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.     

Verification of male infertility biomarkers in seminal plasma by multiplex selected reaction monitoring assay

Seminal plasma is a promising biological fluid to use for noninvasive clinical diagnostics of male reproductive system disorders. To verify a list of prospective male infertility biomarkers, we developed a multiplex selected reaction monitoring assay and measured the relative abundance of 31 proteins in 30 seminal plasma samples from normal, nonobstructive azoospermia and post-vasectomy individuals. Median levels of some proteins were decreased by more than 100-fold in nonobstructive azoospermia or post-vasectomy samples, in comparison with normal samples. To follow up the most promising candidates and measure their concentrations in seminal plasma, heavy isotope-labeled internal standards were synthesized and used to reanalyze 20 proteins in the same set of samples. Concentrations of candidate proteins in normal seminal plasma were found in the range 0.1-1000 μg/ml but were significantly decreased in nonobstructive azoospermia and post-vasectomy. These data allowed us to select, for the first time, biomarkers to discriminate between normal, nonobstructive azoospermia, and post-vasectomy (simulated obstructive azoospermia) seminal plasma samples. Some testis-specific proteins (LDHC, TEX101, and SPAG11B) performed with absolute or nearly absolute specificities and sensitivities. Cell-specific classification of protein expression indicated that Sertoli or germ cell dysfunction, but not Leydig cell dysfunction, was observed in nonobstructive azoospermia seminal plasma. The proposed panel of biomarkers, pending further validation, could lead to a clinical assay that can eliminate the need for testicular biopsy to diagnose the category of male infertility, thus providing significant benefits to patients as well as decreased costs associated with the differential diagnosis of azoospermia.     

Analysis of seminal plasma from patients with non-obstructive azoospermia and identification of candidate biomarkers of male infertility

Infertility affects approximately 15% of couples with equivalent male and female contribution. Absence of sperm in semen, referred to as azoospermia, accounts for 5-20% of male infertility cases and can result from pretesticular azoospermia, non-obstructive azoospermia (NOA), and obstructive azoospermia (OA). The current clinical methods of differentiating NOA cases from OA ones are indeterminate and often require surgical intervention for a conclusive diagnosis. We catalogued 2048 proteins in seminal plasma from men presented with NOA. Using spectral-counting, we compared the NOA proteome to our previously published proteomes of fertile control men and postvasectomy (PV) men and identified proteins at differential abundance levels among these clinical groups. To verify spectral counting ratios for candidate proteins, extracted ion current (XIC) intensities were also used to calculate abundance ratios. The Pearson correlation coefficient between spectral counting and XIC ratios for the Control-NOA and NOA-PV data sets is 0.83 and 0.80, respectively. Proteins that showed inconsistent spectral counting and XIC ratios were removed from analysis. There are 34 proteins elevated in Control relative to NOA, 18 decreased in Control relative to NOA, 59 elevated in NOA relative to PV, and 16 decreased in NOA relative to PV. Many of these proteins have expression in the testis and the epididymis and are linked to fertility. Some of these proteins may be useful as noninvasive biomarkers in discriminating NOA cases from OA.     

Functions and analysis of the seminal fluid proteins of male Drosophila melanogaster fruit flies

The study of insect seminal fluid proteins provides a unique window upon adaptive evolution in action. The seminal fluid of Drosophila melanogaster contains over 80 proteins and peptides, which are transferred together with sperm by mating males. The functions of many of these substances are not yet known. However, those that have been characterized have marked effects on the reproductive success of males and females. For example, seminal fluid proteins and peptides can decrease female receptivity, can increase egg production and can increase sperm storage, and are necessary for sperm transfer and success in sperm competition. In this review we focus on the currently known functions of seminal fluid molecules and on new technologies and approaches that are enabling novel questions about their form and function to be addressed. We discuss how techniques for disrupting the production of seminal fluid proteins, such as homologous recombination and RNA interference, along with the use of microarrays and yeast two hybrid systems, should allow us to address ever more sophisticated questions about seminal fluid protein function. These and similar techniques promise to reveal the function of naturally-occurring variants of these proteins and hence the evolutionary significance of genetic variation for them.     

Immunoreactive vasopressin in bull seminal fluid

Immunoreactive arginine vasopressin (irAVP) was measured in seminal fluid with and without extraction using a specific radioimmunoassay (RIA). A large fraction of irAVP was removed after extraction on octadecasilylsilica cartridges. The measured amount of irAVP corresponded to the levels found in blood plasma. Dilutions of seminal plasma extracts were parallel with the RIA standard curve. On reversed phase HPLC the extracted material coeluted with synthetic AVP. These findings suggest an identity of this immunoreactive material with intact AVP. During incubations of synthetic AVP and its analogue 8-D-arginine vasopressing (8-DAVP) in seminal plasma, immunoreactivity decreased considerably with the former peptide, while the concentration of 8-DAVP was not significantly altered.     

Cytokines and male infertility

Many male infertility cases have no apparent cause, being characterized as idiopathic. Both inflammation and obesity have long been associated with infertility. On one hand, inflammation, such as orchitis and male accessory gland infections (MAGIs), are regulated by inflammatory cytokines. The latter are also produced in the testis by Leydig and Sertoli cells, being associated with gap junctional communication at the blood–testis barrier. Furthermore, they regulate spermatogenesis through cell interaction, Toll-like receptors and production of reactive oxygen species. Additionally, they affect testosterone production, acting at many levels of the pituitary - gonadal axis. Any imbalance in their production may result in infertility. On the other hand, obesity has also been associated with infertility. Adipokines, cytokines produced by white adipose tissue, regulate the lipid and glucose metabolism and the inflammatory system. Recent data on leptin show that it regulates reproduction by adjusting hypothalamus - pituitary - gonadal axis at both the central and peripheral levels. In this regard, resistin, visfatin and the GH secretagogue peptic hormone ghrelin affect spermatogenesis, whereas data on adiponectin are rather scarce. In conclusion, inflammatory cytokines and adipokines seem to have a pivotal role in the regulation of spermatogenesis; any imbalance in this stable environment may lead to infertility. Nevertheless, further studies are needed to clarify their exact role.     

Targeted deletion of the epididymal receptor HE6 results in fluid dysregulation and male infertility

Human epididymal protein 6 (HE6; also known as GPR64) is an orphan member of the LNB-7TM (B(2)) subfamily of G-protein-coupled receptors. Family members are characterized by the dual presence of a secretin-like (type II) seven-transmembrane (7TM) domain and a long cell adhesion-like extracellular domain. HE6 is specifically expressed within the efferent ductules and the initial segment of the epididymis, ductal systems involved in spermatozoon maturation. Here, we report that targeted deletion of the 7TM domain of the murine HE6 gene results in male infertility. Mutant mice reveal a dysregulation of fluid reabsorbtion within the efferent ductules, leading to a backup of fluid accumulation in the testis and a subsequent stasis of spermatozoa within the efferent ducts. The fertility phenotype of HE6 knockout mice identifies this receptor as a potential nonsteroidal, nontesticular target for future male contraceptives and identifies an in vivo function for a member of this unusual gene family.     

Proteomic analysis of Drosophila mojavensis male accessory glands suggests novel classes of seminal fluid proteins

Fruit-flies of the genus Drosophila are characterized by overwhelming variation in fertilization traits such as copulatory plug formation, sperm storage organ use, and nutritional ejaculatory donation. Despite extensive research on the genetic model Drosophila melanogaster, little is known about the molecular underpinnings of these interspecific differences. This study employs a proteomic approach to pin-point candidate seminal fluid proteins in Drosophila mojavensis, a cactophilic fruit-fly that exhibits divergent reproductive biology when compared to D. melanogaster. We identify several classes of candidate seminal fluid proteins not previously documented in the D. melanogaster male ejaculate, including metabolic enzymes, nutrient transport proteins, and clotting factors. Conversely, we also define 29 SFPs that are conserved despite >40 million years of Drosophila evolution. We discuss our results in terms of universal processes in insect reproduction, as well as the specialized reproductive biology of D. mojavensis.     

Quantification of prostaglandins in human seminal fluid

A method is described for measurement of the prostaglandins present in human seminal plasma. The PGEs and the 19-hydroxy-PGEs are converted to the respective PGB-compounds. They are determined with UV-absorbance after purification with two chromatography steps. The PGFs and the 19-hydroxy-PGFs are determines by gas chromatography, which also allows measurement of the 8β-isomers. Recovery of added PG was in the average 99.2% (range 89.9–107.7). Mean variation between duplicate analyses was 5.6% (range 0.9–9.1). The method has been simplified to allow at least limited clinical use.     

Haemolytic factor in bovine seminal vesicle fluid

The haemolytic factor of bovine seminal fluid has a protein nature; it is present in the precipitate obtained by treating the fluid with alcohol-ether, ammonium sulphate and rivanol; it occurs in three out of five protein fractions obtained by chromatographic separation on Sephadex G 100.
of Enzymatic treatment of seminal vesicle fluid by pronase caused total inactivation of the haemolytic factor. Chymotrypsin caused considerable damage, while a number other enzymes did not affect the activity of the haemolytic factor.     

Chromosomal studies of male infertility

Prometaphase and metaphase chromosome analyses performed on 70 consecutive men with primary infertility (for a period of at least 2 years) revealed 8 (11.42%) men with some kind of chromosomal abnormality. The highest frequency of abnormal karyotypes (10%) was found among patients with azpospennia and the most frequent anomaly was 47, XXY chromosomal constitution, found in 6 (8.57%) patients. All the chromosomal aberrations found in this study was sex chromosomal type and we did not find any autosomal aberration. All patients with numerical chromosomal anomalies had azoospermia. The incidence of structural aberration in our study was 1.42%. Fifteen patients had different chromosomal variants (21.38%). We suggest that men with azoospermia should be considered for cytogenetic investigation and we report that "variants of the Y chromosome" have no influence on the sperm count (million/ml) and fertility of men.