Postnatal cadmium administration affects the presence and distribution of carbohydrates in the sperm membrane during maturation in the epididymis in adult Wistar rats.

Cadmium (Cd) is a heavy metal related to a decrease in sperm parameters. The transit of spermatozoa through the epididymis is necessary to generate changes in the sperm membrane, such as the assembly of various carbohydrates that are added to the spermatazoan's surface to prepare it for successful fertilisation of the oocyte. No studies have yet analysed whether Cd alters the presence and distribution of these carbohydrates. We aimed to evaluate the changes induced by Cd in the distribution pattern of N-acetylglucosamine, sialic acid, mannose and fucose on the sperm membrane in the epididymis (e.g. caput, corpus, cauda) and if it alters the epididymal epithelium. Male Wistar pups were treated with Cd doses (0.125, 0.25 and 0.5mg/kg) on postnatal days 1-49. At postnatal day 90, they were humanely killed, sperm samples were obtained from the epididymis and tissue samples were taken for histological analysis. Cd concentrations in the blood and epididymis increased in proportion to the dose administered and decreased the serum testosterone levels and sperm quality. Histological analysis revealed alterations in the epithelium in all Cd-treated groups. Cd altered the distribution patterns of carbohydrates and fluorescence indices. All these alterations affected the structure and functioning of sperm.

Modification of Crocodile Spermatozoa Refutes the Tenet That Post-testicular Sperm Maturation Is Restricted To Mammals.

Competition to achieve paternity has contributed to the development of a multitude of elaborate male reproductive strategies. In one of the most well-studied examples, the spermatozoa of all mammalian species must undergo a series of physiological changes, termed capacitation, in the female reproductive tract before realizing their potential to fertilize an ovum. However, the evolutionary origin and adaptive advantage afforded by capacitation remains obscure. Here, we report the use of comparative and quantitative proteomics to explore the biological significance of capacitation in an ancient reptilian species, the Australian saltwater crocodile (Crocodylus porosus,). Our data reveal that exposure of crocodile spermatozoa to capacitation stimuli elicits a cascade of physiological responses that are analogous to those implicated in the functional activation of their mammalian counterparts. Indeed, among a total of 1119 proteins identified in this study, we detected 126 that were differentially phosphorylated (± 1.2 fold-change) in capacitated versus, noncapacitated crocodile spermatozoa. Notably, this subset of phosphorylated proteins shared substantial evolutionary overlap with those documented in mammalian spermatozoa, and included key elements of signal transduction, metabolic and cellular remodeling pathways. Unlike mammalian sperm, however, we noted a distinct bias for differential phosphorylation of serine (as opposed to tyrosine) residues, with this amino acid featuring as the target for ∼80% of all changes detected in capacitated spermatozoa. Overall, these results indicate that the phenomenon of sperm capacitation is unlikely to be restricted to mammals and provide a framework for understanding the molecular changes in sperm physiology necessary for fertilization.

Low doses of Bisphenol S affect post-translational modifications of sperm proteins in male mice.

BACKGROUND: Bisphenol S (BPS) is increasingly used as a replacement for bisphenol A in the manufacture of products containing polycarbonates and epoxy resins. However, further studies of BPS exposure are needed for the assessment of health risks to humans. In this study we assessed the potential harmfulness of low-dose BPS on reproduction in male mice. METHODS: To simulate human exposure under experimental conditions, 8-week-old outbred ICR male mice received 8 weeks of drinking water containing a broad range of BPS doses [0.001, 1.0, or 100 µg/kg body weight (bw)/day, BPS1-3] or vehicle control. Mice were sacrificed and testicular tissue taken for histological analysis and protein identification by nano-liquid chromatography/mass spectrometry (MS) and sperm collected for immunodetection of acetylated lysine and phosphorylated tyrosine followed by protein characterisation using matrix-assisted laser desorption ionisation time-of-flight MS (MALDI-TOF MS). RESULTS: The results indicate that compared to vehicle, 100 µg/kg/day exposure (BPS3) leads to 1) significant histopathology in testicular tissue; and, 2) higher levels of the histone protein γH2AX, a reliable marker of DNA damage. There were fewer mature spermatozoa in the germ layer in the experimental group treated with 1 µg/kg bw (BPS2). Finally, western blot and MALDI-TOF MS studies showed significant alterations in the sperm acetylome and phosphorylome in mice treated with the lowest exposure (0.001 µg/kg/day; BPS1), although the dose is several times lower than what has been published so far. CONCLUSIONS: In summary, this range of qualitative and quantitative findings in young male mice raise the possibility that very low doses of BPS may impair mammalian reproduction through epigenetic modifications of sperm proteins.

Bisphenol A affects the maturation and fertilization competence of spermatozoa.

Although there are numerous studies on bisphenol A (BPA) on the testis and spermatozoa, the effect of BPA on the physiological link between the testis and maturation of spermatozoa has not been studied. To provide an optimal environment (acidic pH) for sperm maturation in the epididymis, clear cells secrete protons and principal cells reabsorb bicarbonate and the secreted proton. Because of its crucial role in sperm maturation and fertility, functional changes in the epididymis following BPA exposure must be considered to fully understand the mechanisms of BPA on male fertility. Here, we identified the adverse effects of BPA exposure during puberty in male mice. CD-1 male mice were gavaged daily with vehicle (corn oil) and 50 mg BPA/kg-BW for 6 weeks. We determined the changes in epididymis, functional sperm parameters including motility, capacitation status, tyrosine phosphorylation, and fertility-related protein expression and in vitro and in vivo fertility rate following BPA exposure. Expression of vacuolar-type H + -ATPase is necessary for the secretion of protons by clear cells of the caput epididymis and was directly down-regulated following BPA exposure, while there were no changes in the other epithelial cell types in the epididymis. Also, pERK 1/2 signaling pathway was increased significantly in the caput epididymis following BPA exposure. Consequently, the luminal pH slightly increased, resulting in premature capacitation of spermatozoa. Moreover, there was a significant loss of the acrosomal membrane following an increase of protein tyrosine phosphorylation, while PKA activity decreased during sperm capacitation. Fertility-related proteins also showed aberrant expression upon BPA exposure. These modifications resulted in decreased male fertility in vitro and in vivo.

Double-blind, randomised, placebo-controlled trial on the effect of L-carnitine and L-acetylcarnitine on sperm parameters in men with idiopathic oligoasthenozoospermia.

Carnitine is essential for energy metabolism and spermatozoa maturation. Combining L-carnitine and L-acetylcarnitine with micronutrients has been investigated as a treatment for infertility in men. We evaluated the effects of a therapeutic formulation, Proxeed Plus, on sperm parameters in oligoasthenozoospermic men. This prospective, randomised, double-blind, placebo-controlled clinical trial involved 175 males (19-44 years) with idiopathic oligoasthenozoospermia who failed to impregnate their partners (12 months). Males received Proxeed Plus or placebo for 3 and 6 months. Sperm volume, progressive motility and vitality significantly (p < 0.001) improved after 6 months compared to baseline. Sperm DNA fragmentation index significantly decreased compared to baseline (p < 0.001) and the 3-month therapy (p = 0.014) in treated men. Increased seminal carnitine and α-glucosidase concentration also positively correlated with improved progressive motility. Decreased DNA fragmentation index was the good predictor of progressive sperm motility >10%, and simultaneous measurement of changes in sperm vitality and DNA fragmentation index gave the highest probability of sperm motility 10% (AUC = 0.924; 95% CI = 0.852-0.996; p < 0.001). Logistic regression analyses revealed DNA fragmentation index decrease as the only independent predictor of sperm motility 10% (OR = 1.106; p = 0.034). We have demonstrated the beneficial effects of carnitine derivatives on progressive motility, vitality and sperm DNA fragmentation. Combining metabolic and micronutritive factors is beneficial for male infertility.

Influence of antifertility agents Dutasteride and Nifedipine on CatSper gene level in epididymis during sperm maturation in BALB/c mice.

Calcium (Ca2+) signaling is critical for successful fertilization. In spermatozoa, capacitation, hyperactivation of motility and the acrosome reaction are all mediated by increases in intracellular Ca2+ through CatSper (sperm-specific cation channel). The CatSper channel complex contains four pore-forming α subunits (CatSper1-4) and five accessory subunits called ß, δ, ε, γ and ζ. Genetic deletion of any of the four CatSper genes in mice results in loss of hyperactivated motility and male infertility. Despite their vital role in male fertility, almost very little is known about influence of antifertility agents on CatSper gene expression in epididymis and epididymal spermatozoa. Therefore, we performed quantitative real-time qPCR analysis for CatSper expression in the epididymis and epididymal sperm of BALB/c mice after treatment with Dutasteride (DS), a dual 5-α reductase inhibitor and Nifedipine (NF) a calcium channel blocker as positive control. We observed that treatment with antifertility agents Dutasteride and Nifedipine induced significant decreases in the caput and cauda epididymal sperm counts, motility and fertility which could partly be attributed to alteration in the normal morphology of the sperm associated with downregulation/upregulation of CatSper mRNAs in epididymis and epididymal spermatozoa of male BALB/c mice. These can be explained on the basis of interference with mechanisms affecting calcium ion signaling resulting in changes in intracellular calcium required for sperm activity, finally affecting sperm maturation and fertility of male BALB/c mice. These studies provide some novel avenues for developing new male contraceptives in future.

Long-chain fatty acid triglyceride (TG) metabolism disorder impairs male fertility: a study using adipose triglyceride lipase deficient mice.

STUDY QUESTION: Does the deletion of adipose triglyceride lipase (Atgl) gene impair male fertility? SUMMARY ANSWER: The deletion of Atgl gene impaired male fertility but the effect was partially reversed by a low long-chain triglyceride (TG) diet. WHAT IS KNOWN ALREADY: ATGL specifically hydrolyses long-chain fatty acid TG to diacylglycerol and a high level of expression of ATGL in testes has been reported. However, the role of ATGL in male fertility is unknown. STUDY DESIGN, SIZE, DURATION: To investigate the effect of deletion of Atgl gene on male fertility, cauda epididymides and testes were collected from wild-type, heterozygous and homozygous Atgl-deficient mice at 10 weeks of age and epididymal sperm analysis and histological analysis of the testes were performed. To investigate whether a medium-chain triglycerides (MCTs) replacement diet mitigated the impaired male fertility by deletion of Atgl gene, homozygous Atgl-deficient mice were fed a MCT replacement diet, or a standard diet including long-chain triglycerides (LCTs) in a control group, for 6 weeks from 5 weeks of age (n = 22). The systematic and local effects of the MCT replacement diet on spermatogenesis and sperm maturation in the epididymis were analyzed at 10 weeks of age. PARTICIPANTS/MATERIALS, SETTING, METHODS: Hematoxylin and eosin staining in paraffin-embedded sections of testes and Oil Red O staining in frozen sections of testes were performed. The epididymal sperm concentrations were analyzed. Statistical analyses were performed using the Student's t-test or Mann-Whitney U test with Shapiro-Wilk Normality test. MAIN RESULTS AND THE ROLE OF CHANCE: Although heterozygous mice were fertile and showed a similar number of epididymal total and motile sperm concentrations to wild-type mice, the deletion of Atgl gene in homozygous mice led to accumulation of TG deposits in testes and impaired spermatogenesis. The deletion of Atgl gene also impaired the sperm maturation process required for sperm to acquire the ability to move forward in the epididymis. The MCT replacement diet for 6 weeks increased the plasma level of non-esterified fatty acid (NEFA) (1.5-fold, P = 0.005), but not the plasma total cholesterol (T-Cho) and TG levels. In testes, the MCT replacement diet decreased the number of Oil Red O stain positive vacuoles (-40%, P < 0.001) and increased testis tissue weight (1.1-fold, P = 0.012), total sperm concentration (1.5-fold, P = 0.011) and motile sperm concentration (2.1-fold, P < 0.001) compared to the control group. However, there was no significant change in the sperm survival rate between the two groups. LARGE SCALE DATA: None. LIMITATIONS REASONS FOR CAUTION: One previous study reported that Atgl-deficient male mice were fertile. In most studies heterozygous Atgl(+/-) mice were used to generate homozygous Atgl-deficient Atgl(-/-) mice. Although the same gene targeting mice were used in this study and the formation of vaginal plugs were observed after mating with Atgl(-/-) male mice, there were no pregnant wild-type mice observed after mating with Atgl(-/-) male mice. WIDER IMPLICATIONS OF THE FINDINGS: Local TG metabolism in the male reproductive system could affect spermatogenesis and sperm motility in men. The MCT replacement diet could be an effective therapy for idiopathic non-obstructive oligozoospermia or asthenozoospermia in men with low levels of serum NEFA. STUDY FUNDING AND COMPETING INTEREST(S): This study was supported in part by the Japan Society for the Promotion of Science JSPS KAKENHI Grant (Nos. JP24249080, JP25462557, JP16K11086). The authors declare no conflict of interest.

Role of Ca2+ in the IVM of spermatozoa from the sterlet Acipenser ruthenus.

The role of Ca2+ in sturgeon sperm maturation and motility was investigated. Sperm from mature male sterlets (Acipenser ruthenus) were collected from the Wolffian duct and testis 24h after hormone induction. Testicular spermatozoa (TS) were incubated in Wolffian duct seminal fluid (WDSF) for 5min at 20°C and were designated 'TS after IVM' (TSM). Sperm motility was activated in media with different ion compositions, with motility parameters analysed from standard video microscopy records. To investigate the role of calcium transport in the IVM process, IVM was performed (5min at 20°C) in the presence of 2mM EGTA, 100µM Verapamil or 100µM Tetracaine. No motility was observed in the case of TS (10mM Tris, 25mM NaCl, 50mM Sucr with or without the addition of 2mM EGTA). Both incubation of TS in WDSF and supplementation of the activation medium with Ca2+ led to sperm motility. The minimal Ca2+ concentration required for motility activation of Wolffian duct spermatozoa, TS and TSM was determined (1-2nM for Wolffian duct spermatozoa and TSM; approximately 0.6mM for TS). Motility was obtained after the addition of verapamil to the incubation medium during IVM, whereas the addition of EGTA completely suppressed motility, implying Ca2+ involvement in sturgeon sperm maturation. Further studies into the roles of Ca2+ transport in sturgeon sperm maturation and motility are required.

Glycogen synthase kinase 3 regulates acrosomal exocytosis in mouse spermatozoa via dynamin phosphorylation.

The dynamin family of GTPases has been implicated as novel regulators of the acrosome reaction, a unique exocytotic event that is essential for fertilization. Dynamin activity during the acrosome reaction is accompanied by phosphorylation of key serine residues. We now tested the hypothesis that glycogen synthase kinase 3 (GSK3) is the protein kinase responsible for dynamin phosphorylation at these phosphosites in mouse spermatozoa. Pharmacologic inhibition of GSK3 in mature mouse spermatozoa (CHIR99021: IC50 = 6.7 nM) led to a significant reduction in dynamin phosphorylation (10.3% vs. 27.3%; P < 0.001), acrosomal exocytosis (9.7% vs. 25.7%; P < 0.01), and in vitro fertilization (53% vs. 100%; P < 0.01). GSK3 was shown to be present in developing germ cells where it colocalized with dynamin in the peri-acrosomal domain. However, additional GSK3 was acquired by maturing mouse spermatozoa within the male reproductive tract, via a novel mechanism involving direct interaction of sperm heads with extracellular structures known as epididymal dense bodies. These data reveal a novel mode for the cellular acquisition of a protein kinase and identify a key role for GSK3 in the regulation of sperm maturation and acrosomal exocytosis.

Implication of the estrogen receptors GPER, ESR1, ESR2 in post-testicular maturations of equine spermatozoa.

Estrogen receptors ESR1, ESR2 and GPER are present on mature ejaculated horse spermatozoa, suggesting these cells as putative targets for estrogens. Indeed, spermatozoa are exposed to high level of estrogens during the transit in the male and female genital tracts but their roles are not investigated. So, we evaluated in vitro the role of 17ß-estradiol during post-testicular maturations: regulation of motility, capacitation and acrosome reaction. Moreover according to the pseudo-seasonal breeder status of the stallion, we analyzed the putative seasonal variations in the presence of ESRs in spermatozoa. We showed that ESRs are more present on stallion sperm during the breeding season. We showed that capacitation and acrosome reaction are independent of estradiol action in horse. Estradiol can weakly modulate the motility and this effect is strictly associated with GPER and not with ESR1 and ESR2. The subcellular localization of GPER in the neck on stallion sperm is coherent with this effect. It seems that estrogens are not major regulators of sperm maturations associated to mare genital tract, so they could act during the epididymal maturations.

[IMMUNOCYTOCHEMICAL ANALYSIS OF THE DISTURBANCES IN THE STRUCTURE OF SYNAPTONEMAL COMPLEXES IN SPERMATOCYTE NUCLEI IN MICE UNDER EXPOSURE TO ROCKET FUEL COMPONENT].

There was performed an assessment of genotoxic effects of rocket fuel component--unsymmetrical dimethylhydrazine (UDMH, heptyl)--on forming germ cells of male mice. Immunocytochemically there was studied the structure of meiotic nuclei at different times after the intraperitoneal administration of UDMH to male mice. There were revealed following types of disturbances of the structure of synaptonemal complexes (SCs) of meiotic chromosomes: single and multiple fragments of SCs associations of autosomes with a sex bivalent, atypical structure of the SCs with a frequency higher than the reference level. In addition, there were found the premature desinapsis of sex bivalents, the disorder offormation of the genital corpuscle and ring SCs. Established disorders in SCs of spermatocytes, analyzed at 38th day after the 10-days intoxication of animal by the component of rocket fuel, attest to the risk of permanent persistence of chromosomal abnormalities occurring in the pool of stem cells.

Retinoic acid induces multiple hallmarks of the prospermatogonia-to-spermatogonia transition in the neonatal mouse.

In mammals, most neonatal male germ cells (prospermatogonia) are quiescent and located in the center of the testis cords. In response to an unknown signal, prospermatogonia transition into spermatogonia, reenter the cell cycle, divide, and move to the periphery of the testis cords. In mice, these events occur by 3-4 days postpartum (dpp), which temporally coincides with the onset of retinoic acid (RA) signaling in the neonatal testis. RA has a pivotal role in initiating germ cell entry into meiosis in both sexes, yet little is known about the mechanisms and about cellular changes downstream of RA signaling. We examined the role of RA in mediating the prospermatogonia-to-spermatogonia transition in vivo and found 24 h of precocious RA exposure-induced germ cell changes mimicking those that occur during the endogenous transition at 3-4 dpp. These changes included: 1) spermatogonia proliferation; 2) maturation of cellular organelles; and 3), expression of markers characteristic of differentiating spermatogonia. We found that germ cell exposure to RA did not lead to cellular loss from apoptosis but rather resulted in a delay of ∼2 days in their entry into meiosis. Taken together, our results indicate that exogenous RA induces multiple hallmarks of the transition of prospermatogonia to spermatogonia prior to their entry into meiosis.

Most regions of mouse epididymis are able to phagocytose immature germ cells.

The role of the epididymis as a quality control organ in preventing infertile gametes entering the ejaculate has been extensively explored, and it has been suggested that a specific region of mammalian epididymis is able to phagocytose abnormal germ cells. This study examines whether the epithelium of certain zones of the mouse epididymis can act as a selection barrier by removing immature germ cells from the lumen by phagocytosis. To detect the presence of immature germ cells in the epididymis, we generated transgenic mice expressing enhanced green fluorescent protein under the deleted in azoospermia-like (mDazl) promoter to easily identify immature germ cells under fluorescence microscopy. Using this technique, we observed that during the first stage of spermatogenesis in prepuberal mice, a wave of immature germ cells is released into the epididymis and that the immature epididymis is not able to react to this abnormal situation. By contrast, when immature germ cells were artificially released into the epididymis in adult mice, a phagocytic response was observed. Phagosomes appeared inside principal cells of the epididymal epithelium and were observed to contain immature germ cells at different degradation stages in different zones of the epididymis, following the main wave of immature germ cells. In this paper, we describe how the epididymal epithelium controls sperm quality by clearing immature germ cells in response to their artificially induced massive shedding into the epididymal lumen. Our observations indicate that this phenomenon is not restricted to a given epididymis region and that phagocytic capacity is gradually acquired during epididymal development.

Effect of abamectin exposure on semen parameters indicative of reduced sperm maturity: a study on farmworkers in Antalya (Turkey).

Environmental exposure to pesticides may cause serious health risks including fertility and reproductive function. The aim of this study was to highlight whether there is a relationship between exposure to abamectin and male fertility parameters of farmworkers. Twenty male farmworkers who were using abamectin and 20 men not exposed to pesticides were recruited as experimental and control groups, respectively. Semen analysis, molecular markers of sperm maturity and serum reproductive hormone levels were evaluated. In experimental group, high plasma abamectin levels were detected. These men have decreased sperm motility. Moreover, diminished molecular markers of sperm maturity, such as decreased hyaluronic acid (HA) binding of sperm, increased numbers of aniline blue positive sperm and increased percentage of creatine kinase (CK) positive sperm, were observed in abamectin-exposed men. Their serum testosterone, LH and FSH levels did not change significantly. We conclude that exposure to abamectin may impair male fertility by effecting semen quality.

Molecular perspective concerning fluoride and arsenic mediated disorders on epididymal maturation of spermatozoa: A concise review.

Epididymis is a complex tubular structure of male reproductive system where spermatozoa undergo maturation and gain the fertilizing ability. Epididymal pseudostratified columnar epithelium with different cell types play imperative role by their secretory properties and enrich the luminal microenvironment necessary for achieving spermatozoal motility. During epididymal transit several secretory proteins like P26h, SPAG11, HSPD1 and many others are deposited on spermatozoal surface. At the same time spermatozoal proteins are also modified in this intraluminal milieu, which include cyritestin, fertilin, CE9 and others. Natural and anthropogenic activities disclose various environmental pollutants which affect different physiological systems of animals and human being. Likewise, reproductive system is also being affected. Fluoride causes structural alterations of caput and cauda segments of epididymis. Redox homeostasis and functional integrity are also altered due to diminished activities of SOD1, GR, Crisp2, Lrp2 and other important proteins. On the contrary arsenic affects mostly on cauda segment. Redox imbalance and functional amendment in epididymis have been observed with arsenic revelation as evidenced by altered genomic appearance of SOD, GST, catalase, Ddx3Y, VEGF and VEGFR2. This review is dealt with structure-function interplay in normal epididymal spermatozoal maturation along with subsequent complications developed under fluoride and arsenic toxicities.

Endoplasmic reticulum protein 29 (ERp29), a protein related to sperm maturation is involved in sperm-oocyte fusion in mouse.

BACKGROUND: Sperm-oocyte fusion is a critical step in fertilization, which requires a series of proteins from both spermatozoa and oocyte to mediate membrane adhesion and subsequent fusion. A rat spermatozoa membrane protein is endoplasmic reticulum protein 29 (ERp29), which significantly increases on the sperm surface as well as in the cytoplasm of epididymal epithelia from caput to cauda as the sperm undergo epididymal maturation. Moreover, ERp29 facilitates viral infection via mediating membrane penetration. We determined if in addition to promoting sperm maturation ERp29 may also play a role in facilitating gamete fusion during the fertilization process. METHODS: Laser scanning confocal microscopy (LSCM) and Western blot analysis were employed to probe for ERp29 protein in BALB/c mouse epididymal and acrosome-reacted spermatozoa. We prepared rabbit polyclonal antibodies against mouse recombinant ERp29 (rERp29) to characterize: 1) fertilization rate (FR); 2) fertilization index (FI); 3) sperm motility and 4) acrosome reaction (AR). RESULTS: Confocal microscopy indicated that ERp29 was partially localized at the sperm head of the epididymal caput as well as over the whole head and part of the principal piece of the tail region from the epididymal cauda. However, when the acrosome reacted, ERp29 remained in the equatorial and post-acrosomal regions of the sperm head, which is the initial site of sperm-oocyte membrane fusion. Such localization changes were confirmed based on the results of Western blot analysis. Furthermore, the antibodies against mouse rERp29 inhibited the spermatozoa from penetrating into the zona pellucida (ZP)-free oocytes. The functional blocking antibodies reduced both mouse sperm-oocyte FR and FI at concentrations of 100 and 200 micro g/ml compared with pre-immunized rabbit IgG or with anti-mouse recombinant bactericidal/permeability-increasing protein (BPI, a sperm surface protein unrelated to sperm-oocyte fusion) antibodies (100 micro g/ml), but they had no effect on sperm motility and AR. CONCLUSION: This study demonstrates that ERp29 on mouse spermatozoa membrane changes during epididymal transit and AR. Accordingly, in mice this protein may be one of the important factors involved in sperm fertilization by facilitating sperm-oocyte membrane fusion.

Impact of experimental diabetes and insulin replacement on epididymal secretory products and sperm maturation in albino rats.

The present study is aimed to explore the impact of experimental diabetes and insulin replacement on epididymal secretory products, sperm count, motility, and fertilizing ability in albino rats. Prepubertal and adult male Wistar strain rats were made diabetic with a single intraperitoneal injection of streptozotocin (STZ), at 120 and 65 mg/kg body weight for prepubertal and adult rats, respectively. After 3 days of STZ administration, insulin was given to a group of diabetic rats at a dose of 3 U/100 g body weight, subcutaneously and killed after 20 days of treatment. STZ-diabetes significantly reduced the epididymal tissue concentrations of testosterone, androgen-binding protein, sialic acid, glycerylphosphoryl choline, and carnitine, suggesting its adverse effects on the secretory activity and concentrating capacity of epididymal epithelium. Impaired cauda epididymidal sperm motility and fertility (in vivo) of STZ-diabetic rats imply the defective sperm maturation. Insulin replacement prevented these changes either partially or completely. From the above findings, it is evident that STZ-diabetes has an adverse effect on sperm maturation, which may be due to the decrease in the bioavailability of testosterone and epididymal secretory products.

Hypotonic resistance of boar spermatozoa: sperm subpopulations and relationship with epididymal maturation and fertility.

Hypotonic resistance of boar spermatozoa was investigated by measuring the ratio of live/dead spermatozoa (SYBR-14/propidium iodide) by flow cytometry after hypotonic stress. The survival rate of ejaculated spermatozoa incubated in hypotonic solutions ranging from 3 to 330 mmol/kg followed a sigmoid curve that fitted a simple logistic model. The critical osmolality value (Osm(crit)) at which 50% of spermatozoa died was determined with this model. Hypotonic resistance of spermatozoa increased with temperature between 15 and 39 degrees C and decreased after hydrogen superoxide treatment, but was not modified during 8 days of preservation in Beltsville thawing solution. Hypotonic resistance markedly decreased during epididymal maturation and after ejaculation as Osm(crit) at 15 degrees C was 54.7+/-3.2, 68.5+/-10.6, 116.7+/-2.1 and 194.3+/-3.7 mmol/kg for the caput, corpus, cauda and ejaculated spermatozoa respectively. Hypo-osmotic stress of 100 mmol/kg revealed a sperm subpopulation exhibiting increased hypotonic resistance compared with the whole ejaculate (Osm(crit)=67.8+/-2.1 mmol/kg). Consistent differences were observed between lean and standard breeds (Pietrain versus Large White) and between boars within the same breed. According to data collected by artificial insemination centers during a large-scale field trial, hypotonic resistance of ejaculates was found to be positively correlated with in vivo fertility.

Prenatal exposure to betamethasone causes intergenerational impairment of epididymal development in the rat.

BACKGROUND: Studies on epididymal toxicology are scarce. Betamethasone (BM) is a glucocorticoid used in clinical practice for antenatal therapy. We previously reported changes to testicular morphology, altered sperm quality, and fertility in adult rats following intrauterine administration of BM. OBJECTIVES: Given that high levels of corticosteroids during gestation lead to fetal androgen depletion, and the essential role of testosterone during epididymal development, here we investigated epididymal morphology and physiology in the F1 and F2 male offspring of female rats treated with BM during gestation. MATERIALS AND METHODS: Pregnant rats were randomly divided into two experimental groups: control (saline vehicle, n = 11) and BM-treated group (0.1 mg/kg betamethasone 21-phosphate disodium, n = 13). Rats received an intramuscular injection of vehicle or BM on gestational days 12, 13, 18, and 19. This encompasses the beginning of the critical window of male rat reproductive tract development. A subset of three males from each litter (n = 5 litters/group) was used: One rat per litter was euthanized at puberty, one was euthanized at adulthood, while the others were mated with a non-treated female to obtain the F2 generation. The same protocol described for the F1 was applied for F2, except for the mating protocol. RESULTS: In both F1 and F2 generations, prenatal BM exposure resulted in delayed differentiation of the cauda epididymal epithelium, characterized by increased cribriform appearance on PND 45, and displayed weaker or non-detectable Cx43 immunostaining. Furthermore, in the F1 generation only, immunostaining of TP63, a transcription factor expressed in basal cells, appeared more intense with a greater number of TP63-positive cells observed in the cauda epididymis. In adults, the epithelial area was reduced in the F1 BM rats. The contractile activity of isolated epididymal ducts was comparable between groups. DISCUSSION AND CONCLUSION: Prenatal BM exposure leads to intergenerational impairment in the development and structure of the rat epididymis.

Korean red ginseng improves testicular ineffectiveness in aging rats by modulating spermatogenesis-related molecules.

Korean red ginseng (Panax ginseng Meyer) is known to rejuvenate testicular effectiveness and the sperm maturation process by regulating redox proteins in aged rats. This study was performed to investigate the effect of Korean red ginseng water extract (KRG-WE) on the expression level of spermatogenesis-related key biomolecules and sex hormone receptors as well as enzymes regulating oxidation, histone deacetylation, and growth-related activities in aged rat testis. KRG-WE (200mg/kg) mixed with a regular pellet diet was administered to 12-month-old rats for 6months (KRG-AC), whereas the young (YC, 2months) and aged (AC, 12months) controls received the vehicle only. The results showed that the expression levels of spermatogenesis-related key biomolecules (inhibin-α, nectin-2, and cyclic adenosine monophosphate [cAMP] responsive element binding protein [CREB]-1), sex hormone receptors (androgen, luteinizing- and follicle-stimulating hormone receptors [AR, LHR, and FSHR, respectively]), and antioxidant enzymes (glutathione S-transferase mu [GSTm]-5, glutathione peroxidase [GPx]-4, peroxiredoxin [PRx]-3), as well as histone deactylation (silent mating type information regulation 2 homolog 1, SIRT1) and growth-related (mammalian target of rapamycin complex 1, mTORC1) molecules were significantly altered in the AC group rat testes compared with those of the YC group. However, KRG-WE treatment of the AC group significantly (p<0.05) attenuated these molecular changes. From these results, it can be concluded that long-term administration of KRG-WE significantly delayed the aging-induced testicular dysfunction.