The role of S-palmitoylation of C4BPA in regulating murine sperm motility and complement resistance.

The epididymis and epididymosomes are crucial for regulating sperm motility, a key factor in male fertility. Palmitoylation, a lipid modification involving the attachment of palmitic acid to cysteine residues, is essential for protein function and localization. Additionally, this modification plays a vital role in the sorting of proteins into exosomes. This study investigates the role of S-palmitoylation at the Cys15 residue of the C4b binding protein alpha chain (C4BPA) in murine sperm motility. Our findings revealed high expression of C4BPA mRNA in the caput epididymis, with the protein present across all regions of the epididymis. Palmitoylation of C4BPA in epididymal epithelial cells was essential for its enrichment in epididymosomes and on sperm, thereby maintaining sperm motility. Inhibition of palmitoylation significantly reduced sperm motility and the localization of C4BPA on sperm. Additionally, palmitoylated C4BPA in exosomes resisted complement C4 attacks, preserving motility, unlike mutated C4BPA (C15S). These results highlight the critical role of palmitoylated C4BPA in protecting sperm from complement attacks and maintaining motility, suggesting that reversible palmitoylation of epididymal proteins could be explored as a therapeutic strategy for male contraception. Our study underscores the importance of post-translational modifications in sperm function and presents new insights into potential male contraceptive methods.

Quality assurance applied to semen analysis.

Artificial insemination is an important biotechnology employed in livestock production. Production of semen products requires analysis of sperm concentration, motility and morphology. Although adequate analytical procedures are essential to ensure product quality, several multicenter studies have reported large variations in semen analysis results within and across laboratories. Differences in equipment and methodology, inconsistent training and performance testing, and lack of quality assurance programs are likely responsible for these observations. The somewhat pervasive perception that semen analysis is a trivial task must be challenged as it jeopardizes efficient semen production and germplasm utilization. This manuscript reviews the Quality Assurance (QA) procedures recommended to control results obtained during semen analysis. Reference, standard technical specifications for basic semen analysis are also described.

Influence of free and microencapsulated recombinant rabbit nerve growth factor with chitosan on rabbit sperm quality parameters.

ß-nerve growth factor (ßNGF) plays a crucial role in reproductive physiology and sperm quality. Enzymatic activity of seminal plasma and vaginal fluids reduces available ßNGF and it has been demonstrated that chitosan microspheres could protect rrßNGF from degradation. This study examined the effects of microencapsulated rrbNGF with chitosan on rabbit sperm viability, motility and capacitation status. Results showed that 0.5 and 1 µg/mL of microencapsulated rrßNGF, as well as free rrßNGF or empty microspheres, did not adversely affect sperm viability or total motility after 2 h of incubation. However, the highest progressivity kinetic parameters were observed with 1 µg/mL free rrßNGF, while the highest curvilinear velocity (VCL) occurred with 0.5 µg/mL microencapsulated rrßNGF. Empty chitosan microspheres did not induce acrosome reaction (AR), but both concentrations of free and rrßNGFch favoured AR during in vitro incubation. The study suggests that using chitosan spheres did not show any adverse effects on sperm traits, unlike free rßNGF and rrßNGFch promoted capacitation and AR. Further research is needed to explore the potential of rrßNGFch in modifying in vitro capacitation and inducing ovulation during artificial insemination.

Characterizing the Impact of Dysregulated Micrornas on CRISP3 Isoforms in Male Infertility.

microRNAs (miRNAs) have a serious and dynamic function in spermatogenesis. These molecules have been recognized as crucial parts of the control of gene activity, and their involvement in the regulation of target genes has been extensively studied. This research aimed to determine the expression of CRISP3 and miR-493-5p, miR-204-5p, and miR-182-5p in the seminal plasma fluid and spermatozoa and to examine the relationship between CRISP3 and the mentioned miRNAs in 57 infertile men with Asthenozoospermia (AZ) (n = 19), Teratoasthenozoospermia (TAZ) (n = 19), and Normozoospermia (NZ) (n = 19). The selection of these three miRNAs, miR-493-5p, miR-204-5p, and miR-182-5p, was conducted using computational prediction algorithms. These miRNAs were nominated as CRISP3-associated miRNAs that can target CRISP3. We performed the quantitative real-time polymerase chain reaction (qRT-PCR) method to determine the levels of the studied miRNA expression. In the following stage, the expression of two protein isoforms of CRISP3, targeted by these miRNAs, was quantified using western blotting. The results demonstrate significant differences in the levels of miR-182-5p, miR-204-5p, miR-493-5p, and CRISP3 isoforms among the patient groups. In TAZ individuals, miR-182-5p and miR-204-5p expression decreased, while miR-493-5p expression increased compared to the control samples. Additionally, significant differences were observed in the expression levels of unglycosylated and glycosylated CRISP3 isoforms between the AZ and NZ groups. Correlation analysis revealed associations between miRNA expression and the expression of CRISP3 isoforms in the patient groups. Additionally, there were correlations between the expression of CRISP3 isoforms and sperm motility and morphology. These results offer valuable insights into the underlying molecular processes associated with male infertility.

Study of the Association of Semen Parameters With Embryo Quality in a Tertiary Care Center in Bihar: A Retrospective Study.

Introduction Semen quality, characterized by parameters such as sperm count, motility, and morphology, determines successful fertilization and subsequent embryo health. This study investigates the impact of sperm parameters on embryo quality in assisted reproductive technology (ART). Methods The study utilized a retrospective design with 194 male and 194 female participants who underwent fertility treatment from October 2020 to May 2024. Semen analysis included assessment of sperm count, motility, and morphology. Embryo quality was evaluated on days three and five post-fertilizations based on morphological criteria. Statistical analyses, including one-way ANOVA and chi-square tests, explored relationships between semen parameters and embryo quality. Results The study included 388 participants (males: 194 and females: 194). Female participants had a mean age of 31.0 ± 4.6 years and a mean BMI of 23.1 ± 5.3 kg/m², while males had a mean age of 36.6 ± 5.4 years and a mean BMI of 22.7 ± 2.8 kg/m². Paternal age and BMI showed no significant association (p > 0.05) with embryo quality. However, sperm quality parameters such as sperm count, motility, and morphology demonstrated significant associations (p < 0.05) with embryo quality on both day three and day five, indicating that abnormal sperm parameters were linked to poorer embryo quality. Factors such as alcohol consumption, smoking, tobacco use, living in industrial areas, and tea/coffee consumption showed no significant association with embryo quality. Conclusion The findings of the study emphasize the importance of comprehensive semen analysis in fertility assessments and highlight opportunities for improving ART outcomes through targeted interventions and further mechanistic research.

Addition of Mitoquinone (MitoQ) to Fresh Human Sperm Enhances Sperm Motility without Attenuating Viability.

The preparation of human sperm in an andrology laboratory subjects it to oxidative stress. Reactive oxygen species are produced by mitochondria, making it susceptible to oxidative damage; hence, mitochondria-targeted antioxidants like Mitoquinone (MitoQ) might have therapeutic potential for oxidative-damage-associated disorders. The current research aims to establish whether MitoQ has any positive effects during in vitro preparation of fresh human sperm. Viability and motility are evaluated to determine the effective MitoQ concentration and to assess whether MitoQ supplementation is affected by sperm concentration by incubating normospermia semen samples at 37 °C for 2 h and 4 h, respectively. The effect of semen centrifugation following supplementation of 20 × 106 sperm/mL with 200 nM MitoQ is also assessed by measuring viability, motility and sperm DNA fragmentation. The best sperm motility is achieved after 2 h of incubation with 200 nM MitoQ at 37 °C. Sperm concentration of 20 × 106 sperm/mL is the best concentration where 200 nM MitoQ works efficiently. For semen centrifugation at 300× g for 20 min, supplementation with 200 nM MitoQ shows higher sperm motility. The current results demonstrate that MitoQ supplementation during in vitro human semen preparation procedures positively affects fresh sperm motility without affecting viability or increasing DNA fragmentation.

Sperm Migration and Hyaluronic Acid Binding: Implications for Male Fertility Evaluation.

Mature, vital, and motile spermatozoa are essential for reaching the oocyte and binding to hyaluronic acid (HA) in the cumulus oophorus matrix. This study aims to determine the relationship between sperm-migration ability and HA-binding potential, as well as the relationship between sperm concentration and motility. Semen samples were collected from 702 men aged 20-56 years (median 34.8). We evaluated the sperm concentration and motility from basic semen analysis, the swim-up test (expressed as millions per mL and the migration efficiency percentage), and the hyaluronan-binding assay (HBA). A moderate positive correlation was found between the migration test results and HBA (R = 0.48). The highest correlation was observed between the concentration of motile spermatozoa and the migration test results (R = 0.85) and HBA (R = 0.4). The sperm migration efficiency strongly correlated with progressive motility (R = 0.6). Although significantly higher sperm migration was observed in patients with normal HBA results, the results of the functional tests were found to differ in some cases. For infertility treatment, the current diagnostic algorithm should be enhanced with more comprehensive seminological methods that assess the sperm-migration ability and HA-binding potential. We also recommend incorporating the swim-up method into the diagnostic protocol before planning assisted reproductive technology (ART) treatment.

Enhancing canine semen quality through a second centrifugation after 48 hours of storage: a comparative study.

BACKGROUND: Centrifugation is a common procedure to improve the quality of chilled and frozen canine semen by removing debris and seminal plasma and adding semen extenders. The aim of this study was to evaluate the efficacy and influence of a second centrifugation after 48 h of storage at 5 °C on the sperm quality of canine semen. The ejaculates of 45 healthy male dogs, divided into three groups according to body weight, were analyzed for macro- and microparameters such as ejaculate volume, sperm concentration, kinematic parameters, morphology, and integrity of plasma membrane. Samples were analyzed at baseline conditions (T0), after 24 h (T24) and after 48 h (T48) to assess the effects of the different treatments on sperm quality. RESULTS: The results showed a significant effect of a second centrifugation on the improvement of chilled sperm quality compared to the other techniques, especially up to 48 h. CONCLUSIONS: Analysis of the data showed that the semen samples centrifuged and then cooled at 5 °C had acceptable semen parameters, especially in terms of motility, with a gradual decrease in serial evaluations after 24 and 48 h. A second centrifugation after 48 h of storage may lead to better semen quality and improve the kinetics of sperm parameters, the percentage of morphologically normal sperm and the percentage of sperm with intact membranes.

Effect of Two Different Sperm Selection Methods on Boar Sperm Parameters and In Vitro Fertilisation Outcomes.

Porcine in vitro embryo production (IVP) protocols have conventionally used density gradient selection (DGS) by centrifugation to prepare sperm samples and achieve successful fertilisation. However, the possible toxicity of the solutions used and the potential damage caused by the centrifugation step may have a negative effect on the quality of the sample. Microfluidic chip-based sperm (MCS) sorting has been proposed as an alternative technique for the selection of high-quality sperm with the purpose of improving reproductive outcomes in IVF. This device does not require centrifugation or any toxic solution to prepare the sample for fertilisation. The sample is not subjected to unnecessary stress, and the process is less operator-dependent. In this study, we compared the sperm parameters of unselected extender-diluted boar semen samples with selected samples using DGS and MCS methods. The results show an expected reduction in sperm concentration after both methods. All the groups were significantly different from one another, with MCS being the group with the lowest concentration. Though the three groups had a similar overall motility, significant differences were found in progressive motility when comparing the unselected group (control, 19.5 ± 1.4%) with DGS and MCS. Progressive motility in DGS was also significantly higher than in MCS (65.2 ± 4.9% and 45.7% ± 5.3, respectively). However, MCS selection resulted in enriched sperm samples with a significantly lower proportion of morphologically abnormal sperm compared to DGS. After fertilisation, no statistical differences were found between the two methods for embryological parameters such as cleavage rates, blastulation rates, and embryo quality. The number of cells in blastocysts derived from MCS was significantly greater than those derived from DGS sperm. Thus, we demonstrate that MCS is at least as good as the standard DGS for most measures. As a more gentle and reproducible approach for sperm selection, however, it could improve consistency and improve IVP outcomes as mediated by a greater proportion of morphologically normal sperm and manifested by a higher cell count in blastocysts.

Aquaporin-3a Dysfunction Impairs Osmoadaptation in Post-Activated Marine Fish Spermatozoa.

Spermatozoon volume regulation is an essential determinant of male fertility competence in mammals and oviparous fishes. In mammals, aquaporin water channels (AQP3, -7 and -8) have been suggested to play a role in spermatozoon cell volume regulatory responses in the hypotonic female oviduct. In contrast, the ejaculated spermatozoa of marine teleosts, such as the gilthead seabream (Sparus aurata), experience a high hypertonic shock in seawater, initially resulting in an Aqp1aa-mediated water efflux, cell shrinkage and the activation of motility. Further regulatory recovery of cell volume in post-activated spermatozoa is mediated by Aqp4a in cooperation with the Trpv4 Ca2+ channel and other ion channels and transporters. Using a paralog-specific antibody, here, we show that seabream spermatozoa also express the aquaglyceroporin AQP3 ortholog Aqp3a, which is highly accumulated in the mid posterior region of the spermatozoon flagella, in a similar pattern to that described in mouse and human sperm. To investigate the role of Aqp3a in seabream sperm motility, we used a recently developed AQP3 antagonist (DFP00173), as well as the seabream Aqp3a-specific antibody (α-SaAqp3a), both of which specifically inhibit Aqp3a-mediated water conductance when the channel was heterologously expressed in Xenopus laevis oocytes. Inhibition with either DFP00173 or α-SaAqp3a did not affect sperm motility activation but did impair the spermatozoon motion kinetics at 30 s post activation in a dose-dependent manner. Interestingly, in close resemblance to the phenotypes of AQP3-deficient murine sperm, electron microscopy image analysis revealed that both Aqp3a inhibitors induce abnormal sperm tail morphologies, including swelling and angulation of the tail, with complete coiling of the flagella in some cases. These findings suggest a conserved role of Aqp3a as an osmosensor that regulates cell volume in fish spermatozoa under a high hypertonic stress, thereby controlling the efflux of water and/or solutes in the post-activated spermatozoon.

Quantitative assessment of Nigella sativa and conjugated silver nanoparticles against hexavalent chromium toxic effects on sperm function.

BACKGROUND: Infertility has been observed as one of the major issues in humans, one known risk factor is heavy metals. METHODS: The main focus of the present research was to assess the toxic effect of hexavalent chromium (Cr (VI)) on sperm and its mitigation by Nigella sativa seed extract (NS) and its conjugated silver nanoparticles (NS + NP). In the present study, we administered 1.5 mg/kg body of Cr (VI) orally in mice for 60 days routinely, to induce toxicity in testes and effect on sperm production and motility in male mice. NS and NS + NP (50 mg/kg body weight) were administered to evaluate protective action against Cr (VI). The sperm were analyzed by computer-assisted semen analysis (CASA) and chromium concentration in testicular tissue was measured via the atomic absorption spectrophotometer. RESULTS: The CASA analysis showed that Cr (VI) was directly linked with a decline in sperm concentration, motility, distance, velocity, straightness, and head beat frequency attributes. However, the administration of Nigella sativa seed extract and its green synthesized silver nanoparticles improved sperm concentration, motility, distance, velocity, straightness, and head beat frequency. The chromium content in the testes of Cr-exposed animals significantly increased, which negatively affected sperm parameters. However, Nigella sativa and Nigella sativa conjugated silver nanoparticles appeared to help in the removal of Cr content from testes hence improving the sperm parameters in exposed mice. CONCLUSION: The decrease in Cr concentration improved sperm quality and quantity, hence, improve male fertility.

The efficacy and functional consequences of interactions between human spermatozoa and seminal fluid extracellular vesicles.

Abstract: Seminal fluid extracellular vesicles (SFEVs) have previously been shown to interact with spermatozoa and influence their fertilisation capacity. Here, we sought to extend these studies by exploring the functional consequences of SFEV interactions with human spermatozoa. SFEVs were isolated from the seminal fluid of normozoospermic donors prior to assessing the kinetics of sperm-SFEV binding in vitro, as well as the effects of these interactions on sperm capacitation, acrosomal exocytosis, and motility profile. Biotin-labelled SFEV proteins were transferred primarily to the flagellum of spermatozoa within minutes of co-incubation, although additional foci of SFEV biotinylated proteins also labelled the mid-piece and head domain. Functional analyses of high-quality spermatozoa collected following liquefaction revealed that SFEVs did not influence sperm motility during incubation at pH 5, yet SFEVs induced subtle increases in total and progressive motility in sperm incubated with SFEVs at pH 7. Additional investigation of sperm motility kinematic parameters revealed that SFEVs significantly decreased beat cross frequency and increased distance straight line, linearity, straightness, straight line velocity, and wobble. SFEVs did not influence sperm capacitation status or the ability of sperm to undergo acrosomal exocytosis. Functional assessment of both high- and low-quality spermatozoa collected prior to liquefaction showed limited SFEV influence, with these vesicles inducing only subtle decreases in beat cross frequency in spermatozoa of both groups. These findings raise the prospect that, aside from subtle effects on sperm motility, the encapsulated SFEV cargo may be destined for physiological targets other than the male germline, notably the female reproductive tract. Lay Summary: A male's influence over the biological processes of pregnancy extends beyond the provision of sperm. Molecular signals present in the ejaculate can influence the likelihood of pregnancy and healthy pregnancy progression, but the identity and function of these signals remain unclear. In this study, we wanted to understand if nano-sized particles present in the male ejaculate, called seminal fluid extracellular vesicles, can assist sperm in traversing the female reproductive tract to access the egg. To explore this, we isolated seminal fluid extracellular vesicles from human semen and incubated them with sperm. Our data showed that seminal fluid extracellular vesicles act to transfer molecular information to sperm, but this resulted in only subtle changes to the movement of sperm.

Oxidative stress affects sperm health and fertility-Time to apply facts learned at the bench to help the patient: Lessons for busy clinicians.

Background: Increased oxidative stress (OS), resulting from the delicate balance between reactive oxygen species (ROS) production and antioxidant defense, is closely linked to sperm abnormalities and male subfertility. Elevated ROS levels particularly affect sperm quality. The vulnerability of spermatozoa to ROS is due to the absence of DNA repair mechanisms and the high presence of polyunsaturated fatty acids in their membranes. Methods: This article updates and advances our understanding of the molecular damage caused by OS in spermatozoa, including lipid peroxidation, DNA damage, motility, and functionality. Additionally, the review discusses the challenges in diagnosing OS in semen and recommends accurate and sensitive testing methods. Case studies are utilized to demonstrate the effective management of male infertility caused by OS. Main findings: Highlighting the need to bridge the gap between research and clinical practice, this review suggests strategies for clinicians, such as lifestyle and dietary changes and antioxidant therapies. The review emphasizes lifestyle modifications and personalized care as effective strategies in managing male infertility caused by OS. Conclusion: This review calls for early detection and intervention and interdisciplinary collaboration to improve patient care in male infertility cases related to increased OS.

Lipidomic and transcriptomic characteristics of boar seminal plasma extracellular vesicles associated with sperm motility.

Seminal plasma extracellular vesicles (SPEVs) play an important role in regulating sperm motility by delivering various cargoes, such as miRNAs, mRNAs, proteins and metabolites. However, information on the lipid compositions of SPEVs and their roles in semen quality is limited. Here, we performed high-throughput transcriptomic and lipidomic analysis on SPEVs isolated from 20 boars with high or low sperm motility. Then, we evaluated the lipid composition and gene expression characteristics of SPEVs and identified the specific lipids and genes related to sperm motility. As a result, a total of 26 lipid classes were identified in SPEVs, and five subclasses, CerG2, CerG3, LPE, LPS and TG, were significantly different in boars with high and low sperm motility. In addition, 195 important lipids and 334 important genes were identified by weighted gene coexpression analysis (WGCNA) and differential expression analysis. We observed that several important genes and lipids in SPEVs potentially influence sperm motility via glycerophospholipid metabolism, glycerolipid metabolism, the sphingolipid signaling pathway and the ferroptosis pathway. Furthermore, we found a significant correlation between the content of 22 lipids and the expression levels of 67 genes (|cor| > 0.8, P < 0.05). Moreover, we observed that three important gene-lipid linkages (CerG1 (d22:0/24:0) - RCAN3, Cer (d18:1/24:0) - SCFD2 and CerG1 (d18:0/24:1) - SCFD2) were strongly correlated with sperm motility. Based on the results, some genes and lipids in SPEVs may play important roles in sperm motility by interacting with sperm through important pathways.

Supplementation of Rooster Semen Extender with Aqueous Extract of Urtica dioica for a Long Time Preservation by Low Temperature.

The peroxidation of spermatozoa membrane phospholipids is a primary cause of irreversible changes in the preservation of avian semen. To address this issue, the objective of the present study was to assess the potential of Urtica dioica extracts in protecting avian spermatozoa during prolonged storage. Gas chromatography-mass spectroscopic techniques were employed to evaluate the bioactive compounds present in the aqueous and ethanolic extracts obtained from the aerial parts and roots of U. dioica. Semen samples were collected from 16 roosters twice a week and were diluted in Lake's extender containing different concentrations (0, 0.5, and 1 mg/100 mL) of the various extracts. Subsequently, the extended semen samples were cooled and stored at 5°C, and the sperm quality parameters were assessed at 0, 12, 24, and 36 hours of storage. The data from this experiment clearly demonstrate that the addition of nettle root aqueous extracts to the semen diluent, especially at a concentration of 0.5 mg/100 mL, resulted in a significant improvement in various sperm quality parameters. Notably, there were enhancements in total and progressive sperm motilities, viability, fertility, membrane integrity, acrosomal membrane integrity, and a reduction in malondialdehyde production in rooster semen stored in vitro for up to 36 hours. Interestingly, the present study reveals that the beneficial effects of the aqueous extracts from different parts of the nettle were supported not only by the conventional manual method but also by the computer-assisted sperm analysis system. This dual confirmation further emphasizes the positive impact of the aqueous extract on various sperm traits during cooled semen preservation. In conclusion, this study highlights the potential of U. dioica extracts, particularly the aqueous extract from nettle roots at a concentration of 0.5 mg/100 mL, in safeguarding avian spermatozoa during prolonged storage. The significant improvements in various sperm quality parameters and the validation of results through both manual and computer-assisted analysis methods provide strong evidence for the application of U. dioica extracts in avian breeding programs and artificial insemination practices.

Seasonal seminal quality variations and vitrification prospects in black flounder Paralichthys orbignyanus.

Long-term preservation of gametes has been identified as a tool to improve broodstock management and increase the number of juveniles produced by artificial fertilization. Paralichthys orbignyanus is an important commercial and recreational species distributed in marine and estuarine waters from Rio de Janeiro (Brazil) to the San Matías Gulf (Argentina). This work focused on studying the seminal quality of tank-reared P. orbignyanus, demonstrating that males are fluent year-round, with the highest yields at the early reproductive season. Fresh sperm exhibited good forward swimming, and samples could be refrigerated up to 48 h while retaining their motility after activation. The optimal conditions for P. orbignyanus sperm motility activation were established as 950 mOsmol/Kg and pH values between 7 and 7.9. Additionally, a well-defined protocol for semen vitrification was developed to assess the cryotolerance of this species' sperm. We successfully produced high-quality sperm samples, using two vitrification formulations containing trehalose and both z-1000 and x-1000 polymers, that can be used in a near-future in vitro embryo production program.

Feeding spent hemp biomass does not adversely affect fertility in rams.

OBJECTIVE: To determine the reproductive effects of feeding spent hemp biomass (SHB) to rams. Several studies suggest cannabidiols negatively affect reproductive characteristics, and the reproductive effects of SHB ingestion have not been investigated in any species. Spent hemp biomass is high in protein and essential fatty acids, indicating a potential use in livestock diets pending studies investigating its safety in animals. METHODS: Polypay rams approximately 6 months old were randomly assigned to 5 feeding trial groups (n = 7/group): either a low or high concentration of SHB in diet for either 4 or 8 weeks plus a control group. Blood samples were collected for hormone assays. At the conclusion of the feeding trial, the testes were removed, and sperm collected directly from the vas deferens were evaluated for motility (total, progressive, and speed), morphology, and concentration. A section from each ram's testes was formalin fixed, paraffin embedded, and subjected to routine immunohistochemistry to determine the expression of fertility-associated proteins deleted in azoospermia-like and Boule, followed by quantitative image analysis. RESULTS: Rams fed either 10% or 20% SHB for 8 weeks had significantly higher progressive sperm motility compared to controls. There were no significant differences between the treatment and control groups in the other parameters studied. Boule immunoexpression was adversely affected, whereas deleted in azoospermia-like immunoexpression was differentially affected by SHB ingestion. CONCLUSION: We conclude that up to 20% of the diet can be fed as SHB to rams for 8 weeks without adversely affecting testicular or sperm function. CLINICAL RELEVANCE: Supplementing young rams with SHB is unlikely to affect fertility.

Ethanol-related transcriptomic changes in mouse testes.

BACKGROUND: Alcohol consumption is widely known to have detrimental effects on various organs and tissues. The effects of ethanol on male reproduction have been studied at the physiological and cellular levels, but no systematic study has examined the effects of ethanol on male reproduction-related gene expression. RESULTS: We employed a model of chronic ethanol administration using the Lieber-DeCarli diet. Ethanol-fed mice showed normal testicular and epididymal integrity, and sperm morphology, but decreased sperm count. Total RNA sequencing analysis of testes from ethanol-fed mice showed that a small fraction (∼ 2%) of testicular genes were differentially expressed in ethanol-fed mice and that, of these genes, 28% were cell-type specific in the testis. Various in silico analyses were performed, and gene set enrichment analysis revealed that sperm tail structure-related genes, including forkhead box J1 (Foxj1), were down-regulated in testes of ethanol-fed mice. Consistent with this result, ethanol-fed mice exhibited decreased sperm motility. CONCLUSION: This study provides the first comprehensive transcriptomic profiling of ethanol-induced changes in the mouse testis, and suggests gene expression profile changes as a potential mechanism underlying ethanol-mediated reproductive dysfunction, such as impaired sperm motility.

Proteomics and Metabolic Characteristics of Boar Seminal Plasma Extracellular Vesicles Reveal Biomarker Candidates Related to Sperm Motility.

Although seminal plasma extracellular vesicles (SPEVs) play important roles in sperm function, little is known about their metabolite compositions and roles in sperm motility. Here, we performed metabolomics and proteomics analysis of boar SPEVs with high or low sperm motility to investigate specific biomarkers affecting sperm motility. In total, 140 proteins and 32 metabolites were obtained through differentially expressed analysis and weighted gene coexpression network analysis (WGCNA). Seven differentially expressed proteins (DEPs) (ADIRF, EPS8L1, PRCP, CD81, PTPRD, CSK, LOC100736569) and six differentially expressed metabolites (DEMs) (adenosine, beclomethasone, 1,2-benzenedicarboxylic acid, urea, 1-methyl-l-histidine, and palmitic acid) were also identified in WGCNA significant modules. Joint pathway analysis revealed that three DEPs (GART, ADCY7, and NTPCR) and two DEMs (urea and adenosine) were involved in purine metabolism. Our results suggested that there was significant correlation between proteins and metabolites, such as IL4I1 and urea (r = 0.86). Furthermore, we detected the expression level of GART, ADCY7, and CDC42 in sperm of two groups, which further verified the experimental results. This study revealed that several proteins and metabolites in SPEVs play important roles in sperm motility. Our results offered new insights into the complex mechanism of sperm motility and identified potential biomarkers for male reproductive diseases.

The protective effects of antioxidants against endogenous and exogenous oxidative stress on bull sperm.

Oxidative stress, caused by both endogenous and exogenous factors, affects sperm function by damaging morphology and reducing metabolic activity, leading to reduced fertilization ability. The purpose of this study was to investigate the effects of oxidative stress on bull sperm and to evaluate the efficacy of targeted antioxidants in mitigating these detrimental effects. Fresh bull semen samples were subjected to hydrogen peroxide (H2O2) and antimycin treatments to induce oxidative stress, and the antioxidants PQQ, ergothioneine, and vitamin C were applied to counteract the induced stress. Sperm motility, viability, and reactive oxygen species (ROS) levels in the cytoplasm and mitochondria of sperm were assessed using computer-assisted sperm analysis (CASA) and flow cytometry. The treatment with H2O2 rapidly decreased sperm viability, and antimycin-induced mitochondrial ROS mainly decreased sperm motility; PQQ and vitamin C effectively reduced mitochondrial ROS, while ergothioneine and vitamin C reduced cytosolic ROS. In frozen-thawed sperm, oxidative stress was elevated in both cytoplasm and mitochondria, and all three antioxidants improved sperm motility by inhibiting ROS production. Furthermore, the localization of oxidized lipids (4-hydroxynonenal) in sperm was detected using immunofluorescence, indicating that oxidative stress affects the head and midpiece of sperm. These findings highlight the potential of targeted antioxidants to mitigate the detrimental effects of oxidative stress on bull sperm and provide valuable insights to improve semen quality and optimize the use of antioxidants in artificial insemination.