Ejaculated Spermatozoa Research Articles

A new insight into male fertility preservation for patients with completely immotile spermatozoa

BackgroundSperm cryopreservation is the most effective method to preserve male fertility but this is normally used for motile spermatozoa. Thus, only motile spermatozoa are used for cryopreservation in most reproductive medicine centers worldwide. The immotile spermatozoa from some problematic patients are usually discarded, resulting in a missed opportunity of sterility cryopreservation for future assisted reproductive treatments. Many studies have shown that successful fertilization can be obtained after selection of viable sperm from the completely immotile spermatozoa before ICSI. Whether the completely immotile spermatozoa are worth of freezing has not been realized The aim of this study is to explore the clinical value of cryopreservation of immotile spermatozoa.MethodsCompletely immotile spermatozoa were collected and frozen, and subsequently viable but immotile frozen-thawed spermatozoa were selected by laser plus for ICSI. Main outcomes included spermatozoa survival index, fertilization rate and good quality embryo rate.ResultsAfter identification by laser, the fresh samples of spermatozoa presented with a mean survival rate of 54.86% and 26.05%, and this was reduced to 44.13% and 18.13% in frozen-thawed spermatozoa samples, which showed a frozen-thawed spermatozoa survival index of 0.80 and 0.70 in the testicular and ejaculate sperm, respectively. There were no statistically differences in fertilization rate (80% vs80.51%, 75.00% vs 81.48%), cleavage rate (95.45% vs 98.95%, 100.00% vs 95.45%) and good quality embryo rate (40.48% vs 52.13%, 33.33%vs38.10%) between the frozen-thawed immotile spermatozoa group and the routine fresh immotile spermatozoa ICSI group in both testicular and ejaculate sperm, respectively.ConclusionsThe results of the study show that completely immotile spermatozoa can be frozen in order to preserve male fertility as long as viable spermatozoa are present. This procedure provides a further possibility for fertility preservation for patients with completely immotile spermatozoa.

Effects of GnRHR polymorphisms on sperm quality in Chinese water buffalo

Gonadotropin-releasing hormone receptor (GnRHR) plays a critical physiological role in animal reproduction and is a potential marker for improving sperm quality. In the present study, eight SNPs (g.539T>C, g.640A>G, g.655T>C, g.707T>C, g.812A>G, g.18951A>T g.16867T>C and g.18953Indel GGCAAAGTAA) were detected in the GnRHR gene from one-hundred-sixty-five water buffalo by direct sequencing and identification of overlapping peaks. All SNPs were associated significantly with the ejaculate volume and two genes (g.655T>C and g.707T>C) were correlated with sperm abnormalities. Furthermore, three haplotypes (H1:TAI, H2:CT-, and H3:TT-) were identified by linkage disequilibrium analysis and were composed of four combined genotypes. Notably, buffalo with the combined genotypes H1H2 and H1H3 had the higher ejaculate volume compared to the other combined genotypes. Among the eight SNPs and four combined genotypes, the deletion of GGCAAAGTAA at position 18953bp in GnRHR was associated significantly with a higher ejaculate volume. Moreover, the GGCAAAGTAA deletion may lead to the miR8661 binding failure and subsequent changes in GnRHR gene expression. In the present study, we demonstrate that there is a significant association between SNPs in the GnRHR gene and the sperm ejaculate volume of Chinese water buffalo. To the best of our knowledge, this study is the first to address the association between the SNPs in the GnRHR gene and the sperm quality of Chinese buffalo.

The outcomes of intracytoplasmic sperm injection (ICSI) using spermatozoa with Y chromosome microdeletion

AZF microdeletions are the leading genetic cause of male infertility and useful to obtain reliable genetic information for assisted reproductive techniques. Thus, the detection is clinically relevant for appropriate genetic counseling. However, there are only a few reports evaluating clinical course of the patients with AZF microdeletions. The aim of this study was to clarify the results of microdissection testicular sperm extraction (micro TESE) with Y chromosome microdeletion and ICSI outcomes using sperm from those men. A retrospective study in 18 Japanese reproductive center. This study was retrospectively examined a total of 1934 azoospermic or severe oligozoospermic patinets who undergone genetic testing for AZF deletions in 18 Japanese medical centers from April 2007 to June 2016. We examined genetic testing for AZF deletions by Promega Y Chromosome AZF Analysis System (version 2.0). We firstly investigated each type of AZF deletions frequencies, and sperm retrieval rate (SRR) by micro TESE of the azoospermic patients with AZF deletions. In addition, we analyzed the ICSI results using testicular or ejaculatory sperm of men with AZF deletions. AZF microdeletions were found in 189 cases (9.8%): 20 AZFa, 15 AZFb, 72 AZFc, 1 AZF a+b, 58 AZFb+c, and 23 AZFa+b+c. Of men with isolated AZFc deletion, spermatozoa were retrieved in 71.4% (30/42) by micro TESE. We could not find spermatozoa from patients with other AZF deletions. Twenty-nine couples underwent ICSI using testicular sperm retrieved by micro TESE, and 3 couples underwent ICSI using ejaculatory sperm. In totally, 572 metaphase 2 oocytes were retrieved by 62 oocyte retrieval cycles. Fertilization rate was 46.1% (220/477). We performed 62 embryo transfer (ET) cycles, mean replaced embryos were 1.39 per ET. Implantation rate per total transfer embryos were 19.8% (17/86), and clinical pregnancy rate per ET cycles were 27.4% (17/62). Abortion rate was 23.5% (4/17). Although we could not detect significant differences, the results of ICSI using ejaculatory sperm had better propensity in fertilization rate, implantation rate, and clinical pregnancy rate compared with the results of ICSI using testicular sperm (72.7% vs 44.1%, 50.0% vs 17.5%, 75.0% vs 24.1%, respectively). AZFc deletions are associated with severe dysregulations of spermatogenesis, however, sperm retrieval is possible in more than half by micro-TESE. It is useful to obtain reliable genetic information from azoospermic patients and to avoid unnecessary micro TESE. The results of this study provide useful actual clinical information for infertile couples with AZF deletions.

Implications of ejaculatory sperm DNA fragmentation on male infertility management.

Latest guidelines from the World Health Organization (WHO) on the laboratory examination of human semen still focus on traditional semen analysis (SA) (1). However, the more recent recommendations from the major societies in the andrology field suggest to utilize new functional tests in certain conditions (2,3). Although the American Urological Association (AUA) best practice statement (2) and the joint guideline course of European Academy of Andrology (EAA) and the European Association of Urology (EAU) on clinical andrology (2) stated that routine use of ejaculatory sperm DNA fragmentation (SDF) in all infertile couples is not supported by current literature, there is a general consensus that some of the infertile couples may benefit from sperm DNA integrity evaluation. Abnormal SDF, is characterized by more than 15% (fair sperm DNA integrity) to more than 30% (poor sperm integrity) fragmented DNA in ejaculatory sperm (4).

LEM-domain proteins are lost during human spermiogenesis but BAF and BAF-L persist

During spermiogenesis the spermatid nucleus is elongated, and dramatically reduced in size with protamines replacing histones to produce a highly compacted chromatin. After fertilisation, this process is reversed in the oocyte to form the male pronucleus. Emerging evidence, including the coordinated loss of the nuclear lamina (NL) and the histones, supports the involvement of the NL in spermatid nuclear remodelling, but how the NL links to the chromatin is not known. In somatic cells, interactions between the NL and the chromatin have been demonstrated: LEM-domain proteins and LBR interact with the NL and respectively, the chromatin proteins BAF and HP1. We therefore sought to characterise the lamina-chromatin interface during spermiogenesis, by investigating the localisation of six LEM-domain proteins, two BAF proteins and LBR, in human spermatids and spermatozoa. Using RT-PCR, IF and western blotting, we show that six of the proteins tested are present in spermatids: LEMD1, LEMD2 (a short isoform), ANKLE2, LAP2β, BAF and BAF-L, and three absent: Emerin, LBR and LEMD3. The full-length LEMD2 isoform, required for nuclear integrity in somatic cells, is absent. In spermatids, no protein localised to the nuclear periphery, but five were nucleoplasmic, receding towards the posterior nuclear pole as spermatids matured. Our study therefore establishes that the lamina-chromatin interface in human spermatids is radically distinct from that defined in somatic cells. In ejaculated spermatozoa, we detected only BAF and BAF-L, suggesting that they might contribute to the shaping of the spermatozoon nucleus and, after fertilisation, its transition to the male pronucleus.

ICSI outcome in patients with high DNA fragmentation: Testicular versus ejaculated spermatozoa.

Sperm DNA fragmentation (SDF) has emerged as an important biomarker in the assessment of male fertility potential with contradictory results regarding its effect on ICSI. The aim of this study was to evaluate intracytoplasmic sperm injection (ICSI) outcomes in male patients with high SDF using testicular versus ejaculated spermatozoa. This is a prospective study on 36 men with high-SDF levels who had a previous ICSI cycle from their ejaculates. A subsequent ICSI cycle was performed using spermatozoa retrieved through testicular sperm aspiration. Results of the prior ejaculate ICSI were compared with those of the TESA-ICSI. The mean (SD) SDF level was 56.36% (15.3%). Overall, there was no difference in the fertilization rate and embryo grading using ejaculate and testicular spermatozoa (46.4% vs. 47.8%, 50.2% vs. 53.4% respectively). However, clinical pregnancy was significantly higher in TESA group compared to ejaculated group (38.89% [14 of 36] vs. 13.8% [five of 36]). Moreover, 17 live births were documented in TESA group, and only three live births were documented in ejaculate group (p<.0001). We concluded that the use of testicular spermatozoa for ICSI significantly increases clinical pregnancy rate as well as live-birth rate in patients with high SDF.

Is early embryo development as observed by time-lapse microscopy dependent on whether fresh or frozen sperm was used for ICSI? A cohort study.

The aim of this study was to compare timings of key events of embryo development from those originating from either fresh or cryopreserved ejaculate sperm using time-lapse technology. In this retrospective observational cohort study, time-lapse technology was used to monitor 1927 embryos from 234 women undergoing intracytoplasmic sperm injection (ICSI) and utilizing either fresh (n=172 cycles) or cryopreserved ejaculate sperm (n=62 cycles) for insemination were included in the study. Key developmental events as described in time-lapse were compared with the use of generalized estimating equations (GEE) to adjust for any auto-correlation between the observations. In addition, multivariable logit regression models were used to account for any known baseline differences between the two groups. There were no differences in conventional embryo development such as number of 8-cell embryos by 72h (p=0.359), the number of blastocysts by 120h (p=0.417), and the number of top quality blastocysts (p=0.956) between the two groups compared. There were no statistical differences in the timings of any of the key embryo developmental events (PN_t1, NEBD, cytokinesis, t2, t3, t4, t5, t6, t7, t8, tM, tSB, tEB, tHB, s1, s2, s3, cc2, and cc3) when either fresh or cryopreserved ejaculate sperm was used for ICSI. This was also confirmed with conventional morphological assessment. This observational cohort study has shown that there are no differences in the morphokinetic parameters of early embryo development when either fresh or frozen ejaculate sperm are used for ICSI insemination.

KUALITAS SPERMA EJAKULAT PEJANTAN AYAM KUKUAK BALENGGEK PADA PENGANDANGAN TUNGGAL TERISOLASI (Ejaculated Sperm Quality of Isolated Single Caging of Balenggek Chickens)

The Balenggek chicken is a long crower type fowl. This fowl have isolated and caging in single cage for years to result the long crower sound.This study was carried out to identify the influence of fed to the ejaculate sperm quality of balenggek chickens. First, the sperm were collected from isolated caging of Balenggek fowl more or less than three years and un-isolated caging of Balenggek fowl as control group. Second, the sperm were collected from isolated caging of Balenggek fowl more or less than three years and un-isolated caging of Balenggek fowl as control group, after treated by 124 ComFed for three months. The sperm were collected by massages methods and analyze with counting chamber of Improve Neubauer. Slides of sperm were stained by Eosin. Variables observed were sperm ejaculated, colours, and sperm consistency. The results shows that the sperm quality of Balenggek fowl increased (from 2.5 billion sperm/ml to 3.5 billion sperm/ml) significantly higher (p&lt;0.05) in isolated caging fowl with reschedule feeding. Sperm color and its consistency were increased from + (c quality) to ++ (b quality). It can conclude that single caging could decreased the ejaculated sperm of Balengggek chickens quality and it. could reveal increased after it treated by 124 ConFed.

The effect of age, interval collection and season on selected semen parameters and prediction of AI boars productivity

The possibility of planning the production level has a special importance for AI stations, and its inaccuracy should be minimized by eliminating the factors differentiating the quantity and quality of the ejaculate. The aim of the study was to determine the effect of age, collection interval and season on selected quantitative and qualitative semen parameters and to develop equations for estimating production capacity for AI stations. The study included 48 117 ejaculates. Experimental groups were created according to: age of boar (8–9, 10–12, 13–18, 19–24, 25–30, >30 months), collection interval (≤2, 3, 4, 5, 6, 7, ≥8 days), and season (Winter, Spring, Summer, Autumn). Selected quantitative and qualitative parameters were analyzed: semen volume (ml), spermatozoa concentration (×106ml−1), total number of spermatozoa in ejaculate (×109), number of motile spermatozoa in ejaculate (×109), and number of insemination doses obtained from one ejaculate (n). The most important role in shaping the results was played by interactions. The highest semen volumes were achieved for the oldest boars after 7-day interval collection during Autumn (about 300ml). Interestingly, the lowest semen volumes were also noted during Autumn from the youngest boars after 1–2 and 7-day interval collection (a little over 160ml). The highest spermatozoa concentrations (515×106ml−1) were observed in Autumn for 10–12 month-old boars with the longest interval collection, which clearly demonstrates the positive impact of long resting on young boars. The lowest spermatozoa concentrations – below 300×106ml−1 – were reported for the oldest boars after 1–2 and 3-day collection intervals. The highest total number of spermatozoa in ejaculate and motile spermatozoa were noted during the longest collection interval in Winter at the ages of 13–18 months and 25–30 months, while the lowest occurred in the same season with a short interval from the 10–12 month boars. Almost all correlations were statistically proven at the level of P≤0.05 and P≤0.01, except for the correlation between the number of motile spermatozoa and the age of boars. Developed regression equations enable, with a high degree of accuracy, the estimation of the selected ejaculate parameters on the basis of age, collection interval and season. The solution proposed in the article is a useful tool, especially for AI stations or large farms with a high rate of boars producing semen for AI. AI stations assessing the long-term production capacity of boars should take into account the factors examined in this study. Underestimation of the interaction may, in fact, reduce the number of insemination doses by up to 5 portions of a single ejaculate.

Efficacy of treatment with pseudoephedrine in men with retrograde ejaculation.

The use of pseudoephedrine, an alpha agonist, for the treatment of retrograde ejaculation is well-known, however, there is no clear consensus from the literature regarding its efficacy and treatment protocol. We evaluated the efficacy of pseudoephedrine treatment in patients with retrograde ejaculation, utilizing a yet undescribed short-period treatment protocol. Twenty men were medically treated with pseudoephedrine for retrograde ejaculation between January 2010 and May 2016 (12 with complete retrograde ejaculation and 8 with partial retrograde ejaculation). All patients had a semen analysis and post-ejaculatory urinalysis before and after treatment. The treatment protocol consisted of 60mg of pseudoephedrine every 6h on the day before semen analysis and two more 60mg doses on the day of the semen analysis. Diabetes was the most common etiology for complete retrograde ejaculation (60%), whereas an idiopathic cause was the most common etiology for partial retrograde ejaculation (82%). Of the 12 complete retrograde ejaculation patients treated with pseudoephedrine prior to semen analysis, 7 (58.3%) recovered spermatozoa in the antegrade ejaculate, with a mean total sperm count of 273.5±172.5 million. Of the eight patients with partial retrograde ejaculation, five (62.5%) had a ≥50% increase in the antegrade total sperm count. In this group, the mean total sperm count increased from 26.9±8.5 million before treatment to 84.2±24.6 million after treatment, whereas the percentage of spermatozoa in the urine declined from 43.2±9% to 17±10%, respectively (both p<0.05). Overall, in men with retrograde ejaculation treated with a pseudoephedrine regimen prior to ejaculation, some improvement in seminal parameters occurred in 14 (70%) patients, with 10 patients (38.5% of all patients) achieving antegrade total sperm counts over 39 million.

MP89-05 THE ASSESSMENT OF HORMONE REPLACEMENT THERAPY SUCCESS DUE TO HYPOGONADOTROPHIC HYPOGONADISM

You have accessJournal of UrologyInfertility: Therapy I1 Apr 2017MP89-05 THE ASSESSMENT OF HORMONE REPLACEMENT THERAPY SUCCESS DUE TO HYPOGONADOTROPHIC HYPOGONADISM Emre Salabas, Ebubekir Buyuk, Caner Bese, Mazhar Ortac, and Ates Kadioglu Emre SalabasEmre Salabas More articles by this author , Ebubekir BuyukEbubekir Buyuk More articles by this author , Caner BeseCaner Bese More articles by this author , Mazhar OrtacMazhar Ortac More articles by this author , and Ates KadiogluAtes Kadioglu More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.2803AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Hypogonadatrophic hypogonadism is a rare disease in which medical treatment has high success rates for infertility. The aim of this study was to analyze the efficacy of hormone replacement therapy on the infertile patients with hypogonadotrophic hypogonadism (HH) diagnosis. METHODS A total of 250 patients had been diagnosed as HH between years 2002-2016 underwent Cella Pituitary MR imaging to exclude intracranial pathology, genetic and biochemical(hormone) analysis. Patients ,who had been diagnosed as HH with low testosterone and FSH levels but without surgical requirement, were administered LH analogues (Recombinant choriogonadotropin) and Recombinant FSH as hormone replacement therapy. Patient followed up was carried out with quarterly semen analysis and hormonal profiles. Detection of sperm in ejaculate and achievement of pregnancy spontaneously or with assisted reproductive tecniques were the predetermined primary and secondary endpoints. RESULTS The patients with regular follow-ups (135) had been selected out of the HH database with 250 patients and retrospectively investigated. During the treatment period(8-24 months), 94 (70%) patients had sperm appearance in their ejaculate. Out of the 72 married males with sperm appearing on their ejaculate, 40 patients (55%) obtained pregnancy spontaneously; 7 patients (%10) via assisted reproductive techniques. Forty one patients (%30) who had no sperm in their ejaculate yet are under routine control with hormonal treatment. CONCLUSIONS The medical approach of males ,with hypogonadotropic hypogonadism and azoospermia, is a successful treatment modality both in terms of sperm presence in ejaculate and pregnancy rates. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e1203-e1204 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Emre Salabas More articles by this author Ebubekir Buyuk More articles by this author Caner Bese More articles by this author Mazhar Ortac More articles by this author Ates Kadioglu More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Extreme spermatogenesis failure: andrological phenotype and intracytoplasmic sperm injection outcomes

Patients with very low sperm count through direct sperm examination can exhibit extreme oligozoospermia or cryptozoospermia (after centrifugation). The management of these patients is a real challenge for both clinicians and biologists. In this retrospective and comparative cohort study, we compared the andrological phenotype of patients with extreme alterations of spermatogenesis and assessed whether the origin of spermatozoa (testicular or ejaculate) had any influence on intracytoplasmic sperm injection (ICSI) outcomes. A total of 161 ICSI cycles were performed using ejaculated spermatozoa from 75 patients with extreme oligozoospermia (EOS) or cryptozoospermia (CS) and 150 ICSI cycles using extracted testicular spermatozoa from 74 patients with non-obstructive azoospermia (NOA). Physical, hormonal, ultrasound assessments, and ICSI outcomes were performed in each group. Cryptorchidism was significantly more frequent in the NOA group (60.8% vs. 22.6%, p=0.001). FSH levels were significantly higher [18.9IU/L (5.9-27.0) vs. 15.3IU/L (9.0-46.5), p=0.001] and the majority of inhibin B levels measured were found mostly undetectable in the NOA group as compared to EOS/CS group (31.1% vs. 10.7%, p=0.0004). Moreover, we found no significant differences in the respect to the fertilization rates (48.9% and 43.3%, p=0.43), implantation rates (17.4% and 15.9%, p=0.77), and percentage of top quality embryo (22.4% and 20.4%, p=0.73) between the two groups. The clinical pregnancy rates per embryo transferred were comparable in both groups (28.3% and 27.4%, p=0.89). In this study, we showed for the first time a different andrological phenotype between EOS/CS and NOA groups. Indeed, cryptorchidism was significantly more frequent with more severe endocrine parameters found in the NOA group. These results reflect a more profound alteration in spermatogenesis in NOA patients. However, there was no difference in ICSI outcomes between NOA and EOS/CS groups.

Testicular sperm aspiration (TESA) for infertile couples with severe or complete asthenozoospermia.

The aim of the study was to evaluate reproductive outcomes in a cohort of infertile couples with severe and complete asthenozoospermia undergoing TESA (testicular sperm aspiration) with ICSI. We conducted a retrospective study of 28 couples with complete or severe asthenozoospermia who underwent TESA between January 2010 and December 2015. We compared TESA-ICSI outcomes of these couples to ejaculate ICSI outcomes of 40 couples with severe asthenozoospermia treated during the same time period at our institution. Couples with female factor infertility and/or female aged ≥39 were excluded. Sperm retrieval rates and ICSI outcomes [(MII oocytes, fertilization rate, good embryo rate (transferred and frozen), couples with embryo transfer (per cycle started), clinical pregnancy (per embryo transfer)] were recorded. Patients were grouped based on whether they had ejaculated (Ej-group) or testicular (TESA-group) spermatozoa used. Testicular sperm patients were further classified based on whether they had complete asthenozoospermia (0% total motility) (Tc-group) or severe asthenozoospermia (≤1% progressive motility) (Ts-group). Mean (±SD) male and female ages were 36±6 and 32±4, respectively. Sperm recovery by testicular sperm aspiration (TESA) was successful in 100% (28/28) of the men. The overall clinical pregnancy rate (CPR) per cycle started was 34% (23/68) with a mean of 1.1±0.4 embryos transferred per transfer. Fertilization rates were significantly lower in TESA-group compared to Ej-group (52% vs. 67%, respectively; p=0.001), while male age was significantly higher in TESA-group compared to Ej-group (34±6 vs. 37±6, respectively; p=0.03). Moreover, female age was significantly higher in Tc-group compared to Ts-group (30±4 vs. 33±3, respectively; p=0.0285). However, there were no significant difference in clinical pregnancy rate per embryo transfer in the Tc-group, Ts-group, and Ej-group (50% vs. 45% vs. 57%, respectively; p=0.8219). The data suggest that testicular sperm-ICSI is no better than ejaculated sperm-ICSI in couples with severe or complete asthenozoospermia. Randomized, controlled trials comparing ejaculated vs. testicular spermatozoa are needed to assess the true benefit of TESA-ICSI in these couples.

Testosterone serum profile, semen characteristics and testicular biometry of Mangalarga Marchador stallions in a tropical environment.

This study was conducted to characterize the daily profile of testosterone secretion and its mean concentrations in the four seasons as well as to evaluate the semen characteristics and testicular biometry of Mangalarga Marchador stallions throughout the year in a tropical region. Three stallions were submitted to semen collections and evaluation of testicular biometry every 14days along a year. Blood samples were collected once at the middle of each season, in a 20-min interval during 24hr in order to evaluate the testosterone secretion profiles among seasons. Testosterone concentrations along the day were higher at the beginning of the afternoon (from 12:00 to 15:00hr), but a circadian secretion was not clearly observed. Mean testosterone concentrations did not differ among seasons (p>.05), but a pattern of secretion along the day showed variations with higher concentrations in the afternoon during the winter. Ejaculate volume was higher during summer; however, sperm motility decreased in summer and spring. Total sperm in ejaculate, sperm morphology and testicular biometry kept constant along the year showing no differences among the seasons. The results demonstrated that in a tropical region, reproductive aspects of stallions did not show a clearly defined seasonal variation, and months of autumn and winter were not unsuitable for reproduction of the males.

Effects of season on semen parameters and relationships between selected semen characteristics in Hypor boars

The aim of this study was to evaluate the effect of the season of the year on the quality of Hypor boar semen and to determine the correlations between selected semen characteristics in particular seasons. Ejaculates from Hypor boars were analyzed for volume, sperm concentration, percentage of spermatozoa with normal motility, total number of spermatozoa, and the number of insemination doses per ejaculate. Microscopic slides were prepared from semen samples. Each sample was evaluated for the morphology of spermatozoa. Spermatozoa with normal morphology were counted, as well as those with both major and minor abnormalities. Spermatozoa were measured for morphometric traits. Ejaculates were grouped by the season of collection: winter, spring, summer, and autumn. Correlation coefficients were calculated between sperm morphometric traits and the concentration and total count of spermatozoa in ejaculates. In spring and winter, sperm concentrations are higher compared to ejaculates collected in summer and autumn. In spring and summer, ejaculates contain spermatozoa of greatest motility. In spring, sperm cells have largest head dimensions. In autumn, spermatozoa are smallest in terms of dimensions. Significant correlations were observed between sperm concentration and total number of sperm in ejaculates vs. morphometric dimensions of sperm heads in Hypor boars' semen in the autumn, winter, and summer.

Varicocele and non-obstructive azoospermia – where to start?

Background. Male etiology is responsible for nearly half of cases of infertility. Frequency of azoospermia in infertile men reaches 30 %; varicocele is diagnosed in almost 40 % of the patients. Varicocele- associated infertility has traditionally been considered an indication for surgical treatment. At the same time, the main method of sperm retrieval in azoospermia is testicular biopsy (testicular sperm extraction, TESE). Objective: to assess the efficacy of surgical treatment in patients with non-obstructive azoospermia (NOA) and varicocele. Materials and methods . The study included 327 patients with NOA and varicocele aged between 22 and 48 years treated in the Department of Andrology and Urology at the V. I. Kulakov National Medical Research Center of Obstetrics, Gynecology, and Perinatology in 2012– 2016. Study participants were divided into two comparable groups. Patients in the first group underwent microsurgical varicocelectomy (Marmara procedure) on the first stage and testicular biopsy (micro-TESE) on the second stage. Patients in the second group underwent micro-TESE with no surgical correction of varicocele prior to it. Results. In the first group, the appearance of spermatozoa in ejaculate was observed in 84 (44.4 %) patients 6 month post microsurgical varicocelectomy. Upon micro-TESE, spermatozoa suitable for use in assisted reproductive technologies were obtained in 60 (56.7 %) men. Treatment was effective in 144 (76.2 %) patients from the first group. In the second group, spermatozoa were retrieved in 66 (47.8 %) men upon micro-TESE. Conclusion . Microsurgical varicocelectomy increases treatment efficacy in males with NOA and varicocele. In the absence of severe degeneration of the seminiferous epithelium, 72.6 % of patients had the opportunity of becoming a genetic parent, avoiding in some cases testicular biopsy.

Asthenozoospermi ve Oligoasthenozoospermi Erkek Olguların Ejekulat Spermatozoalarında NRF2 mRNA Ekspresyon Düzeyi

Asthenozoospermi ve Oligoasthenozoospermi Erkek Olguların Ejekulat Spermatozoalarında NRF2 mRNA Ekspresyon Düzeyi

Effect of seminal plasma from high- and low-fertility bulls on cauda epididymal sperm function.

The aim of this study was to characterise the effect of seminal plasma (SP) from bulls of high or low fertility on sperm function. First, the effect of SP on the motility of fresh cauda epididymal spermatozoa (CES) and frozen-thawed ejaculated spermatozoa was assessed (Experiment 1a). Seminal plasma was then collected from bulls of known high and low fertility. Pooled CES were incubated in the SP from each bull, diluted and assessed for motility and viability on Days 1, 2, 3 and 5 after packaging as fresh semen (Experiment 1b). Also assessed were motility, kinematics, viability and mitochondrial membrane potential after thawing (Experiment 1c) as well as hypotonic resistance (Experiment 2) and fertilisation potential using in vitro fertilisation (Experiment 3). Seminal plasma increased the motility of CES (P<0.05); however, there was no effect of SP on the motility and viability of fresh CES or on CES post-thaw motility, viability and mitochondrial membrane potential (P>0.05). The hypotonic resistance of CES was reduced by SP (P<0.05), irrespective of whether the SP was from high- or low-fertility bulls. Seminal plasma from high- or low-fertility bulls had no effect on cleavage or blastocyst rates (P>0.05). In conclusion, SP affects the physiological function of CES but there is no difference between SP from high- or low-fertility bulls.

Developmental competence of Dromedary camel oocytes fertilized in vitro by frozen-thawed ejaculated and epididymal spermatozoa.

The present study aimed to compare the in vitro fertilizing capacity of frozen-thawed ejaculated and epididymal spermatozoa in order to standardize the semen preparation protocol for camel in vitro fertilization (IVF). Semen samples were collected from 7 Dromedary camels by means of artificial vagina (AV). Ten cauda epididymes were obtained from slaughtered adult camels, isolated, incised and rinsed for obtaining the sperm rich fluid. Ejaculated and epididymal spermatozoa were processed for cryopreservation. Fresh and frozen-thawed ejaculated and epididymal spermatozoa were evaluated for motility, livability, membrane and acrosomal integrities. Frozen-thawed ejaculated and epididymal spermatozoa were used to fertilize camel mature oocytes in vitro. The results showed that, the progressive motility of freshly collected epididymal spermatozoa was significantly (P<0.05) higher than ejaculated spermatozoa (49.25 ± 1.75 vs. 38.50 ± 1.50%, respectively). The viability index of epididymal spermatozoa was significantly (P<0.05) higher than that of ejaculated spermatozoa (96.63 ± 2.45 vs. 84.00 ± 4.08, respectively). The post-thaw acrosome and membrane integrities of epididymal spermatozoa were significantly (P<0.05) higher than those of ejaculated spermatozoa. Morula and blastocyst rates of camel oocytes in vitro fertilized by frozen-thawed epididymal spermatozoa (59.4 ± 0.8, 19.12 ± 0.7 and 10.29 ± 0.7%, respectively) were significantly (P<0.05) higher than those fertilized by frozen-thawed ejaculated spermatozoa (48.27 ± 3.1, 11.63 ± 1.1 and 5.43 ± 0.8%, respectively). In conclusion, the Dromedary camel frozen epididymal spermatozoa have the capacity to endure cryopreservation, fertilize oocytes and produce embryos in vitro better than ejaculated sperm.

Spermaurin, an La1-like peptide from the venom of the scorpion Scorpio maurus palmatus, improves sperm motility and fertilization in different mammalian species.

Is it possible to identify original compounds that are able to enhance sperm motility from the venom of the scorpion Scorpio maurus palmatus? We identified a potent disulfide-rich peptide (DRP) of 73 amino acids that significantly improved the motility of fresh and frozen-thawed sperm in different mammalian species, including human, and improved fertilization outcome in mouse IVF experiments. Any disturbance of sperm motility has a strong impact on fertilization and can lead to subfertility or infertility. Significant efforts have, therefore, been made to identify pharmacological drugs that might improve sperm motility. Such compounds are particularly useful in azoospermia to improve testicular sperm extraction and in the domain of cryopreservation because the motility of frozen-thawed sperm is reduced. This was a basic science/medical research study aimed at identifying original compounds from a library of venoms able to enhance mammalian sperm motility, including human. We first identified in the venom of a scorpion S. m. palmatus a fraction able to potently activate sperm motility. We next purified and characterized the compound by liquid chromatography, mass spectrometry and peptide synthesis. Finally, the potency and toxicity of both purified and synthetic versions of the identified compound on sperm motility were assessed using different in vitro tests in different mammalian species. For human sperm, biological samples were collected from normozoospermic donors and subfertile patients attending a reproduction department for diagnostic semen analysis. Testicular sperm was collected from cynomolgus monkeys (Macaca fascicularis) euthanized for the needs of specific authorized research projects. The peptide was also tested on bovine and mouse epidydimal sperm. We measured different sperm motility parameters with a computer-assisted sperm analysis system in the presence or absence of the peptide. Size exclusion chromatography enabled us to isolate a fraction of the venom of S. m. palmatus able to increase sperm motility. By liquid chromatography and mass spectrometry, a peptide comprising 73 amino acids with 4 disulfide bridges was identified as responsible for the biological activity and called 'spermaurin'. The identity of spermaurin was confirmed by chemical synthesis. We showed that the peptide increased the motility of fresh and frozen-thawed human sperm. We observed that the potency of the peptide was higher on fresh ejaculated spermatozoa with a low motility, achieving a 100% increase of curvilinear velocity in poorly performing sperm. We also demonstrated that peptide is effective on bovine and mouse fresh epididymal, bovine frozen-thawed ejaculated and fresh non-human primate testicular sperm. Finally, in mouse IVF, the production of 2-cell embryos was increased by 24% when sperm were treated with the peptide. This work is an in vitro evaluation of the ability of spermaurin to improve sperm motility parameters. Another limitation of this study is the small number of human sperm samples tested with the natural (n = 36) and synthetic (n = 12) peptides. Moreover, the effect of the peptide on IVF outcome was only tested in mouse and further tests with human and bovine gametes are required to confirm and extend this result in other mammalian species. This work confirms our initial study showing that venoms represent an interesting source of molecules that are able to modify sperm physiology. Moreover, this work presents the first demonstrated biological action of a venom peptide from the scorpion S. m. palmatus with sequence similarities to La1 peptide from Liocheles australasiae (Wood scorpion), a widespread family of DRPs. Not applicable. This work is part of the project 'LAB COM-14 LAB7 0004 01-LIPAV', funded by the program LabCom 2014 from the French Research Agency (ANR). Dr Arnoult reports grants from IMV Technologies during the conduct of the study. In addition, Drs Arnoult, Martinez, Ray and Schmitt have a patent EP16305642.7 pending containing some of the information presented in this manuscript.