbreviatus, a major pest of numerous plant species of economic importance. Using radiolabeled-methionine (35S), we found that D. abbreviatus males synthesized proteins de novo in their reproductive tissues after mating. Males that were fed radiolabeled methionine transferred radioactivity to females beginning within the first 10 min of mating. Male-derived substances are absorbed from the female's reproductive tract into the hemolymph and circulated throughout the body, but are found primarily in the eggs and ovaries. As a result, SFMs may be a useful means of both horizontal (to mates) and vertical transfer (to offspring) of control agents between conspecifics.     

Identification and partial purification of a heart mitochondrial membrane proteinase

Membrane-bound proteinase activity was demonstrated by a solid-phase assay system in both beef heart and rat liver mitochondria. The activity was sensitive to SH reagents and assorted proteinase inhibitors. Although stimulated by nonionic detergents, it became labile when solubilized by detergents. The proteinase activity from heart mitochondria copurified with the ADP:ATP translocator protein. Gel electrophoresis of this preparation revealed the translocator polypeptide as well as a number of minor components. In solubilized mitochondria the ADP:ATP translocator polypeptide slowly disappeared upon standing at 0°C as revealed by polyacrylamide gel electrophoresis under denaturing conditions. The loss of this polypeptide was prevented by addition of proteinase inhibitors as well as the translocator affinity ligand, carboxyatractylate. These observations confirm the presence of an integral membrane proteinase in mitochondria and suggest a structural and enzymatic interaction between the proteinase and the ADP:ATP translocator.Abbreviations PMSF phenylmethanesulfonyl fluoride - TPCK l-1-tosylamido-2-phenylethylchloromethyl ketone - TLCK 1-chloro-3-tosylamido-7-amino-l-2-heptanone - NEM N-ethylmaleimide - PCMBS p-chloromercuriphenylsulfonic acid - SDS sodium dodecyl sulfate - MOPS morpholinopropane sulfonate - [I₅₀] concentration of inhibitor required to give 50% inhibition     

beta-Microseminoprotein binds CRISP-3 in human seminal plasma

beta-Microseminoprotein (MSP) and cysteine-rich secretory protein 3 (CRISP-3) are abundant constituents of human seminal plasma. Immunoprecipitation and gel filtration of seminal plasma proteins combined with examination of the proteins in their pure form showed that MSP and CRISP-3 form stable, non-covalent complexes. CRISP-3 binds MSP with very high affinity, as evidenced by surface plasmon resonance. Due to far higher abundance of MSP in prostatic fluid, it manifests large overcapacity for CRISP-3 binding. Structural similarity with an MSP-binding protein from blood plasma suggests that CRISP-3 binds MSP through its aminoterminal SCP-domain.     

Cross-generational fitness benefits of mating and male seminal fluid

In many species, the physical act of mating and exposure to accessory gland proteins (Acps) in male seminal fluid reduces female survival and offspring production. It is not clear what males gain from harming their sexual partners or why females mate frequently despite being harmed. Using sterile strains of Drosophila melanogaster that differ in their production of Acps, we found that both the physical act of mating and exposure to male seminal fluid in mothers increase the fitness of daughters. We show that the changes in daughter fitness are mediated by parental effects, not by sexual selection involving good genes or owing to variation in maternal egg production. These results support the idea that male harm of females might partly evolve through cross-generational fitness benefits.     

Characterization of a latent cysteine proteinase from ascitic fluid as a high molecular weight form of cathepsin B

The latent cysteine proteinase present in ascitic fluid of patients with neoplasia and released from ascites cells in culture has been partially purified and the enzyme after pepsin activation was shown to be immunologically related to the lysosomal proteinase, cathepsin B. The latent form was characterized as a single chain of M_r 40 000 as determined by SDS-polyacrylamide gel electrophoresis under reducing conditions followed by Western blotting and immune staining with an antiserum to human cathepsin B. Using the same techniques the enzyme after pepsin activation gave a single band of M_r 33 000. Analysis by isoelectric focusing showed that the latent enzyme before and after pepsin treatment is composed of several acidic isoenzymes. These findings suggest that this latent proteinase represents a precursor form of cathepsin B which is released extracellularly rather than being processed and directed to the lysosome.     

Expression of sarcolectin in the human pituitary gland and amniotic fluid

Sarcolectin (SCL) is a 55 kDa protein cross-reacting with a cytokeratin 7 monomer found in placental blood, sarcomas and various tissues. It blocks the synthesis of interferon-dependent secondary proteins, induces cell DNA activation and sensitizes cells to viral infection. SCL is a potent promoter of tissue growth. In the present report, we demonstrate that SCL is expressed in the human pituitary gland at the mRNA and protein levels. We show also its presence in human amniotic fluid in high titres while interferon titres is weak. These results allow to postulate a potential role of SCL as a growth factor participating in human foetal development.     

Effect of Nandrolone decanoate induced-oxidative stress on rat testes,prostate, and seminal vesicle: Biochemical,morphometric and histopathological studies

Nandrolone decanoate (Nd) is a highly abused androgenic-anabolic steroid among body builders. Even though it has weak androgenic effects, its prolonged use may have harmful impact on male reproductive system which needs to be evaluated. This study aimed to reinvestigate its possible oxidative stress induced alteration on male rat reproductive system. Twenty-eight male rats were divided into two groups. Nd treated group (n = 18) injected intramuscular with 10 mg/kg body weight once a week for four weeks. While, the control group (n = 10) was injected with physiologic saline by the same route for four weeks. Body weight was recorded for all rats and after animal dissection weight of testes, prostate and seminal vesicles were also recorded. The results showed that the average testicular weight was decreased in treated group compared to the control. The average weights of the prostate and seminal vesicles were increased compared to the control. Morphometric study revealed that in Nd treated group, there was a decrease in the width of seminiferous tubules and the height of spermatogenic cell layer compared to the control. Testicular degeneration was expressed by presence of spermatid giant cells, vacuolation, and degenerated spermatozoa. Tunnel technique showed scattered positive reaction among the spermatogenic cell layers and interstitial cells. Severe alterations of the prostate were expressed by benign prostate hyperplasia and retained secretions. Lipid peroxidation products (malonaldehyde concentration as ng/g of testicular tissue) were increased in treated group compared to the control and suggested the occurrence of oxidative damage. Nd induced severe alterations in the male genital organs were resulted from oxidative stress. It is concluded that the male genital organs are highly sensitive to the anabolic steroids and there is a high extent of reproductive risk associated with the use of AASs.     

细菌性前列腺炎患者前列腺液菌群调查及耐药谱分析

目的了解细菌性前列腺炎患者菌群分布及其耐药现状,更有效地对细菌性前列腺炎进行预防控制。方法对我院2005年10月至2007年9月拟诊为细菌性前列腺炎的患者266例,取前列腺液做细菌需氧及5%CO2环境培养,以Phoenix 100型全自动细菌分析系统进行菌种鉴定及药物敏感试验,综合分析结果。结果266例患者中前列腺液培养阳性者204例,阳性率为76.69%。其中葡萄球菌152株,占检出菌株数的74.51%。检出主要细菌为：溶血葡萄球菌82株（40.2%）,表皮葡萄球菌34株（16.67%）,粪肠球菌25株（12.25%）,头状葡萄球菌10株（4.9%）,金黄色葡萄球菌5株（2.45%）。葡萄球菌对氨苄西林、青霉素耐药率达到100%,对万古霉素、呋喃妥因、吗啉噁酮敏感率分别为100%、95.9%、95.1%。结论细菌性前列腺炎患者多由条件致病菌感染所致,尤其以凝固酶阴性葡萄球菌感染多见,患者年龄呈逐渐年轻化趋势;万古霉素、呋喃妥因、吗啉噁酮对葡萄球菌高度敏感。     

Evaluation of seminal plasma HSPA2 protein as a biomarker of human spermatogenesis status

In mammals, testicular Heat shock-related 70 kDa protein 2 (HSPA2) is a chaperon strictly linked to spermatogenesis status, whereas its presence in spermatozoa ensures successful oocyte fertilization. However, there is little information on this protein in seminal plasma in infertile males. Based on our previous two independent studies, we have selected HSPA2 to evaluate this seminal plasma protein is a potential biomarker of correct spermatogenesis.Using immunoblotting and mass spectrometry (MS) we have screened human seminal plasma samples for the presence of HSPA2. Samples were obtained from individuals with normozoospermia, cryptozoospermia, non-obstructive and obstructive azoospermia.Our results showed a lack of HSPA2 in seminal plasma in all azoospermic males however, in cryptozoospermia the results were extremely diversified. Additionally, the application of 2-dimensional gel electrophoresis (2-DE) indicated the presence of additional protein isoforms suggesting possible mechanisms underlying the male infertility.Our findings suggest seminal plasma HSPA2 protein as a possible biomarker not only of spermatogenesis status, especially in cryptozoospermic males, but also as a biomarker predicting the success of reproductive treatment including assisted reproductive techniques (ART).     

Sperm survival in female stalk-eyed flies depends on seminal fluid and meiotic drive

Sperm competition is common in many insect species; however, the mechanisms underlying differences in sperm precedence are not well understood. In the stalk-eyed fly, Cyrtodiopsis whitei (Diptera, Diopsidae), sperm precedence is influenced by the presence of sex chromosome meiotic drive. When drive-carrying males compete with non-driving males for fertilizations within a female, the number of progeny sired by drive males is significantly fewer than predicted by sperm mixing alone. Thus, drive males apparently suffer not only a reduction in the number of viable sperm produced, but also a reduction in sperm competitive ability. In this study, we manipulated the amount and source of seminal fluid and sperm received by females by interrupting copulations before sperm, but after seminal fluid, was transferred. We find that seminal fluid from another male influences the number of progeny sired by a drive-carrying male when both males mate with the same female. Sperm viability staining reveals that sperm from drive males are incapacitated by seminal fluid from other males within the female reproductive tract. These results suggest that multiple mating by females enables seminal fluid products to interact differentially with sperm and may reduce the transmission advantage of the drive chromosome.     

Ascaris suum: electrophoretic characterization of reproductive tract and perienteric fluid polypeptides, and effects of seminal and uterine fluids on spermiogenesis.

Fluids isolated from the testis, seminal vesicle, uterus, and pseudocoelomic cavity of Ascaris suum were characterized using sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and measured for protein concentration, pH, and osmolarity. The testis and seminal fluids display much homology and share major polypeptide components having molecular weights of 15,000 and 35,000. A cytoplasmic extract of spermatids from the seminal vesicle exhibited a banding pattern nearly identical to that of testis fluid. The seminal fluid has unique major components of 57,000 and 150,000, and seminal fluid from individual worms showed differences in major band concentration and distribution of minor components. The uterine fluid has major polypeptides of 14,000, 16,000, 66,000, 74,000, 120,000, and 140,000, and exhibits more similarity to the perienteric fluid then either the seminal or testis fluids. Electrophoretic comparisons of four uterine regions revealed nearly identical banding patterns although somewhat higher concentrations of four major components occurred in certain segments. The male and female perienteric fluids have major bands at 40,000, 120,000, and 140,000, and the female fluid has more intense minor components of 90,000 and 115,000. Perienteric fluid from individual worms differed only in minor band distribution. The reproductive fluids have numerous minor components mostly from 20,000 to 70,000, while the perienteric fluid minor bands are mainly located in the 80,000 to 120,000 range. The pH of the seminal fluid (6.5) differs from that of the uterine fluid (7.7), and both seminal and uterine fluids are of lower osmolarity than the perienteric fluid. In vitro studies demonstrate that uterine fluid does not induce spermatid transformation into bipolar, ameboid spermatozoa, while the seminal fluid induces only lipid granule coalescence in either seminal vesicle or terminal testis spermatids.