Technology Updates in Male Infertility Management

Background: Technologies are replacing manpower many fields including medical field. Several devices have been marketed for replacing/reducing manpower in the medical field including male infertility. Here we reviewed several technologies that developed in male infertility. Review: Computer assisted sperm analysis (CASA), Automatic assessment of biochemical marker of seminal plasma, B-mode ultrasound, and automatic sperm cryopreservation can be applied routinely. Several updates i.e. automatic histopathology assessment, ultrasound strain elastography, magnetic resonance imaging (MRI) stronger than 3.0 T, Artificial intelligence for predicting the presence of sperm in azoospermia cases, automatic sperm selection, and automatic intracytoplasmic sperm injection (ICSI) need more studies before their application. Summary: Prudent choice based on valid studies is needed in order to give a comprehensive management to patient with male infertility without using useless technology.

Cyclic FEE Peptide Improves Human Sperm Movement Parameters without Modification of Their Energy Metabolism

Cyclic fertilin peptide (cFEE: phenylalanine, glutamic acid; glutamic acid) improves gamete interaction in humans. We investigate whether it could be via improvement of sperm movement parameters and their mitochondrial ATP production. Sperm movement parameters were studied using computer-assisted sperm analysis (CASA) in sperm samples from 38 patients with normal sperm in medium supplemented with cyclic fertilin against a control group. Sperm mitochondrial functions were studied using donor’s sperm, incubated or not with cFEE. It was evaluated by the measurement of their ATP production using bioluminescence, their respiration by high resolution oxygraphy, and of mitochondrial membrane potential (MMP) using potentiometric dyes and flow cytometry. cFEE significantly improved sperm movement parameters and percentage of hyperactivated sperm. Impact of inhibitors showed OXPHOS as the predominant energy source for sperm movement. However, cFEE had no significant impact on any of the analyzed mitochondrial bioenergetic parameters, suggesting that it could act via a more efficient use of its energy resources.

Evaluation of Bali Cattle Semen Quality During Cryopreservation with Coconut Water-Based Extenders

The aim of this research was to evaluate the Bali cattle semen quality during cryopreservation with different types of extenders in term of live, total motility, progressive motility, and abnormality of post-thawed bull sperms. The treatments were AndroMed® (T0), Tris-based egg yolk diluent (T1), Tris-based coconut water diluent (T2), Coconut water egg yolk diluent (T3). Bulls’ semen was collected from two adult Bali cattle maintained at the semen production facility at Bali Artificial Insemination Center, Tabanan Bali. The age of the bulls were 6 years old. Sperm live, total motility, progressive motility, and abnormality were analyzed with computer assisted sperm analysis (CASA) post-dilution, before and after thawed. The study was replicated five times, and data were analyzed using analysis of variance (ANOVA). The results showed that AndroMed ® and Tris-based egg yolk had significantly higher sperm live, total motility, progressive motility and abnormalities of spermatozoa for post-dilution, after equilibration and post thawed than Tris egg yolk coconut water and Coconut water egg yolk diluent. It was concluded that AndroMed® and Tris-based egg yolk can be considered as the best suitable extender for Bali cattle sperm cryopreservation. Coconut water had a deleterious effect when supplemented with 20% in tris and egg yolk.

Effects of Three Semen Extenders, Breeding Season Month and Freezing–Thawing Cycle on Spermatozoa Preservation of Portuguese Merino Sheep

This study aimed to evaluate and compare the effect of three semen extenders (S-EXT) on 22 spermatozoa (SPZ) parameters (subjective and computer-assisted sperm analysis evaluations), before and after semen cryopreservation throughout different months of the breeding season in the Portuguese Merino breed. According to the multivariable model, the SPZ viability (alive %), kinetics subjective individual motility, total motility, total progressive motility and its subpopulations, and beat cross frequency) were higher in the egg yolk-based S-EXT improved by Estação Zootécnica National (Portugal) than in Ovixcell® or Andromed® extenders. All the differences were only observed in thawed semen, except for total motility and total progressive motility, in which Ovixcell® also showed the poorest results on fresh semen. An interaction effect between S-EXT and semen processing was observed on 72.3% (17/22) of the evaluated parameters, evidencing a variable cryoprotective action between S-EXT. The SPZ viability was poorer in the onset of the breeding season (end of April/early May) than in the previous middle breeding season (November/early December), suggesting the influence of a short anoestrous season on ejaculate quality, even though the volume and SPZ concentration of the ejaculates remained stable throughout the experiment. Additionally, S-EXT x semen processing x month interaction effect on 59.1% (13/22) of the evaluated parameters evidenced the importance of SPZ time collection in a natural environment to cryopreserve ram’s semen. We concluded that, overall, the egg yolk-based S-EXT provided a greater value to the cryopreservation of Merino rams´ semen. Nevertheless, the causes of the interaction effect between S-EXT, semen processing and/or month on several SPZ parameters should be addressed, including SPZ molecular research in new studies, in order to improve egg yolk-based as well as in egg yolk-free-based S-EXT.

Post-Thaw Sperm Quality and Functionality in the Autochthonous Pig Breed Gochu Asturcelta

Genetic resource banks (GRB) preserve the genetic material of endangered, valuable individuals or genetically relevant breeds. Semen cryopreservation is a crucial technique to reach these goals. Thus, we aimed to assess the sperm parameters of semen doses from the native pig breed Gochu Asturcelta stored at the GRB of Principado de Asturias (GRB-PA, Gijón, Spain), focusing on intrinsic and extrinsic (boar, season) factors. Two straws per boar (n = 18, 8–71 months of age) were thawed, pooled, and assessed after 30 and 150 min at 37 °C by CASA (computer-assisted sperm analysis system; motility and kinematic parameters) and flow cytometry (viability, acrosomal status, mitochondrial activity, apoptosis, reactive oxygen species, and chromatin status). The effects of age, incubation, and season on post-thawing quality were determined using linear mixed-effects models. Parameters were on the range for commercial boar breeds, with chromatin status (SCSA: fragmentation and immaturity) being excellent. Incubation decreased sperm quality and functionality. The boar age did not have a significant effect (p > 0.05), but the between-boar variability was significant (p < 0.001). The season significantly affected many parameters (motility, kinematics, viability, acrosomal status, mitochondrial activity), especially after 150 min of incubation. In general, samples collected in spring and summer showed higher quality post-thawing, the lowest in winter. In conclusion, the sperm doses from the Gochu Asturcelta breed stored at the GRB-PA showed excellent chromatin status and acceptable characteristics after thawing. Therefore, boar and seasonal variability in this autochthonous breed could be relevant for cryobank management.

Διερεύνηση της επίδρασης το πρωτόζωου Toxoplasma gondii στη γονιμοποιητική ικανότητα του κριού

Σκοπός αυτής της μελέτης ήταν η διερεύνηση της επίδρασης της πειραματικής μόλυνσης με Toxoplasma gondii στην ποιότητα του σπέρματος του κριού. Τριάντα δύο άνηβοι κριοί ηλικίας 5 μηνών χωρίστηκαν σε 4 ομάδες (n=8). Η ομάδα Α αποτέλεσε την ομάδα ελέγχου, ενώ οι υπόλοιπες τρεις ομάδες (Β, Γ, Δ) μολύνθηκαν από το στόμα με 5000 ώριμες ωοκύστεις του πρωτόζωου T. gondii. Στην ομάδα Γ χορηγήθηκε σουλφαδιμιδίνη (33mg/kg ΣΒ, i.m., ανά διήμερο, για 8 ημέρες) 2 μήνες μετά τη μόλυνση και στην ομάδα Δ η ίδια αγωγή δύο φορές (24 ώρες μετά τη μόλυνση και 2 μήνες αργότερα). Η ομάδα Β δεν έλαβε φαρμακευτική αγωγή. Δείγματα αίματος συλλέγονταν ανά 15 ημέρες για την ανίχνευση ειδικών αντισωμάτων IgG (ELISA). Τα ζώα όλων των ομάδων θανατώθηκαν 4 μήνες μετά τη μόλυνση και από τις επιδιδυμίδες τους συλλέχθηκαν δείγματα σπέρματος τα οποία υποβλήθηκαν σε α) ανίχνευση του παρασίτου με PCR, β) εξετάσεις ποιοτικών χαρακτηριστικών: πυκνότητα (αιμοκυτταρόμετρο Neubauer), ζωτικότητα και μορφολογία (χρώση Εωσίνης–Νιγροσίνης), κινητικότητα (computer–assisted sperm analysis system — CASA), βιοχημική λειτουργικότητα των μεμβρανών των σπερματοζωαρίων (δοκιμή HOS) και ακεραιότητα της δομής της χρωματίνης (DNA) του πυρήνα των σπερματοζωαρίων (δοκιμή πορτοκαλόχρωμης ακριδίνης). Επιπλέον, λήφθηκαν ιστοτεμάχια από τους όρχεις για ιστοπαθολογική εξέταση. Στα μολυσμένα ζώα ο τίτλος αντισωμάτων αυξήθηκε τη 2η εβδομάδα μετά τη μόλυνση και παρέμεινε υψηλός για 4 μήνες. Η χορήγηση σουλφαδιμιδίνης i.m. 24 ώρες μετά τη πειραματική μόλυνση άνηβων αρσενικών αμνών με T. gondii, επέφερε καθυστέρηση στην ανοσιακή απόκριση κατά 1 εβδομάδα, αλλά και χαμηλότερες τιμές ειδικών αντισωμάτων IgG. Το DNA του παρασίτου ανιχνεύθηκε σε 3 δείγματα σπέρματος μολυσμένων κριών (12,5%), 15 εβδομάδες μετά τη μόλυνση. Σημαντικά υψηλότερες τιμές παρατηρήθηκαν στη ζωτικότητα και στη βιοχημική λειτουργικότητα των μεμβρανών των σπερματοζωαρίων στην ομάδα ελέγχου σε σύγκριση με τις μολυσμένες ομάδες, επίπεδο σημαντικότητας p <0,05. Oι μορφολογικές ανωμαλίες ήταν περισσότερες στις ομάδες Γ, Δ έναντι της Α και στη Γ έναντι της Β (p<0,05). Η πειραματική μόλυνση με T. gondii δεν επέφερε κάποια σοβαρή επιβάρυνση στα κύρια χαρακτηριστικά κίνησης του σπέρματος των κριών και δεν επηρέασε την ακεραιότητα της δομής της χρωματίνης (DNA) του πυρήνα των σπερματοζωαρίων τους. Η ιστοπαθολογική εξέταση στους όρχεις αποκάλυψε όμοια ευρήματα με ελάχιστες διακυμάνσεις μεταξύ των μολυσμένων ομάδων, οι οποίες χαρακτηρίζονταν κυρίως από αυξημένο διάμεσο συνδετικό ιστό, μη πυώδη φλεγμονή και παρουσία σπερματικών σωληναρίων με διαταραχή της σπερματογένεσης, τα οποία αυξάνονταν βαθμιαία από την Δ στην Γ και στην ομάδα Β. Δεν ανευρέθηκαν κύστεις τοξοπλάσματος σε κανένα από τα δείγματα ιστών από τα μολυσμένα ζώα. Συνοψίζοντας, η ολοκλήρωση της παρούσας διατριβής οδήγησε στo συμπέρασμα ότι η πειραματική μόλυνση κριών κατά την προ–εφηβική ηλικιακή περίοδο με ώριμες ωοκύστεις του Τ. gondii υποβαθμίζει την ποιότητα του σπέρματος κατά την εφηβεία, συνεπώς και τη γονιμοποιητική ικανότητά του, ενώ η χορήγηση σουλφαδιμιδίνης δεν μπορεί να αποτρέψει αυτό το αποτέλεσμα.

Differences in the proteome of stallion spermatozoa explain stallion-to-stallion variability in sperm quality post-thaw†

The identification of stallions and or ejaculates that will provide commercially acceptable quality post-thaw before cryopreservation is of great interest, avoiding wasting time and resources freezing ejaculates that will not achieve sufficient quality to be marketed. Our hypothesis was that after bioinformatic analysis, the study of the stallion sperm proteome can provide discriminant variables able to predict the post-thaw quality of the ejaculate. At least three ejaculates from 10 different stallions were frozen following a split sample design. Half of the ejaculate was analyzed as a fresh aliquot and the other half was frozen and then analyzed as a frozen-thawed aliquot. Computer-assisted sperm analysis and flow cytometry were used to analyze sperm quality. Detailed proteomic analysis was performed on fresh and frozen and thawed aliquots, and bioinformatic analysis was used to identify discriminant variables in fresh samples able to predict the outcome of cryopreservation. Those with a fold change &gt; 3, a P = 8.2e-04, and a q = 0.074 (equivalent to False discovery rate (FDR)) were selected, and the following proteins were identified in fresh samples as discriminant variables of good motility post-thaw: F6YTG8, K9K273, A0A3Q2I7V9, F7CE45, F6YU15, and F6SKR3. Other discriminant variables were also identified as predictors of good mitochondrial membrane potential and viability post-thaw. We concluded that proteomic approaches are a powerful tool to improve current sperm biotechnologies.