Search Results (1,073)

Oxidative stress is one of the few defined causes of male infertility affecting at least one third of patients attending infertility clinics. Human spermatozoa are vulnerable to this form of attack because their stripped-down architecture means that they possess limited antioxidant protection and little capacity for biochemical repair. They also compound their vulnerability by being active generators of reactive oxygen species (ROS) and possessing multiple substrates for oxidative damage. The major sources of ROS in these cells are their mitochondria, an L-amino acid oxidase (IL4I1) and a calcium-dependent NADPH oxidase (NOX5). Spermatozoa tolerate the risks associated with ROS generation because their biology is heavily dependent on redox regulation. ROS are important mediators of sperm capacitation, stimulating the generation of cAMP and prostaglandins, inhibiting protein phosphatases and encouraging removal of cholesterol from the plasma membrane. Furthermore, during fertilization, the ability of ROS to activate metalloproteinases facilitates penetration of the zona pellucida and sperm–oocyte fusion. While ROS are physiologically important for sperm function, the over-production of these metabolites can impair sperm function. Antioxidants have therefore assumed some importance as a possible therapy for the infertile male. However, before this potential can be realized, we need to optimize the composition and dose of reagents used in such formulations and develop improved methods of diagnosing oxidative stress within the patient population. Full article

Although the ATP-binding cassette (ABC) transporter superfamily of proteins typically facilitates the movement of compounds across cellular membranes, the ABC E subfamily (ABCE) influences protein synthesis via non-transporter roles in ribosome biogenesis. Despite this essential role, our understanding of the impact that ABCE proteins have on insect physiology is limited. Here, we identified and characterized the ABCE gene from Lygus hesperus, a major agricultural pest of crops in North America. LhABCE transcripts were constitutively expressed throughout development and were present in all adult tissues tested. RNA interference (RNAi)-mediated knockdown of LhABCE transcripts in fifth instar nymphs resulted in high nymphal mortality and an incomplete molt. LhABCE knockdown in adults disrupted gametogenesis and reduced longevity. In females, oogenesis was impaired and oocytes did not progress beyond the pre-vitellogenic phase. In males, LhABCE knockdown reduced both spermatozoa abundance and male fertility. LhABCE knockdown, however, had little to no impact on hemolymph protein levels or the levels of circulating vitellogenin. Taken together, the results indicate that LhABCE is critical for the normal progression of processes like molting and gametogenesis that require coordinated bursts of protein synthesis and suggest that ABCE may play an important role in the mechanisms underlying those bursts. Full article

Ambient heat exposure reduces male fertility in mammals with scrotal testes. Our previous work has demonstrated that some stallions are more susceptible to ambient heat-related subfertility than others, yet the mechanism for heat-induced subfertility remains uncertain, limiting both diagnosis and preventative measures. This study sought to define how the phenotype of stallions susceptible to heat-induced subfertility differs from that of more resilient animals, by measuring the systemic (blood plasma) and localized (reproductive tract) inflammatory and oxidative stress markers of sperm concentration, sperm motility assessments, total antioxidant capacity (TAC; in blood and seminal plasma), malondialdehyde (MDA; in blood and seminal plasma), oxidized guanine species (8-OH-2dG; in blood plasma and spermatozoa DNA), sperm DNA damage (assessed via Halo, SCSA (Sperm Chromatin Structure Assay) and CMA3 (Chromomycin A3)), and c-reactive protein (CRP; in blood plasma). Post-breeding dismount semen samples (n = 357) and blood plasma samples (n = 97) were collected from 31 stallions at commercial thoroughbred studs throughout one breeding season (NSW, Australia). A subset of stallions (16%) was deemed heat-induced subfertility-susceptible (HISS) stallions. These animals showed reduced seminal plasma antioxidant capacity, increased systemic and localized lipid peroxidation, and distinct systemic inflammatory response. Seminal antioxidant capacity was found to be strongly associated with impaired sperm motility (r = 0.739 * vs. r = −0.059). The plasma c-reactive protein of heat-susceptible stallions correlated to heat exposure (r = 0.597 *) and affected sperm motilities (r = −0.527 **, r = −0.434 *). Systemic oxidative DNA damage (8-OH-2dG) also increased following heat events (r = 0.862 ***) and correlated with fertility losses (FCP: r = −0.740 **, PCP: r = −0.603 *). Non-HISS stallions displayed greater variability in systemic antioxidant status and robust response following heat exposure (r = 0.307 *) and localized antioxidant capacity was more strongly correlated to systemic antioxidant capacity than in the heat-susceptible group (r = 0.897 *** vs. r = 0.482 **). We demonstrate that impaired antioxidant responses, altered redox balance and suppressed acute-phase inflammatory signalling are key features associated with heat-induced subfertility in stallions and highlight biomarkers that could be used to identify animals with heat-susceptible fertility. Full article

This study systematically elucidated the developmental characteristics and molecular regulatory mechanisms of the testis during the critical period of sexual maturation in Kazakh horses by combining histological observation of one- and two-year-old testicular tissues with transcriptomic sequencing. In the testes of one-year-old horses, no obvious lumen was observed, and the interior is mainly comprising supporting cells and spermatogonia on the basement membrane; in contrast, in the testes of two-year-old horses, the tubular lumen was complete with spermatogonia, spermatocytes, and spermatozoa, indicating that spermatogenic function had approached maturity. Transcriptome profiling identified 979 differentially expressed genes (DEGs), with 209 up-regulated genes, including CYP11A1 and CATSPER2, and 770 down-regulated genes, including CD9. Gene Ontology (GO) annotation indicated primary enrichment of DEGs in biological processes related to multicellular organism development, cell membrane composition, and ion binding. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed significant enrichment of DEGs in the calcium signaling pathway, cell adhesion molecules, and neuroactive ligand–receptor interaction, among other key pathways. Protein–protein interaction (PPI) network analysis further highlighted core genes, including TNF, CATSPER2, and CDH13. Validation by RT-qPCR confirmed the reliability of the RNA-Seq data. Our findings reveal the dynamics of testicular development in Kazakh horses through histological and molecular analyses, thereby providing a theoretical framework and candidate genes to further elucidate regulatory mechanisms and guide genetic improvement in reproductive traits. Full article

Background/Objectives: Round spermatid injection (ROSI) is considered an experimental “last resort” option for couples with severe male factor infertility when mature spermatozoa cannot be obtained. We aimed to identify which stage of the clinical chain most strongly constrains overall success in routine practice and to describe the observed safety signal. Methods: We conducted a retrospective single-center cohort study of 221 consecutive ROSI-evaluated cycles (2021–2024). Outcomes were analyzed using a chain-of-outcome framework with explicit denominators: cycle-level feasibility (≥1 injected oocyte), two pronuclei (2PN) formation per injected oocyte, blastocyst development per 2PN, transfer per blastocyst cycle, and clinical pregnancy per transfer and per initiated cycle. Exact (Clopper–Pearson) 95% confidence intervals (CIs) were reported. Results: ROSI feasibility was observed in 5 of 221 initiated cycles (2.3%; exact 95% CI 0.7–5.2). Among the five transfer procedures performed after successful progression through upstream stages, clinical pregnancy occurred in four (80.0%; exact 95% CI 28.4–99.5). At the initiated-cycle level, overall clinical pregnancy was 4 of 221 cycles (1.8%; exact 95% CI 0.5–4.6). Conclusions: The overall effectiveness of ROSI remained low at the initiated-cycle level because very few cycles reached procedural feasibility and early attrition remained substantial. Conditional downstream outcomes may appear favorable only among the rare cycles reaching fertilization and transfer, while safety inference remains highly imprecise due to small denominators. Because only five cycles reached feasibility, all downstream conditional estimates remained highly unstable and sensitive to single-case variation. Full article

Human infertility affects approximately 17.5% of the global population, with male factors accounting for nearly half of all cases. Identifying reliable molecular biomarkers is crucial for improving the diagnosis and assessment of male fertility. This study established and refined an untargeted high-performance liquid chromatography–electrospray ionization–tandem mass spectrometry (HPLC-ESI-MS/MS) protocol for a comprehensive lipidomic and metabolomic analysis of human spermatozoa, using only 1.25 million cells per sample. Compared with previous reports, our optimized method achieved an unparalleled level of analytical depth, identifying 473 lipid species and 955 structurally annotated metabolites. This corresponds to nearly a 7600-fold improvement in detection efficiency per cell compared with previously published approaches. Lipidomic analysis revealed that the most abundant lipid classes were glycerophospholipids (39%), cholesterol (20%) and fatty acids (19%), with cholesterol representing the single most abundant compound. This observation is consistent with the structural complexity of the sperm plasma membrane. Metabolomic profiling similarly identified glycerophospholipids (44%), eicosanoids (14%) and N-acyl amino acids (12%) as the major metabolite classes. The integration of lipidomic and metabolomic data highlighted functionally interconnected pathways related to membrane dynamics, energy metabolism, and hormone biosynthesis. Overall, this work establishes a robust, sensitive, and scalable analytical framework that enables the high-coverage molecular characterization of spermatozoa from limited sample material, laying the groundwork for future biomarker discovery and clinical applications in male infertility research. Full article

This study aimed to evaluate the impact of coenzyme Q10 (CoQ10) supplementation during in vitro maturation (IVM) and/or vitrification of immature dromedary camel oocytes on their subsequent in vitro developmental competence. Additionally, total antioxidant capacity (TAC) and malondialdehyde (MDA) concentrations were assessed in the IVM spent medium. In experiment 1, cumulus–oocyte complexes (COCs, n = 808) collected from slaughtered dromedary camel ovaries were cultured in IVM media supplemented with either 0, 25, 50, or 100 μM CoQ10 for 36 h. Matured oocytes were then fertilized in vitro with epididymal camel spermatozoa. Eighteen hours post-insemination (pi), presumptive zygotes were cultured in vitro for 7 days. In experiment 2, a total of 875 COCs were randomly allocated to one of four experimental groups, namely (a) Vit−/IVM− (control) group, where COCs were vitrified in vitrification solution (VS; 25% DMSO + 25% EG) and matured in IVM media without CoQ10 supplementation; (b) Vit+/IVM− group, in which COCs were vitrified in a VS supplemented with 50 µM CoQ10 and matured in IVM media without CoQ10 supplementation; (c) Vit−/IVM+ group, where COCs were vitrified in VS without CoQ10 supplementation and matured in IVM media supplemented with 50 µM CoQ10; and (d) Vit+/IVM+ group, where COCs were vitrified in VS and matured in IVM media, both supplemented with 50 µM CoQ10. Following vitrification and warming, oocyte viability was evaluated morphologically and by trypan blue staining. Viable oocytes were then matured, fertilized, and cultured in vitro. In experiment 3, TAC and MDA concentrations in the IVM spent media were analyzed. Results showed that 50 µM CoQ10 supplementation to IVM media enhanced cumulus expansion, nuclear maturation, cleavage, and blastocyst rates (experiment 1). Adding 50 µM CoQ10 to the VS enhanced (p ≤ 0.05) oocyte viability compared to those vitrified in CoQ10-free media. Cumulus cell expansion and nuclear maturation rates were higher (p ≤ 0.05) in Vit−/IVM+ than in Vit+/IVM+ and Vit−/IVM− groups. Furthermore, cleavage and blastocyst rates were the highest (p ≤ 0.05) in Vit−/IVM+ group (experiment 2). The concentrations of TCA were higher, and the concentrations of MDA were lower (p ≤ 0.05) in Vit−/IVM+ than in other groups (experiment 3). In conclusion, supplementation of CoQ10 in the maturation medium of vitrified–warmed immature dromedary camel oocytes may enhance their in vitro developmental competence. Full article

The recovery and cryopreservation of epididymal spermatozoa enable genetic preservation in male dogs that die unexpectedly or require castration; however, sperm collection is typically performed immediately after surgery, and artificial insemination is often surgical. This study aimed to evaluate whether epididymides stored at 4–5 °C for 24 h prior to sperm recovery retain fertilizing capacity after cryopreservation and whether pregnancy can be achieved using endoscopically guided transcervical intrauterine insemination. Testes and epididymides from an eight-month-old German Shepherd dog were stored in physiological saline at 4–5 °C for 24 h following castration. Spermatozoa were recovered from the cauda epididymis using single-incision aspiration, evaluated, frozen according to the Uppsala method, and stored in liquid nitrogen for two months. After thawing, 62 × 10⁶ progressive motile spermatozoa were inseminated once into a bitch in heat using transcervical endoscopic guidance. Pregnancy was confirmed by ultrasonography, and nine healthy puppies were delivered at term. These findings demonstrate that 24 h refrigerated storage of the epididymis does not impair post-thaw fertilizing ability and that non-surgical transcervical intrauterine insemination represents an effective alternative to surgical techniques for the use of frozen–thawed epididymal semen in dogs. Full article

Background: Cryopreservation induces oxidative stress, membrane disruption, and mitochondrial injury in spermatozoa, leading to impaired motility and fertility. Selenium, as an essential trace element, protects cells from oxidative damage through selenoproteins such as glutathione peroxidase 4 (GPX4), a critical enzyme that detoxifies lipid hydroperoxides and inhibits ferroptosis. This study investigated whether supplementation with L-selenomethionine (L-SeMet), an organic selenium source with superior bioavailability and lower toxicity than inorganic forms, could alleviate cryo-induced sperm injury by suppressing ferroptosis. Methods & Results: Dairy goat sperm were cryopreserved with 0, 2, 4, 6, 8, 10 μM L-SeMet. Supplementation with 6 μM L-SeMet significantly improved motility, membrane and acrosome integrity, and mitochondrial membrane potential. Biochemical assays showed reduced iron, ROS, and MDA levels, alongside increased ATP, SOD, and GSH contents. Proteomic analysis identified 148 differentially expressed proteins, including up-regulation of GPX4, FTH1, VDAC2, and VDAC3—core ferroptosis regulators. Metabolomic profiling further revealed enrichment in unsaturated fatty acid biosynthesis, amino acid metabolism, and the TCA cycle, pathways closely linked to ferroptosis regulation. Transmission electron microscopy confirmed that L-SeMet preserved mitochondrial ultrastructure. Mechanistically, L-SeMet mirrored the ferroptosis inhibitor N-acetyl-L-cysteine and reversed RSL3-induced oxidative damage. Western blotting verified activation of the NRF2–SLC7A11–GPX4 antioxidant axis and inhibition of KEAP1 expression. Conclusions: Collectively, these findings demonstrate that L-SeMet protects spermatozoa from cryo-induced injury by stabilizing redox homeostasis, maintaining mitochondrial function, and inhibiting ferroptosis. The results highlight ferroptosis as a critical mechanism of sperm cryodamage and identify L-SeMet as a promising metabolic intervention to enhance post-thaw sperm quality and fertility. Full article

Semen quality and fecal microbial composition were compared between native Oula rams reared on the Qinghai–Tibet Plateau and Hu sheep rams introduced from lowland regions. Semen quality was analyzed in eight adult Oula rams and eight Hu rams, and fecal microbial composition was assessed via 16S rRNA sequencing. Results indicated that sperm acrosome integrity was significantly higher in Hu sheep than in Oula sheep (p < 0.001); other semen parameters showed no significant differences. Significant differences were also observed in fecal microbial communities between the two breeds. Compared with Hu sheep, Oula sheep exhibited higher microbial abundance and diversity at the phylum level, particularly Campylobacterota, Euryarchaeota, Planctomycetota, Verrucomicrobiota, Myxococcota, and Deferibacterota (p < 0.05). At the genus level, Oula sheep had significantly higher abundances of Treponema, Campylobacter, Methanobrevibacter, UCG-009, Family_XIII_AD3011_group, [Eubacterium]nodatum group, Candidatus Soleaferrea, Akkermansia, and unidentified_Ruminococcaceae (p < 0.05). Correlation analysis indicated associations between sheep semen quality and the top 30 abundant fecal microbial genera. Six genera showed significant positive correlations with acrosome integrity rate, and eight genera exhibited significant negative correlations (p < 0.05). Two genera were correlated positively with plasma membrane integrity rate (p < 0.05). Prevotellaceae_UCG-004 was positively correlated with sperm motility and Progressive Motility spermatozoa proportion (p < 0.05); Ruminococcus showed a significant positive correlation with sperm linear motility and a significant negative correlation with acrosome integrity rate (p < 0.05). These findings indicate that the microbial groups enriched in Oula sheep fecal samples and exhibiting negative correlations with acrosome integrity—including Ruminococcus, Treponema, Akkermansia, and Euryarchaeota—are associated with sperm quality through physiological adaptation mechanisms specific to high-altitude environments. Full article

Male accessory gland inflammation (MAGI) can impair male fertility through inflammation-driven oxidative stress and direct sperm damage; nutraceutical approaches may be useful when antibiotics are not indicated. Here, we evaluated a 3-month treatment with a Graminex™-based dietary supplement (Deprox-HP) in twenty MAGI patients integrating conventional semen analysis and oxidative stress assessment with sperm proteomics before and after therapy. After treatment, total and progressive sperm motility increased significantly, whereas sperm concentration and sperm morphology showed a non-significant upward trend. Sperm lipid peroxidation decreased markedly, while the antioxidant capacity showed a non-significant increase. Analysis of the sperm proteome demonstrated a clear PRE–POST clustering, consistent with treatment-associated remodeling. POST samples showed upregulation of proteins linked to sperm motility, redox homeostasis, mitochondrial metabolism and membrane remodeling. Two pregnancies occurred during the treatment period; in both cases, lipid peroxidation decreased along with an increase of morphologically typical spermatozoa, and sperm proteomics showed a concordant post-treatment shift enriched in flagellar and mitochondrial respiratory/redox compartments. Moreover, we found a selective enrichment POST treatment in these two patients of TEX50, a crucial protein involved in acrosome/head-stability during epididymal transit. Overall, Deprox-HP was associated with reduced oxidative membrane damage and a coordinated sperm proteomic shift consistent with improved motility. Full article

Efficient liquid storage of turkey semen is critical for artificial insemination, but its use is limited by bacteriospermia and oxidative damage. This study evaluated the effects of gentamicin supplementation in Glutac and Sperm Motility Medium (SMM) on bacterial load and sperm quality after 2 and 24 h of liquid storage. Semen from turkeys (n = 40) was assessed for motility, viability, plasma membrane and acrosome integrity, mitochondrial and metabolic activity, oxidative profile, apoptosis, DNA integrity, and microbiological status. The sperm motility and kinematic parameters declined significantly after 24 h in all the groups. However, both extenders (particularly SMM) maintained significantly higher motility than the untreated control. Gentamicin further improved the motility, viability, and plasma membrane and acrosome integrity. The mitochondrial activity and mitochondrial membrane potential were significantly higher in the extender-treated groups than in the controls at 2 and 24 h, whereas the superoxide and total ROS production were significantly higher in the controls. The total antioxidant capacity declined markedly in the untreated controls, especially after 24 h. Gentamicin significantly reduced bacterial load, most effectively in SMM, and decreased DNA fragmentation compared with the untreated controls. In conclusion, gentamicin supplementation—particularly in SMM—reduces bacteriospermia and oxidative stress while preserving turkey sperm quality during liquid storage. Full article

The functional genomic mechanisms contributing to aging-associated decline in fertility in men remain insufficiently elucidated. This study investigated age-related alterations in the sperm metabolome of healthy fertile Arab men across three groups: young adult (21–30 years, n = 6), late adult (31–40 years, n = 7), and advanced age (41–51 years, n = 5). Metabolomics was performed using LC-MS/MS. Statistical/functional analyses were performed using MetaboAnalyst-Pro. A total of 380 metabolites were identified, of which 164 showed significant differences (p < 0.05) across age groups. Principal component analysis, partial least squares-discriminate (PLS-DA), and sparse PLS-DA consistently demonstrated distinct metabolomic clustering between young adult and advanced age groups. Notably, in the advanced-age spermatozoa, L-homocysteine was undetectable, while methyloctadecanoyl-CoA was uniquely abundant. Biomarker analysis identified 137 potential aging-sperm biomarkers (AUC = 1), including upregulated (e.g., pentadecanoyl-CoA, (3S)-3-hydroxylinoleoyl-CoA, CDP-DG(LTE4/20:4(8Z11Z14Z17Z)), uracil) and downregulated (e.g., (S)-hydroxyoctanoyl-CoA, DG(22:6/18:4), L-homocysteine, N-myristoyl serine) metabolites. These biomarkers participate in key sperm domains, including motility, energy metabolism, membrane remodeling, oxidative-stress regulation, and fertilization. In conclusion, advancing age disrupts sperm “metabolostasis” (metabolite homeostasis essential for normal function), compromising their physiological integrity and fertilization competence. The identified biomarkers offer promising targets for interventions to preserve sperm health and mitigate age-related fertility decline. Full article

Omega-3 polyunsaturated fatty acids (n-3 PUFAs) are recognized for their beneficial effects on male fertility. This study evaluated the protective effects of dietary n-3 PUFAs from extruded linseed, alone or combined with the alga Padina pavonica, against in vitro lipopolysaccharide (LPS)-induced sperm dysfunction in rabbits. Twelve bucks were fed for 60 days a control diet (CNT), a diet containing 5% extruded linseed (L), or 5% extruded linseed plus 0.2% P. pavonica extract (LPP). Ejaculates were exposed in vitro to increasing LPS concentrations (0, 400, and 600 µg/mL), and sperm motility was evaluated at 0, 1, 2, and 4 h using computer-assisted sperm analysis. LPS markedly impaired sperm motility in the CNT group, increasing the percentage of static spermatozoa (p < 0.001) and reducing sperm progressive motility (p < 0.001), with complete immobility observed at 600 µg/mL after 4 h. Conversely, sperm from L and LPP groups maintained significantly higher progressive motility, lower static sperm, and improved kinematic parameters throughout the LPS challenge (p < 0.05). Dietary n-3 PUFA supplementation also attenuated LPS-induced TLR4 activation and reduced lipid peroxidation, as indicated by lower seminal TBARS levels. No histological alterations were detected in the male reproductive tract. These findings indicate that n-3 PUFA supplementation, particularly linseed combined with algae, mitigates LPS-induced sperm dysfunction in vitro. Full article

Male infertility contributes to nearly half of infertile couples, with asthenozoospermia and oligoasthenoteratozoospermia as predominant factors. Despite advancements in sperm processing techniques, the outcomes remain limited in severe cases, particularly concerning motility, mitochondrial function, and DNA integrity. Platelet-rich plasma (PRP), an autologous concentrate rich in platelets (>1 × 10⁶/μL) and growth factors, has recently gained attention as an adjunctive therapy in andrology and assisted reproduction. This review systematically evaluated studies published between 2015 and 2025 investigating PRP use in sperm processing, including in vitro experiments, clinical trials, animal models, and mechanistic studies. PRP demonstrated concentration-dependent benefits, with 5% PRP yielding optimal improvements: motility increased by 15–30%, mitochondrial activity increased by up to 80% (p = 0.002), and oxidative stress was significantly reduced (p < 0.001). PRP’s effects on DNA integrity differ by application method: intratesticular administration improves spermatogenesis, producing sperm with reduced DNA fragmentation (~33% relative reduction after 3 months, p < 0.001), while in vitro supplementation provides limited protection against processing-induced damage. Mechanisms involve antioxidant action, mitochondrial protection via AMP-activated protein kinaseNuclear Factor kappa-light-chain-enhancer of activated B cells (AMPK/NF-κB) modulation, membrane stabilization, and the selective preservation of higher-quality spermatozoa. PRP shows consistent biological efficacy and safety but lacks methodological standardization. Fewer than 20% of studies report platelet concentrations, limiting reproducibility. Standardized protocols distinguishing leukocyte-poor from leukocyte-rich preparations and randomized trials focusing on live birth rates are recommended. This review proposes an eight-point characterization checklist for future research. Full article