Search Results (114)

F₄-Neuroprostanes (F₄-NeuroPs), oxidative metabolites of docosahexaenoic acid, act as bioactive lipid mediators enhancing sperm motility and induce capacitation-like changes in vitro. Their biological action is proposed to involve sperm ion channels, in particular ryanodine receptors (RyRs), which regulate intracellular calcium homeostasis. We evaluated the effects of dantrolene, a RyR inhibitor, on motility and vitality of a selected spermatozoa at different concentrations (10, 30, 50, 100 μM). Then sperm motility, acrosome integrity, and RyR localization following co-incubation with dantrolene (D50 or D100 μM) and 4-/10-F_4t-NeuroPs (7 ng) were investigated. Acrosomal status was assessed using Pisum sativum agglutinin (PSA) staining and RyR localization by immunofluorescence. D50 was identified as the minimum effective dose to induce significant reductions in sperm motility. F₄-NeuroPs significantly increased rapid progressive motility versus controls. Co-incubation with F₄-NeuroPs + D50 reduced rapid motility and increased in situ and circular movement. The acrosome staining appeared altered or absent to different percentages, and RyR localization was also seen in the midpiece. These findings suggested that F₄-NeuroPs enhance sperm motility via RyR-mediated pathways, as confirmed by dantrolene inhibition. Accordingly, our results underscore the physiological relevance of RyRs in sperm function and suggest new insights into lipid-based mechanisms regulating sperm motility. Full article

Polystyrene microplastics (PS-MPs) released in the environment reportedly affect the reproduction of various organisms, induced oxidative stress and apoptosis, resulting in altered sperm parameters. In this in vitro study, we tested the cytotoxicity and genotoxicity of PS-MPs by exposing human semen samples to PS-MPs levels (105 and 210 μg/mL) for 30–60–90 min. Semen parameters, genome stability, sperm DNA fragmentation (SDF) and reactive oxygen species (ROS) production were analyzed before and after exposure. Moreover, we also evaluated the expression level of spermatozoa-specific expressed genes essential for the fusion with oocyte (DCST1, DCST2, IZUMO1, SPACA6, SOF1, and TMEM95). After PS-MP exposure, semen concentration and morphology did not differ, while sperm vitality and motility decreased in a time-dependent manner. In addition, sperm agglutination was observed in the groups exposed to both PS-MPs concentrations tested. A time- and concentration-dependent reduction in genomic stability, as well as increased SDF and ROS production, was also observed. Moreover, all investigated transcripts were down-regulated after PS-MP exposure. Our results confirm the oxidative stress-mediated genotoxicity and cytotoxicity of PS-MPs on human spermatozoa. The sperm agglutination observed after treatment could be due to the aggregation of PS-MPs already adhered to the sperm membranes, hindering sperm movement and fertilizing capability. Interestingly, the downregulation of genes required for sperm–oocyte fusion, resulting from data on the in vitro experimental system, suggests that PS-MP exposure may have implications for sperm functionality. While these findings highlight potential mechanisms of sperm dysfunction, further investigations using in vivo models are needed to determine their broader biological implications. Possible environmental and working exposure to pollutants should be considered during the counselling for male infertility. Full article

The physiological functions of mammalian sperm, such as motility, hyperactivation, and capacitation, require substantial energy. This study investigates the effects of two classic cryopreservation extenders—TCG (tris-citrate-glucose) and LEY (lactose-egg yolk)—on the energy metabolism of frozen–thawed boar semen. By comparing the quality indicators, key metabolite levels, and the activities of critical enzymes involved in glycolysis and the tricarboxylic acid cycle, we aim to understand how these different semen extenders influence the spermatozoa vitality of frozen–thawed boar semen. Following thawing, the LEY-cryopreserved sperm demonstrated significantly elevated motility parameters (viability, VCL, VSL, and VAP) and enhanced plasma membrane and acrosomal integrity compared with the TCG group (p < 0.05), though both cryopreserved groups exhibited significantly reduced performance relative to fresh semen controls. Cryopreservation markedly reduced intracellular adenosine triphosphate (ATP), pyruvate, and acetyl coenzyme A (A-CoA) levels (fresh > LEY > TCG; p < 0.05). The LEY-preserved spermatozoa retained higher activities of glycolysis-related enzymes (phosphofructokinase, PFK; pyruvate kinase, PK) compared with the TCG group, which, in turn, showed elevated lactate dehydrogenase (LDH) activity. Critically, TCG-suppressed pyruvate dehydrogenase (PDH) activity (p < 0.05) coincided with diminished A-CoA, indicating impaired mitochondrial oxidative phosphorylation. These results demonstrate LEY’s superior preservation of motility and membrane stability but highlight cryodamage-induced energy metabolism dysregulation, particularly TCG’s disruption of the glycolysis–TCA cycle coordination essential for spermatozoa function. In conclusion, the choice of semen extender has a significant impact on the energy metabolism and overall quality of frozen–thawed semen, highlighting the importance of optimizing cryopreservation protocols for improved spermatozoa viability and functionality. Full article

Artificial insemination (AI) is essential in intensive pig production, which significantly depends on semen quality from boars selected for health, genetics, and fertility. While AI aims to improve productivity, larger litters often result in smaller and less resistant piglets. Beyond fertility and genetic traits, boars also influence offspring health. This study investigated the relationship between sperm parameters of highly fertile boars and both reproductive outcomes and piglet physiological indicators. Multivariate analysis revealed significant paternal effects on blood markers reflecting organ function, including those of the pancreas, liver, and kidneys, as well as on glucose homeostasis, lipid metabolism, oxidative stress, protein and carbohydrate metabolism, muscle contraction, and neural signaling. Notably, sperm velocity was correlated with mitochondrial function, which is crucial for sperm motility, capacitation, DNA integrity, and embryo development—factors likely linked to healthier, more resilient offspring. Boars transmitting superior sperm velocity, erythropoiesis efficiency, and oxygen transport capacities produced piglets with better glucose regulation, growth, and resistance to neonatal hypoglycemia. These findings underscore the broader impact of sperm quality on offspring vitality and suggest that advanced sperm analysis could improve boar selection and enable more effective, health-oriented breeding strategies. Full article

This study aimed to investigate the effects of adding different levels of punicalagin or oleuropein to TRIS diluent on the quality of frozen–thawed semen from Najdi rams. Semen was diluted using TRIS-based diluter with 15% egg yolk (control group); supplemented with 0.1, 0.5, or 1 mg/100 mL punicalagin (in Experiment 1); or supplemented with 1, 2.5, or 5 mg/100 mL oleuropein (in Experiment 2). The collected semen was evaluated and cryopreserved, with the motility and concentration of sperm assessed using a CASA system. The results showed that the total motile spermatozoa (TMS), percentage of progressive motile spermatozoa (PMS), curvilinear velocity (VCL), rectilinear velocity, average path velocity (VAP), linearity coefficient, straightness index, minor defects, and sperm vitality were higher in the 0.1 mg/100 mL punicalagin group (p < 0.05) than in other groups. HOST% post-thawing was significantly higher in all punicalagin groups compared to the control group (p < 0.001). The percentages of PMS, TMS, VCL, minor defects, and sperm vitality were higher in the 1 mg/100 mL oleuropein group (p < 0.05) than in other groups. Oleuropein supplementation at 5 mg/100 mL decreased VAP in cooled sperms, while all levels increased VAP post-thawing. HOST-positive sperms% post-thawing was higher in all oleuropein-treated groups than the control group (p < 0.001). Moreover, oleuropein nonsignificantly increased the acrosome integrity in cooled sperms, while higher studied concentrations of oleuropein (2.5 and 5 mg/100 mL) decreased the acrosome integrity in frozen sperms. In conclusion, adding punicalagin (0.1 mg/100 mL) or oleuropein (1 mg/100 mL) to TRIS diluent improved the quality of frozen–thawed semen from rams. Full article

Because male obesity may result in reproductive failure, we aimed to examine the possible links among body mass index (BMI), the waist–hip ratio (WHR), and basic semen parameters, the oxidation–reduction potential of semen, the total antioxidant capacity of seminal plasma, the ability of sperm to bind hyaluronic acid, and sperm DNA fragmentation (SDF). This study was performed on semen (n = 543) collected from volunteers classified as follows: normal weight (BMI 18.5–24.9 kg/m²), overweight (BMI 25.0–29.9 kg/m²), obese (BMI ≥ 30.0 kg/m²), with a normal WHR (<1) or abnormal WHR (≥1). No significant differences in standard semen parameters were found between men with a normal BMI and those with overweight/obesity. However, compared with overweight men, obese men had a higher SDF index prevalence and risk for an SDF index > 20%. Compared with men with WHR < 1, those with WHR ≥ 1 had significantly lower sperm motility, morphology, and vitality and an increased SDF index, prevalence and risk for an SDF index > 20%. In conclusion, abnormal WHR had a greater negative impact on conventional semen parameters than abnormal BMI. Both BMI ≥ 30.0 and WHR ≥ 1 negatively influenced sperm chromatin integrity only. Obesity is a potential risk factor for sperm DNA damage. Full article

This study explores the endocrine-disrupting effects of climbazole (CBZ), an environmental and lifestyle stressor, on male fertility. The impact of CBZ on sperm vitality, motility, and molecular pathways related to hormone receptors and apoptosis was evaluated, in non-capacitated and capacitated conditions. Gene expression of key components, including hormone receptors (ESR1, ESR2, FSHR, AR), apoptosis-related genes (BAX, BCL2), and COX4l1 (involved in mitochondrial function), was analyzed. Protein tyrosine phosphorylation, a marker of capacitation, was also examined using immunofluorescence and Western blot analysis. We demonstrated that CBZ significantly reduced sperm vitality at concentrations above 25 µM and motility at 1 and 10 µM in non-capacitated and capacitated conditions. Changes in tyrosine phosphorylation patterns were also observed. Gene expression analysis revealed an upregulation of ESR1, ESR2, FSHR, and BAX, while AR and COX4l1 expression were downregulated. These findings offer new insights into the potential endocrine-disrupting and cytotoxic effects of CBZ, highlighting its potential role in compromising male reproductive health. Full article

Feed additives are crucial in livestock production, enhancing performance, health, and reproductive efficiency. Recently, there has been a shift toward natural biomolecules as feed additives, specifically targeting improved reproductive outcomes and sperm quality. This transition arises from concerns about antibiotic misuse, antimicrobial resistance, and consumer preferences for eco-friendly products, along with the superior bioavailability, lower toxicity, and reduced environmental impact of natural biomolecules compared to synthetic alternatives. Collaboration among researchers, veterinarians, nutritionists, and regulators is essential to ensure safe and effective livestock management. The review explores advancements in using vital biomolecules in reproductive processes, including plant-derived bioactives such as phytochemicals and antioxidants. It investigates not only the mechanisms but also the intricate interactions of these compounds with animals’ hormonal and physiological systems. Additionally, the review critically assesses challenges and prospects related to incorporating natural biomolecules into livestock practices. The potential benefits include enhanced reproductive efficiency and improved sperm quality. However, successful implementation requires understanding factors like precise dosing, potential interactions, and long-term health impacts. Overall, this comprehensive review highlights recent research, technological strides, and the future potential of integrating natural biomolecules into animal diets. Full article

Infertility is increasingly recognized as being closely linked to obesity in humans. The successful production of fertile spermatozoa requires adequate spermatogenesis within the testis and proper spermatozoa maturation through the epididymis. This study aimed to evaluate the impact of body adiposity on male fertility, focusing on sperm parameters, epididymal sperm maturation, and sperm capacitation in Wistar rats. Male rats were randomized into three dietary groups over four weeks: a control group receiving less than 4% lard, regular chow, a 10% lard group, and a 60% lard group. Following dietary interventions, fertility tests were conducted across the groups. The epididymis was dissected into caput, corpus, and cauda regions to assess sperm concentration, vitality capacitation, carbohydrate distribution, tyrosine phosphorylation, and phosphatidylserine levels. Additionally, serum testosterone levels were measured to evaluate hormonal influences on fertility. The rats subjected to high-fat diets leading to overweight and obesity exhibited significant alterations in fertility. These changes were characterized by impaired epididymal sperm maturation, as evidenced by lower testosterone levels, decreased sperm viability, and capacitation. Furthermore, increased adiposity was associated with a lack of asymmetry in the plasma membrane, alteration in carbohydrate distribution, and changes in tyrosine phosphorylation. This study underscores the adverse effects of high-fat diets on male fertility, particularly through mechanisms affecting sperm maturation in the epididymis. The evidence suggests that obesity-induced alterations in sperm parameters and hormonal profiles may contribute to reduced fertility in male rats, which could have implications for understanding similar human processes. Full article

Mitochondria play a crucial role in sperm development; however, the mechanisms regulating their function in sperm remain poorly understood. Developing a method to regulate the expression of a target gene within the mitochondria of sperm is a vital step in this area of research. In this study, we aimed to create a system for expressing a transgene in the mitochondria of sperm. As a proof of concept, we generated transgenic mice that express green fluorescent protein (GFP) fused with a mitochondrial localization signal (MLS) driven by the phosphoglycerate kinase 2 (PGK2) promoter, which facilitates the transgene expression in the sperm. Although the PGK2 promoter has previously shown to drive gene expression in spermatocytes and spermatids, the novelty of our approach lies in the combination of PGK2-driven MLS-GFP expression to study mitochondria in vivo. We established two founder lines of transgenic mice through pronuclear microinjection, and MLS-GFP expression was confirmed in the mitochondria of sperm cells using fluorescence microscopy and flow cytometry. Consequently, we provide a novel platform for investigating mitochondrial function in sperm, where GFP can be substituted with other genes of interest to examine their effects on mitochondria. This system specifically targets sperm mitochondria, offering an innovative approach for studying mitochondrial function in vivo. Full article

Extracellular vesicles (EVs) are gaining recognition for their essential role in enhancing gamete quality and improving outcomes in assisted reproductive technologies. These nanosized particles, released by cells, carry proteins, lipids, and RNAs, facilitating critical cell communication and offering the potential to enhance gamete maturation and improve fertilization rates. Most research on males has concentrated on seminal plasma, a complex fluid produced by the testes and accessory glands vital in modulating sperm fertility potential. The components of seminal plasma significantly affect sperm functionality, embryo survival, and placental development, making this a prominent area of interest in reproductive biology. The EVs within seminal plasma contribute to maintaining sperm membrane stability, enhancing motility, and promoting capacitation, which may influence the female reproductive tract following mating. In females, EVs have been identified in both the follicular and uterine environments, where effective embryo–maternal communication is crucial. The oviduct epithelium supports gamete transport and early embryonic development, with EVs found in oviductal fluid playing a key role in reproductive processes. These EVs support the embryo’s growth in the nutrient-rich uterine environment. These important studies underscore the significant role of EVs in transporting essential molecular compounds to gametes and embryos, leading to an enhanced understanding and potential manipulation of reproductive processes. This review aims to summarize the current research on the benefits of EVs in gamete manipulation and embryo development, highlighting their promising implications for reproductive health. Full article

Peroxisome Proliferator-Activated Receptor Gamma (PPARγ) is a regulating agent in antioxidant response also involved in controlling inflammation. The impact of varicocele and urogenital infections on sperm PPARγ expression was studied. The PPARγ gene expression was investigated in spermatozoa of 26 normozoospermic men grouped according to their clinical conditions: normal semen parameters (N), normal semen parameters and varicocele (N + V), and normal semen parameters and urogenital infections (N + UI). Sperm PPARγ expression was correlated with F₂-isoprostanes (F₂-IsoPs), as markers of lipid peroxidation, and Resolvin D1 (RvD1), a pro-resolving mediator in inflammation. Sperm PPARγ expression was evaluated through comparative real-time PCR, and F₂-IsoPs and RvD1 were quantified in the seminal plasma via GC/NCI-MS/MS and immunoassay, respectively. PPARγ expression correlates positively with sperm morphology and vitality and negatively with F₂-IsoPs and RvD1. Sperm morphology positively correlates with vitality and negatively with F₂-IsoP and RvD1 levels. Despite the normozoospermia in the three examined groups, sperm morphology and PPARγ expression were significantly reduced in N + V and N + UI groups compared to the N group. Additionally, F₂-IsoP and RvD1 levels were elevated in N + V and N + UI patients. These data suggest that PPARγ expression is compromised by inflammation and lipoperoxidation, providing new insights to further explore new possibilities of targeted treatment of male infertility. Full article

Spermatogenesis is a complex process requiring precisely controlled metabolic adaptations. Although the genetic and cellular aspects of spermatogenesis have been extensively studied, the underlying metabolic mechanisms remain largely underexplored. In this study, we utilized STA-PUT technology to separate three key cell types involved in mouse spermatogenesis: pachytene spermatocytes (PAC), round spermatids (RS), and elongated spermatids (ES). A comprehensive untargeted metabolomic analysis revealed significant metabolic changes during spermatogenesis, such as reduced methylation-related metabolites and increased glycolytic intermediates and TCA cycle metabolites during ES. Moreover, metabolic differences between germ cells and somatic cells (Leydig and Sertoli cells) were highlighted, particularly in steroidogenesis and lipid metabolism. To investigate clinical relevance, we analyzed human seminal plasma. Samples from individuals with azoospermia displayed significant metabolic abnormalities, including reduced methionine, tryptophan, and arginine, which play vital roles in sperm development. Pathway enrichment analysis revealed disturbances in the metabolism of nucleotide, amino acid, and energy in azoospermia, suggesting potential biomarkers of male infertility. Our findings provide a comprehensive metabolic profile of spermatogenesis and suggest that metabolic alterations may be significant contributors to male infertility, particularly in cases of azoospermia. Full article

Sperm morphology assessment plays a vital role in semen analysis and the diagnosis of male infertility. By quantitatively analyzing the morphological characteristics of the sperm head, midpiece, and tail, it provides essential insights for assisted reproductive technologies (ARTs), such as in vitro fertilization (IVF). However, traditional manual evaluation methods not only rely on staining procedures that can damage the cells but also suffer from strong subjectivity and inconsistent results, underscoring the urgent need for an automated, accurate, and non-invasive method for multi-sperm morphology assessment. To address the limitations of existing techniques, this study proposes a novel method that combines a multi-scale part parsing network with a measurement accuracy enhancement strategy for non-stained sperm morphology analysis. First, a multi-scale part parsing network integrating semantic segmentation and instance segmentation is introduced to achieve instance-level parsing of sperm, enabling precise measurement of morphological parameters for each individual sperm instance. Second, to eliminate measurement errors caused by the reduced resolution of non-stained sperm images, a measurement accuracy enhancement method based on statistical analysis and signal processing is designed. This method employs an interquartile range (IQR) method to exclude outliers, Gaussian filtering to smooth data, and robust correction techniques to extract the maximum morphological features of sperm. Experimental results demonstrate that the proposed multi-scale part parsing network achieves 59.3% $A{P}_{\mathrm{vol}}^p$, surpassing the state-of-the-art AIParsing by 9.20%. Compared to evaluations based solely on segmentation results, the integration of the measurement accuracy enhancement strategy significantly reduces measurement errors, with the largest reduction in errors for head, midpiece, and tail measurements reaching up to 35.0%. Full article

Male reproductive health is governed by an intricate interplay of genetic, epigenetic, and environmental factors. Epigenetic mechanisms—encompassing DNA methylation, histone modifications, and non-coding RNA activity—are crucial both for spermatogenesis and sperm maturation. However, oxidative stress, driven by excessive reactive oxygen species, disrupts these processes, leading to impaired sperm function and male infertility. This disruption extends to epigenetic modifications, resulting in abnormal gene expression and chromatin remodeling that compromise genomic integrity and fertilization potential. Importantly, oxidative-stress-induced epigenetic alterations can be inherited, affecting the health and fertility of offspring and future generations. This review investigates how oxidative stress influences epigenetic regulation in male reproduction by modifying DNA methylation, histone modifications, and non-coding RNAs, ultimately compromising spermatogenesis. Additionally, it discusses the transgenerational implications of these epigenetic disruptions and their potential role in hereditary infertility and disease predisposition. Understanding these mechanisms is vital for developing therapeutic strategies that mitigate oxidative damage and restore epigenetic homeostasis in the male germline. By integrating insights from molecular, clinical, and transgenerational research, this work emphasizes the need for targeted interventions to enhance male reproductive health and prevent adverse outcomes in progeny. Furthermore, elucidating the dose–response relationships between oxidative stress and epigenetic changes remains a critical research priority, informing personalized diagnostics and therapeutic interventions. In this context, future studies should adopt standardized markers of oxidative damage, robust clinical trials, and multi-omic approaches to capture the complexity of epigenetic regulation in spermatogenesis. Such rigorous investigations will ultimately reduce the risk of transgenerational disorders and optimize reproductive health outcomes. Full article