Sperm Selection Research Articles

The effect of advanced sperm selection methods on clinical pregnancy results in cases with diminished ovarian reserve

The effect of advanced sperm selection methods on clinical pregnancy results in cases with diminished ovarian reserve

Lab-on-chip (LoC) application for quality sperm selection: An undelivered promise?

Quality sperm selection is essential to ensure the effectiveness of assisted reproductive techniques (ART). However, the methods employed for sperm selection in ART often yield suboptimal outcomes, contributing to lower success rates. In recent years, microfluidic devices have emerged as a promising avenue for investigating the natural swimming behavior of spermatozoa and developing innovative approaches for quality sperm selection. Despite their potential, the commercial translation of microfluidic-based technologies has remained limited. This comprehensive review aims to critically evaluate the inherent potential of lab-on-chip technology in unraveling sophisticated mechanisms encompassing rheotaxis, thermotaxis, and chemotaxis. By reviewing the current state-of-the-art associated with microfluidic engineering and the swimming of spermatozoa, the goal is to shed light on the multifaceted factors that have impeded the broader commercialization of these cutting-edge technologies and recommend a commercial that can surmount the prevailing constraints. Furthermore, this scholarly exploration seeks to enlighten and actively engage reproductive clinicians in the profound potential and implications of microfluidic methodologies within the context of human infertility.

Age-Dependent Alterations in Semen Parameters and Human Sperm MicroRNA Profile.

The trend to delay parenthood is increasing, impacting fertility and reproductive outcomes. Advanced paternal age (APA), defined as men's age above 40 years at conception, has been linked with testicular impairment, abnormal semen parameters, and poor reproductive and birth outcomes. Recently, the significance of sperm microRNA for fertilization and embryonic development has emerged. This work aimed to investigate the effects of men's age on semen parameters and sperm microRNA profiles. The ejaculates of 333 Portuguese men were collected between 2018 and 2022, analyzed according to WHO guidelines, and a density gradient sperm selection was performed. For microRNA expression analysis, 16 normozoospermic human sperm samples were selected and divided into four age groups: ≤30, 31-35, 36-40, and >40 years. microRNA target genes were retrieved from the miRDB and TargetScan databases and Gene Ontology analysis was performed using the DAVID tool. No significant correlation was found between male age and conventional semen parameters, except for volume. Fifteen differentially expressed microRNAs (DEMs) between groups were identified. Enrichment analysis suggested the involvement of DEMs in the sperm of men with advanced age in critical biological processes like embryonic development, morphogenesis, and male gonad development. Targets of DEMs were involved in signaling pathways previously associated with the ageing process, including cellular senescence, autophagy, insulin, and mTOR pathways. These results suggest that although conventional semen parameters were not affected by men's age, alterations in microRNA regulation may occur and be responsible for poor fertility and reproductive outcomes associated with APA.

An intracellular, non-oxidative factor activates in vitro chromatin fragmentation in pig sperm

BackgroundIn vitro incubation of epididymal and vas deferens sperm with Mn2+ induces Sperm Chromatin Fragmentation (SCF), a mechanism that causes double-stranded breaks in toroid-linker regions (TLRs). Whether this mechanism, thought to require the participation of topoisomerases and/or DNAses and thus far only described in epididymal mouse sperm, can be triggered in ejaculated sperm is yet to be elucidated. The current study aimed to determine if exposure of pig ejaculated sperm to divalent ions (Mn2+ and Mg2+) activates SCF, and whether this has any impact on sperm function and survival. For this purpose, sperm DNA integrity was evaluated through the Comet assay and Pulsed Field Gel Electrophoresis (PFGE); sperm motility and agglutination were assessed with computer assisted sperm analysis (CASA); and sperm viability and levels of total reactive oxygen species (ROS) and superoxides were determined through flow cytometry.ResultsIncubation with Mn2+/Ca2+ activated SCF in a dose-dependent (P < 0.05) albeit not time-dependent manner (P > 0.05); in contrast, Mg2+/Ca2+ only triggered SCF at high concentrations (50 mM). The PFGE revealed that, when activated by Mn2+/Ca2+ or Mg2+/Ca2+, SCF generated DNA fragments of 33–194 Kb, compatible with the size of one or multiple toroids. Besides, Mn2+/Ca2+ affected sperm motility in a dose-dependent manner (P < 0.05), whereas Mg2+/Ca2+ only impaired this variable at high concentrations (P < 0.05). While this effect on motility was concomitant with an increase of agglutination, neither viability nor ROS levels were affected by Mn2+/Ca2+ or Mg2+/Ca2+ treatments.ConclusionMn2+/Ca2+ and Mn2+/Ca2+ were observed to induce SCF in ejaculated sperm, resulting in DNA cleavage at TLRs. The activation of this mechanism by an intracellular, non-oxidative factor sheds light on the events taking place during sperm cell death.

IZUMO1 Receptor Localization during Hyaluronic Acid Selection in Human Spermatozoa.

IZUMO1 is an acrosome transmembrane protein implicated in the adhesion and fusion of gametes. This study aims to describe the distribution of IZUMO1 in human sperm under different physiological conditions: before capacitation (NCS), at one-hour capacitation (CS1), after a hyaluronic acid (HA) selection test (mature, MS1 and immature, IS1), and induced acrosome reaction from one-hour-capacitated sperm (ARS1). The data obtained in NCS, CS1, and MS1 significantly highlight dotted fluorescence in the acrosomal region (P1) as the major staining pattern (~70%). Moreover, we describe a new distribution pattern (P2) with a dotted acrosomal region and a labelled equatorial region that significantly increases in HA-bound spermatozoa, suggesting the onset of the migration of IZUMO1. In contrast, unbound spermatozoa presented an increase in P3 (equatorial region labelled) and P4 (not labelled). Finally, costaining to observe IZUMO1 distribution and acrosome status was performed in ARS1. Interestingly, we reported a variety of combinations between the IZUMO1 staining patterns and the acrosomal stages. In conclusion, these data show as a novelty the diffusion of the IZUMO1 protein during different physiological conditions that could contribute to the improvement in sperm selection techniques.

The birth of a healthy child in a married couple with a history of multiple unsuccessful attempts at ART by selecting sperm on oocyte-cumulus complexes. Case report

In male infertility, couples must use assisted reproductive technologies with the intracytoplasmic sperm injection (ICSI) technique. A significant disadvantage of ICSI is bypassing natural selection barriers since selective processes in the female reproductive tract are replaced by the choice of an embryologist based on sperm motility and morphological criteria. Various selection methods are being actively studied, bringing fertilization by ICSI closer to physiological, ensuring the selection of sperm with high DNA quality and good potential for fertilization. The article presents a clinical case of the birth of a healthy child in a married couple with severe male infertility and many ineffective attempts at assisted reproductive technologies using the method of sperm selection on oocyte-cumulus complexes. The child is physically healthy and develops according to age.

Biointerfacial behavior of stallion spermatozoa adhered to hydrogel surfaces: Impact of the hydrogel chemical composition and the culture medium

Novel soft materials based on hydrogel are proposed to enhance the selection of high-quality stallion sperm based on their adhesion capacity. The hydrogel surfaces are derived from polyacrylamide (PAAm), which is copolymerized with neutral and ionic co-monomers to modify the interfacial properties. The hydrogels undergo characterization through FTIR spectroscopy, assessment of swelling capacity, and wettability under various experimental conditions. Sperm adhesion capacity on the hydrogels is examined through several parameters including the percentage of bound sperm (%Sp) to hydrogels, tail oscillation intensity and flagellar movement. The biointerfacial properties of sperm-hydrogel systems vary based on the chemical composition of hydrogel as well as the components present in the culture medium. High %Sp and excellent metabolic activity of the spermatozoa are observed on hydrogel surfaces that possess moderate hydrophilicity. Specifically, a cationic hydrogel in BGM3 culture medium and a neutral surface in BGM3 medium supplemented with BSA exhibit favorable outcomes. Scanning Electron Microscopy (SEM) reveals the normal morphology of the head and tail in spermatozoa adhered to the hydrogel. Therefore, these hydrogel surfaces are potential materials for selecting stallion sperm with high quality, and their application could be extended to the study of other mammalian reproductive cells.

Exposure to Cumulus Cell Secretome Improves Sperm Function: New Perspectives for Sperm Selection In Vitro.

In the literature, there is a well-known correlation between poor semen quality and DNA sperm integrity, which can turn into negative outcomes in terms of embryo development and clinical pregnancy. Sperm selection plays a pivotal role in clinical practice, and the most widely used methods are mainly based on sperm motility and morphology. The cumulus oophorus complex (COC) during natural fertilization represents a barrier that spermatozoa must overcome to reach the zona pellucida and fertilize the oocyte. Spermatozoa that can pass through the COC have better structural and metabolic characteristics as well as enhanced acrosome reaction (AR). The present study aimed to evaluate the exposure of sperm to cumulus cell secretome during swim-up treatment (SUC) compared with the routinely used swim-up method (SU). To determine the effectiveness of this method, biological factors critical for the ability of sperm to fertilize an oocyte, including capacitation, AR, tyrosine phosphorylation signature, DNA integrity, and mitochondrial functionality, were assessed. The SUC selection assures recovery of high-quality spermatozoa, with enhanced mitochondrial functionality and motility compared with both SU-selected and unselected (U) sperm. Furthermore, using this modified swim-up procedure, significantly reduced sperm DNA damage (p < 0.05) was detected. In conclusion, the SUC approach is a more physiological and integrated method for sperm selection that deserves further investigation for its translation into clinical practice.

Antioxidant Properties and Toxic Risks of Using Metal Nanoparticles on Health and Productivity in Poultry

Metal nanoparticles (NPs) are introduced into various fields of science, particularly poultry farming. Supplementation of metal salts in nanoform can increase the profitability of poultry farming by enhancing meat and egg production. Although their toxic parameters pose limitations on their use, many studies have evaluated the effects of using metal NPs in modern poultry farming on health, productivity, metabolism, and especially antioxidant properties. In addition, the peculiarities of their toxicokinetic and recommended doses that meet safety criteria in practical activities are highlighted. Zinc oxide NPs are one of the most studied compounds in the poultry industry. Their pronounced antioxidant properties, positive effect on productivity and homeostasis of poultry, egg quality, and immune status have been experimentally confirmed. Copper oxide NPs have similar properties but are limited in usage due to their toxicokinetics. Silver and gold NPs emerge as potential alternatives to antibiotics and could solve the resistance problem of microorganisms to antibiotics. Other important NPs used in poultry are Iron and Calcium. In their nanoform, these NPs exhibit high bioavailability, which allows for efficient absorption and utilization by poultry. The methods used to synthesize these nanoparticles make it economically viable to incorporate them into poultry diets, reducing overall expenses compared to similar macroergic compounds. Manganese and chromium NPs positively affect sperm survival in turkeys during refrigerated storage and contribute to increasing the resistance of the broiler chickens' body to heat stress and normalizing the metabolism of sex hormones. In conclusion, the application of metal nanoparticles to poultry is a promising research direction, aiming at the development of feed additives, antibiotics, and growth stimulants due to their antioxidant, bactericidal, and immunomodulatory effects.

Stain-Free Sperm Analysis and Selection for Intracytoplasmic Sperm Injection Complying with WHO Strict Normal Criteria.

This multi-center study evaluated a novel microscope system capable of quantitative phase microscopy (QPM) for label-free sperm-cell selection for intracytoplasmic sperm injection (ICSI). Seventy-three patients were enrolled in four in vitro fertilization (IVF) units, where senior embryologists were asked to select 11 apparently normal and 11 overtly abnormal sperm cells, in accordance with current clinical practice, using a micromanipulator and 60× bright field microscopy. Following sperm selection and imaging via QPM, the individual sperm cell was chemically stained per World Health Organization (WHO) 2021 protocols and imaged via bright field microscopy for subsequent manual measurements by embryologists who were blinded to the QPM measurements. A comparison of the two modalities resulted in mean differences of 0.18 µm (CI -0.442-0.808 µm, 95%, STD-0.32 µm) for head length, -0.26 µm (CI -0.86-0.33 µm, 95%, STD-0.29 µm) for head width, 0.17 (CI -0.12-0.478, 95%, STD-0.15) for length-width ratio and 5.7 for acrosome-head area ratio (CI -12.81-24.33, 95%, STD-9.6). The repeatability of the measurements was significantly higher in the QPM modality. Surprisingly, only 19% of the subjectively pre-selected normal cells were found to be normal according to the WHO2021 criteria. The measurements of cells imaged stain-free through QPM were found to be in good agreement with the measurements performed on the reference method of stained cells imaged through bright field microscopy. QPM is non-toxic and non-invasive and can improve the clinical effectiveness of ICSI by choosing sperm cells that meet the strict criteria of the WHO2021.

Can in vitro embryo production be estimated from semen variables in Senepol breed by using artificial intelligence?

Thoroughly analyzing the sperm and exploring the information obtained using artificial intelligence (AI) could be the key to improving fertility estimation. Artificial neural networks have already been applied to calculate zootechnical indices in animals and predict fertility in humans. This method of estimating the results of reproductive biotechnologies, such as in vitro embryo production (IVEP) in cattle, could be valuable for livestock production. This study was developed to model IVEP estimates in Senepol animals based on various sperm attributes, through retrospective data from 290 IVEP routines performed using 38 commercial doses of semen from Senepol bulls. All sperm samples that had undergone the same procedure during sperm selection for in vitro fertilization were evaluated using a computer-assisted sperm analysis (CASA) system to define sperm subpopulations. Sperm morphology was also analyzed in a wet preparation, and the integrity of the plasma and acrosomal membranes, mitochondrial potential, oxidative status, and chromatin resistance were evaluated using flow cytometry. A previous study identified three sperm subpopulations in such samples and the information used in tandem with other sperm quality variables to perform an AI analysis. AI analysis generated models that estimated IVEP based on the season, donor, percentage of viable oocytes, and 18 other sperm predictor variables. The accuracy of the results obtained for the three best AI models for predicting the IVEP was 90.7, 75.3, and 79.6%, respectively. Therefore, applying this AI technique would enable the estimation of high or low embryo production for individual bulls based on the sperm analysis information.

Oral 1 - Does Sperm Selection with Intracytoplasmic Sperm Injection (ICSI) improve outcomes in men with raised Sperm DNA Fragmentation? A Systematic Review and Meta-analysis

Oral 1 - Does Sperm Selection with Intracytoplasmic Sperm Injection (ICSI) improve outcomes in men with raised Sperm DNA Fragmentation? A Systematic Review and Meta-analysis

Nuclear sperm integrity and ICSI prognosis in Tunisian patients with MMAF syndrome (multiple morphological abnormalities of the sperm flagella)

Multiple Morphological Abnormalities of the Sperm Flagella (MMAF) is a severe form of teratozoospermia associated with several sperm flagellar abnormalities. The study included 52 patients with MMAF syndrome and a control group of 25 fertile men. The impact of nuclear sperm quality on intracytoplasmic sperm injection (ICSI) results was studied in 20 couples. TUNEL assay was used to assess sperm DNA fragmentation and aniline-blue staining was used to assess chromatin condensation. To investigate chromosomal meiotic segregation, we used fluorescence in situ hybridization (FISH). Semen morphology analysis revealed a mosaic of multiple flagella morphological abnormalities, including 46.73% short flagella, 16.22% bent flagella, 22.07% coiled flagella, and 10.90% absent flagella, all of which were associated with a high percentage of sperm head abnormalities. The mean DNA fragmentation index was substantially higher in patients compared to controls (p = 0.001), whereas the rate of aniline blue-reacted spermatozoa was not significantly different. There was a significant difference in aneuploidy frequencies between the two groups (p < 0.05). Infertile males with MMAF syndrome had lower sperm nuclear quality, which affected ICSI results. As a result, better sperm selection procedures are being employed to increase the success rate of assisted reproductive technologies (ART).

Rotation of liquid-preserved artificial insemination doses on roller benches affects sperm quality during storage in stallions.

Appropriate stallion semen handling is of great importance in equine artificial insemination (AI) industry. Optimal treatment of AI doses is aiming for best sperm preservation by excluding strong environmental influences and adding media that favour sperm survival. One method widely used in stallion sperm handling is the rotation of liquid-preserved semen samples on roller benches during storage. As previous studies in boars give rise to the fact that rotation should not be considered beneficial for spermatozoa anymore, the present study investigated the influence of continuous rotation of diluted stallion AI portions on sperm quality. Ejaculates (n = 15) were collected at a German AI centre and diluted with the two extenders EquiPlus and Gent (Minitüb GmbH) to a final concentration of 50 × 106 sperm/mL. Afterwards, samples were placed separately on roller benches at 5°C in the dark, obtaining a rotation frequency of 5 revolutions per minute (rpm) and 36 rpm for four consecutive days following a split-sample design. Both groups were analysed daily in comparison to a control group (0 rpm) with an extended spectrum of spermatological methods including computer-assisted sperm analysis and flow cytometry. Statistical analyses were based on the calculation of generalized linear mixed models for each sperm parameter. The research revealed a decrease in sperm quality parameters of rotated samples compared to non-rotated control groups, visible in total sperm motility (p < .001), decreased thermo-resistance (p < .01) and a drop in pH (p < .001). Interestingly, no differences (p > .05) were detected between rotation frequencies of 5 and 36 rpm. We conclude that the fertilizing capacity of stallion semen was negatively affected by rotation during storage in vitro, irrespective of the rotation frequency. Further studies need to investigate whether field fertility in horses is similarly affected by semen rotation on roller benches in vivo.

Esculeoside A alleviates reproductive toxicity in streptozotocin-diabetic rats’ s model by activating Nrf2 signaling

This examination studied if Esculeoside A (ESA) alleviates reproductive toxicity in a type 1 diabetes mellitus (T1DM) rat model and if activating Nrf2 underlies this protection. T1DM was established by a single injection of STZ. Aged-matched adult control and STZ-DM rats were administered either the vehicle (5% carboxymethyl cellulose) or ESA (100 mg/kg). An additional group [STZ-DM + ESA (100 mg) + brusatol (2 m/kg] was added. All treatments were conducted for 16 weeks. ESA failed to attenuate weight loss, hyperglycemia, and hypoinsulinemia but significantly attenuated the associated dyslipidemia in STZ-DM rats. In parallel, ESA also enhanced total sperm count, motility, survival, reduced head and tail sperm abnormalities, increased circulatory concentrations of follicular stimulating hormone (FSH), testosterone, and Luteinizing hormone (LH), and stimulated the testicular expression of several steroidogenic enzymes (StAR, CYP11A1, CYP17A1, 3β-HSD1) in STZ-DM rats. These observations were associated with a higher testicular increase in the transcription, protein levels, and nuclear activities of Nrf2 that coincided with a reduction in the total levels of MDA and keap1 and a significant increase in the total levels of some antioxidants such as HO-1, SOD, and GSH. In concomitance, ESA reduced the testicular mRNA and nuclear concentrations of NF-κB and depressed the levels of TNF-α and IL-6. Brusatol prevented all these protective effects of ESA. In conclusion, activation of Nrf2 triggers the protective potential of ESA against reproductive toxicity in STZ-DM rats.

Strategies for Bacterial Eradication from Human and Animal Semen Samples: Current Options and Future Alternatives.

The primary role of semen processing and preservation is to maintain a high proportion of structurally and functionally competent and mature spermatozoa, that may be used for the purposes of artificial reproduction when needed, whilst minimizing any potential causes of sperm deterioration during ex vivo semen handling. Out of a multitude of variables determining the success of sperm preservation, bacterial contamination has been acknowledged with an increased interest because of its often unpredictable and complex effects on semen quality. Whilst antibiotics are usually the most straight-forward option to prevent the bacterial contamination of semen, antimicrobial resistance has become a serious threat requiring widespread attention. As such, besides discussing the consequences of bacteriospermia on the sperm vitality and the risks of antibiotic overuse in andrology, this paper summarizes the currently available evidence on alternative strategies to prevent bacterial contamination of semen prior to, during, and following sperm processing, selection, and preservation. Alternative antibacterial supplements are reviewed, and emphasis is given to modern methods of sperm selection that may be combined by the physical removal of bacteria prior to sperm preservation or by use in assisted reproductive technologies.

WITHDRAWN: A proof-of-concept prospective study of applying artificial intelligence for sperm selection in the IVF laboratory

WITHDRAWN: A proof-of-concept prospective study of applying artificial intelligence for sperm selection in the IVF laboratory

Morphological analyses of sperms causing severe male infertility with light microscopy, motile sperm organellar morphology examination and electron microscopy tecnics.

To evaluate abnormal sperm morphology by three different methods including light microscope, real-time motile sperm organelle morphology assessment at 8050x high magnification and electron microscope and to determine degree of affect on morphologic defects level. Beside the light and electron microscopic techniques, the motile sperm organelle examination can offer information about sperm quality in a simple and quick way and by the IMSI developed through the use of sperm selected by this technique can contribute to the success of in vitro fertilization centers. We believe that sperm selection at high magnifications can increase IVF pregnancy rates in patients with sperm morphology that causes severe male infertility.

Semen Collection in a Device Specifically Designed for Human Semen Improves Sample Physiological and Morphological Parameters

Objectives: The current series of studies examined the use of the device for improved semen collection (DISC) and its effects on semen quality specifically designed for human semen collection. The system was designed to lessen cellular shock by providing an environment that optimized conditions for sperm survival. Materials and Methods: A series of experiments were conducted to examine the effects of the DISC, on sperm cell physiology, biochemistry and, in a small FDA-approved clinical trial, to prove equivalence for pregnancy outcomes. Results: Samples collected into the DISC maintained sample motility as well as other movement parameters for extended periods of time when compared to samples collected in a traditional standard specimen cup (P &lt; 0.05). Further, the samples appeared to maintain biochemical functions, such as the acrosome reaction (P &lt; 0.001) and mitochondrial function (P &lt; .001), suggesting the DISC improved sample environment, which protects cells during the collection process.

MACS-DGC sperm preparation method resulted in high-quality sperm, top-quality embryo, and higher blastocyst rate in male factor infertile couples with high DNA fragmented sperm

Conventional sperm selection based on motility and morphology fails to provide detailed information on sperm functional and molecular status. Magnetic-activated cell sorting (MACS) protocol aims to optimize this process by selecting apoptotic sperm cells. Phospholipase C zeta-1 (PLCz1) is a physiological stimulus for oocyte activation and early embryonic development. The purpose of this study was to examine seminal parameters, DNA fragmentation index (DFI), and PLCz1 expression levels in MACS-DGC sorted specimens (DFI > 30%) and assess early development in resulting embryos. Semen specimens from 60 patients diagnosed with male factor infertility were collected and processed by either density gradient centrifugation (DGC) or MACS-DGC protocols. Pre and post-preparation analysis was performed. PLCz1 expression was assessed using the RT-PCR method. Retrieved eggs from their partners were divided into two groups in which they were injected with different sorted sperm. The fertilization rate and embryonic development were evaluated. While sperm’s progressive motility and morphology significantly improved, there was a substantial decline in DFI following MACS-DGC. Fertilization rates were almost the same between the groups, and the latter resulted in remarkably more top-quality embryos and more blastocysts. PLCz1 expression was considerably higher in the MACS-DGC group. By eliminating apoptotic cells, the MACS-DGC technique could sort highly PLCz1-expressed sperm, optimize sperm selection in individuals with elevated DFI, development of resulting embryos.