Sperm Oxidative Damage Research Articles

Non-thermal plasma promotes boar sperm quality through increasing AMPK methylation

Boar sperm quality, as an important indicator of reproductive efficiency, directly affects the efficiency of livestock production. Here, this study was conducted to improve the boar sperm quality by using a non-thermal dielectric barrier discharge (DBD) plasma. Our results showed that DBD plasma exposure at 2.1 W for 15 s could improve boar sperm quality by increasing exon methylation level of adenosine monophosphate-activated protein kinase (AMPK) and thus improving the glycolytic flux, mitochondrial function, and antioxidant capacity without damaging the integrity of sperm DNA and acrosome. In addition, DBD plasma could rescue DNA methyltransferase inhibitor decitabine-caused low sperm quality through reducing the oxidative stress and mitochondrial damage. Therefore, the application of non-thermal plasma provides a new strategy for reducing sperm oxidative damage and improving sperm quality, which shows a great potential in assisted reproduction to solve the problem of male infertility.

Effects of Astragalus polysaccharide to boar sperm on bisphenol A exposure.

Bisphenol A (BPA) is reported to impair male fertility. The alleviating effect of Astragalus polysaccharide (APS) on sperm oxidative damage caused by BPA exposure was analysed for the first time. In this study, the effect of APS (0.25, 0.5, 0.75, 1mg/mL) on motility of BPA-exposed sperm, energy metabolism indexes, and antioxidant parameters was evaluated. In addition, the effects of APS supplementation on protein tyrosine phosphorylation of BPA-exposed sperm were assessed. The results showed that the addition of APS (0.5 and 0.75 mg/mL) significantly increased motility of BPA-exposed sperm by decreasing the content of malondialdehyde and improving activities of superoxide dismutase and catalase (p < .05). Administration of different doses of APS to BPA-exposed sperm improved mitochondrial membrane potential and energy production (p < .05). Moreover, APS protected and alleviated tyrosine phosphorylation protein on the principal-pieces of BPA-exposed sperm flagella. In conclusion, supplementation with APS enhanced the antioxidant capacity of BPA-exposed sperm and improved in vitro capacitation, thereby improving the reproductive capacity of sperm exposed to environmental hormones.

In Vitro Combination of Ascorbic and Ellagic Acids in Sperm Oxidative Damage Inhibition

It is known that an altered redox balance interferes with normal spermatic functions. Exposure to genotoxic substances capable of producing oxidative stress (OS) can cause infertility in humans. The use of antioxidants to reduce oxidative stress contributes to the improvement in reproductive function. This study focused on an antigenotoxic evaluation of ellagic acid (EA) and ascorbic acid (AA) in combination against benzene genotoxic action on human spermatozoa in vitro. In addition to the evaluation of sperm parameters, damage in sperm genetic material and intracellular ROS quantification were assessed after AA, EA and benzene co-exposure using the TUNEL technique and DCF assay. The results showed that the combination of the two antioxidants generates a greater time-dependent antigenotoxic action, reducing both the sperm DNA fragmentation index and the oxidative stress. The genoprotective effect of AA and EA association in sperm cells lays the foundations for a more in-depth clinical study on the use of antioxidants as a therapy for male infertility.

Sperm oxidative status varies with the level of sperm competition and affects male reproductive success

Sperm competition theory predicts that males should strategically adjust the allocation of sperm in response to the level of sperm competition (i.e. the number of competing ejaculates). Predictions of sperm competition models have mostly focused on sperm numbers. However, whether males strategically adjust other important traits of their ejaculate such as sperm antioxidants in response to potential rivals and the consequences of such allocation strategies on sperm integrity (e.g. oxidative damage accumulation) and offspring viability is poorly investigated. Using an experimental design in which I manipulated the perceived level of sperm competition experienced by male field crickets, Gryllus bimaculatus , I show that besides sperm numbers, sperm oxidative status can also vary in response to the level of sperm competition. Male crickets increased the antioxidant content of their ejaculates in response to sperm competition risk, an effect that presumably led to less sperm oxidative damage, at least when the level of competition was low. Moreover, I provide evidence suggesting that such strategic adjustments can have fitness consequences for males, as eggs sired by males with increased sperm damage had lower viability. Overall, my results suggest that sperm competition drives plasticity in nonsperm components affecting sperm oxidative status and support the idea that oxidative stress may play a key role in postcopulatory sexual selection. • Male crickets adjusted sperm numbers as expected by sperm competition theory. • Males also increased sperm antioxidants in response to other potential rivals. • Males under a low level of sperm competition had reduced sperm oxidative damage. • Eggs fertilized by males with reduced sperm oxidative damage were more viable.

Vitamin E Delivery Systems Increase Resistance to Oxidative Stress in Red Deer Sperm Cells: Hydrogel and Nanoemulsion Carriers.

Oxidative stress has become a major concern in the field of spermatology, and one of the possible solutions to this acute problem would be the use of antioxidant protection; however, more studies are required in this field, as highly contradictory results regarding the addition of antioxidants have been obtained. Vitamin E is a powerful biological antioxidant, but its low stability and high hydrophobicity limit its application in spermatology, making the use of organic solvents necessary, which renders spermatozoa practically motionless. Keeping this in mind, we propose the use of hydrogels (HVEs) and nanoemulsions (NVEs), alone or in combination, as carriers for the controlled release of vitamin E, thus, improving its solubility and stability and preventing oxidative stress in sperm cells. Cryopreserved sperm from six stags was thawed and extended to 30 × 106 sperm/mL in Bovine Gamete Medium (BGM). Once aliquoted, the samples were incubated as follows: control, free vitamin E (1 mM), NVEs (9 mM), HVEs (1 mM), and the combination of HVEs and NVEs (H + N), with or without induced oxidative stress (100 µM Fe2+/ascorbate). The different treatments were analyzed after 0, 2, 5, and 24 h of incubation at 37 °C. Motility (CASA®), viability (YO-PRO-1/IP), mitochondrial membrane potential (Mitotracker Deep Red 633), lipid peroxidation (C11 BODIPY 581/591), intracellular reactive oxygen species production (CM-H2DCFDA), and DNA status (SCSA®) were assessed. Our results show that the deleterious effects of exogenous oxidative stress were prevented by the vitamin E-loaded carriers proposed, while the kinematic sperm parameters (p ˂ 0.05) and sperm viability were always preserved. Moreover, the vitamin E formulations maintained and preserved mitochondrial activity, prevented sperm lipid peroxidation, and decreased reactive oxygen species (ROS) production (p ˂ 0.05) under oxidative stress conditions. Vitamin E formulations were significantly different as regards the free vitamin E samples (p < 0.001), whose sperm kinematic parameters drastically decreased. This is the first time that vitamin E has been formulated as hydrogels. This new formulation could be highly relevant for sperm physiology preservation, signifying an excellent approach against sperm oxidative damage.

Mixture Analysis of Associations between Occupational Exposure to Polycyclic Aromatic Hydrocarbons and Sperm Oxidative DNA Damage.

This study aimed to determine (i) associations between levels of the polycyclic aromatic hydrocarbon (PAH) mixture with 16 targeted PAH compounds in the personal breathing zone area and sperm oxidative DNA damage, (ii) associations between levels of individual PAH compounds and sperm oxidative DNA damage, (iii) oxidative stress as the mode of action for the genotoxic effects on sperm, and (iv) any dose-response relationship between exposure to the PAH mixture and/or individual PAH compounds and sperm oxidative DNA damage. Sixteen targeted PAH compounds in the personal breathing zone area of 38 coke-oven workers and 24 control subjects were quantified using gas chromatography-mass spectrometry. Sperm oxidative damage and status were evaluated by measuring levels of sperm 7,8-dihydro-8-oxoguanie (8-oxodGuo), seminal malondialdehyde (MDA) and seminal reactive oxygen species (ROS). Bayesian kernel machine regression with hierarchical variable selection process was employed to determine associations of the PAH mixture and the biomarkers of sperm oxidative damage. A novel grouping approach needed for the hierarchical variable selection process was developed based on PAH bay region and molecular weight. The PAH mixture exhibited a positive trend with increased sperm 8-oxodGuo levels at their lower percentiles (25th-50th). The exposure of the PAH mixture was associated with increased MDA levels in sperm. Bay and bay-like regions of the PAH mixture were the most important group for estimating the associations between the PAH mixture and sperm oxidative stress status. Benzo[a]anthracene was the main individual PAH compound that was associated with increased MDA levels. Sperm oxidative DNA damage induced by occupational exposure to the PAH mixture had a suggestive association with increased MDA levels in coke-oven workers. Finally, the study identified that the individual PAH compound, benzo[a]anthracene, was the primary driver for the suggestive association between the PAH mixture and sperm oxidative damage.

Influences of contamination of Aeromonas hydrophila, on quality, oxidative damage, and ultrastructure in cryopreserved sperm of the silver barb, Barbonymus gonionotus

This study investigated the effects of pathogenic bacteria, Aeromonas hydrophila, contamination on the changes in sperm oxidative damage, ultrastructure, and quality in the silver barb (Barbonymus gonionotus) following cryopreservation. Suspensions of silver barb sperm were in vitro inoculated with A. hydrophila (0, 103, and 105 CFU/mL; named as control, T1, and T2, respectively) and incubated at 25 ± 1 °C for 5 min, before equilibration with 10% DMSO. Sperm samples were frozen and cryostored within a dry shipper at the upper and lower chamber, respectively. The number of A. hydrophila incubated with sperm caused sperm quality decrease and was inversely correlated with unfrozen sperm motility (r = −0.5987) and viability (r = −0.5277) and post-thaw sperm motility (r = −0.5710) and viability (r = −0.7573). The lipid peroxidation (LPO) levels were significantly (p < 0.05) affected by the in vitro inoculation of A. hydrophila and freezing condition (unfrozen or frozen). Significant (p < 0.05) inverse correlations were observed between LPO levels and sperm quality (motility and viability) of silver barb sperm. Ultrastructural damage of silver barb sperm (plasma membrane, mitochondria, flagella, and nucleus) was significantly (p < 0.05) affected by the freezing-thawing process and A. hydrophila contamination. In addition, the inoculation of A. hydrophila and cryopreservation process caused the alteration of sperm morphology at the ultrastructure level including swelling and rupture of the plasma membrane, vesicular to swollen-mitochondria, tail coiling, disarranged flagellum, chromatin condensation, and nuclear fragmentation. Our study revealed that a high level of A. hydrophila contamination and freezing-thawing process during cryopreservation process can activate the necrosis and apoptosis-like phenomenon and autophagy pathway in silver barb sperm.

Sperm DNA damage and seminal antioxidant activity in subfertile men.

Supraphysiological ROS levels can lead to apoptosis, lipid peroxidation, and DNA and protein damage. This pilot study aimed to investigate the sperm oxidative damage in subfertile men, to describe the relationship between the antioxidant system and ROS. Sixty-four semen samples were categorised according to the evaluated routine parameters (WHO, WHO laboratory manual for the examination and processing of human semen, 2010). Results were cross-referenced with the DNA damage [Comet (n=53) and TUNEL (n=49) assays], antioxidant enzyme activity [SOD (n=51), CAT (n=48) and GST (n=48)], and content of total thiols (n=36), lipid hydroperoxides (n=35) and MDA (n=31). Compared to pathospermic samples, normozoospermic presented 40%-45% fewer spermatozoa with fragmented DNA, 19% fewer hydroperoxides, and slightly higher total thiols and MDA levels. Asthenozoospermic/asthenoteratozoospermic samples had the lowest GST activity. SOD and CAT showed a similar trend. Our results evidenced significant positive correlations between DNA damage and immotile spermatozoa; SOD and CAT, GST and total thiols; CAT and GST; total thiols and sperm concentration; and MDA levels and head/midpiece abnormalities and hydroperoxides. This work contributes to the existing body of knowledge by showing that the oxidative status correlates with the classic sperm analysis parameters. Oxidative stress and DNA damage evaluation might be a valuable diagnostic and prognostic tool in cases of idiopathic male subfertility.

Nanotechnology in reproduction: Vitamin E nanoemulsions for reducing oxidative stress in sperm cells

Vitamin E is considered a powerful biological antioxidant; however, its characteristics such as high hydrophobicity and low stability limit its application. We propose to use nanotechnology as an innovative tool in spermatology, formulating nanoemulsions (NE) that accommodate vitamin E, protecting it from oxidation and promoting its release into the medium. The protective effect of the NE against oxidative stress was assessed in red deer epididymal sperm incubated at 37 °C. Cryopreserved sperm from eleven stags were thawed and extended to 400 × 106 sperm/ml in Bovine Gamete Medium (BGM). Once aliquoted, the samples were supplemented with the NE at different concentrations (0, 6 and 12 mM), with or without induced oxidative stress (100 μM Fe2+/ascorbate). The samples were evaluated after 0, 2 and 4 h of incubation at 37 °C. Motility (CASA), viability, mitochondrial membrane potential, acrosomal status, lipoperoxidation (C11 BODIPY 581/591), intracellular reactive oxygen species (ROS) production and DNA status (SCSA®) were assessed. After 2 and 4 h of incubation, the NE were able to prevent the deleterious effects of oxidative stress, thus improving total and progression motility (P ˂0.05). Moreover, the highest concentration tested (12 mM) improved almost every sperm kinematic variable (P ˂0.05) and preserved sperm viability in samples subjected to oxidative stress. In addition, 12 mM of NE protected the acrosomes integrity, maintained and protected mitochondrial activity, prevented sperm lipoperoxidation and reduced ROS production (P ˂0.05) in samples subjected to oxidative stress. This work indicates for the first time that vitamin E formulated in NE could be a new approach against sperm oxidative damage. This could be highly relevant for sperm physiology preservation in the context of assisted reproduction techniques.

Use of antioxidant could ameliorate the negative impact of etoposide on human sperm DNA during chemotherapy.

A previous study showed that N-acetylcysteine (NAC), used after in-vitro exposure to the gonadotoxic chemotherapeutic drug etoposide, has the ability to decrease DNA damage in human spermatozoa; however, it showed no benefit when used before exposure. This study aimed to evaluate the impact of the NAC on the preservation of sperm quality during in-vitro exposure to etoposide. Twenty semen samples were submitted to four experimental conditions: control, NAC-only incubation, etoposide-only incubation, and concomitant etoposide and NAC incubation. After in-vitro incubation, semen parameters, sperm chromatin condensation, sperm DNA fragmentation, sperm oxidative stress and sperm metabolism were used to evaluate the role of NAC in protecting human spermatozoa from etoposide. Etoposide did not affect semen parameters, nor did it cause sperm oxidative damage or alterations in glycolytic profile. However, it induced chromatin decondensation and DNA fragmentation, which were fully prevented by NAC. NAC was able to protect sperm DNA integrity during etoposide treatment in vitro, suggesting that NAC may be useful as an adjuvant agent in preserving male fertility during chemotherapy treatments.

Semen quality and sperm DNA damage associa –revised – final-finalted with oxidative stress in relation to exposure to polycyclic aromatic hydrocarbons

The cross-sectional study aimed to assess whether oxidative stress induced by exposure to polycyclic aromatic hydrocarbons (PAHs) affect semen quality and sperm DNA integrity. A total of 106 who met the inclusion criteria, were recruited from a coke-oven plant during their annual health checkup. The human subjects were grouped into the high exposure group, the low exposure group and the control based on PAH concentrations surrounding their employment locations. Semen quality, oxidative stress status, and sperm DNA damage [DNA fragmentation and 8-hydroxy-2′-deoxyguanosine (8-oxodGuo)] were assessed. Urinary 1-hydroxypyren (1-OHP) was used to assess human subject exposure to PAHs. The high exposure group experienced significantly lower sperm motility and normal morphology than the control (P = 0.046 and 0.049, respectively). The high exposure group also had significantly higher 8-oxoGuo concentrations in sperm than the control (P = 0.027). Urinary 1-OHP concentration was associated with decreased motility and less normal morphology, along with increased sperm oxidative damage and ROS concentrations. Oxidative stress induced by exposure to PAHs was associated with decreased sperm quality.

Peroxiredoxin 6 is the primary antioxidant enzyme for the maintenance of viability and DNA integrity in human spermatozoa.

Are all components of the peroxiredoxins (PRDXs) system important to control the levels of reactive oxygen species (ROS) to maintain viability and DNA integrity in spermatozoa? PRDX6 is the primary player of the PRDXs system for maintaining viability and DNA integrity in human spermatozoa. Mammalian spermatozoa are sensitive to high levels of ROS and PRDXs are antioxidant enzymes proven to control the levels of ROS generated during sperm capacitation to avoid oxidative damage in the spermatozoon. Low amounts of PRDXs are associated with male infertility. The absence of PRDX6 promotes sperm oxidative damage and infertility in mice. Semen samples were obtained over a period of one year from a cohort of 20 healthy non-smoking volunteers aged 22-30 years old. Sperm from healthy donors was incubated for 2 h in the absence or presence of inhibitors for the 2-Cys PRDXs system (peroxidase, reactivation system and NADPH-enzymes suppliers) or the 1-Cys PRDX system (peroxidase and calcium independent-phospholipase A2 (Ca2+-iPLA2) activity). Sperm viability, DNA oxidation, ROS levels, mitochondrial membrane potential and 4-hydroxynonenal production were determined by flow cytometry. We observed a significant decrease in viable cells due to inhibitors of the 2-Cys PRDXs, PRDX6 Ca2+-iPLA2 activity or the PRDX reactivation system compared to controls (P ≤ 0.05). PRDX6 Ca2+-iPLA2 activity inhibition had the strongest detrimental effect on sperm viability and DNA oxidation compared to controls (P ≤ 0.05). The 2-Cys PRDXs did not compensate for the inhibition of PRDX6 peroxidase and Ca2+-iPLA2 activities. Not applicable. Players of the reactivation systems may differ among mammalian species. The Ca2+-iPLA2 activity of PRDX6 is the most important and first line of defense against oxidative stress in human spermatozoa. Peroxynitrite is scavenged mainly by the PRDX6 peroxidase activity. These findings can help to design new diagnostic tools and therapies for male infertility. This research was supported by The Canadian Institutes of Health Research (MOP 133661 to C.O.), and by RI MUHC-Desjardins Studentship in Child Health Research awarded to M.C.F. The authors have nothing to disclose.

The Comet assay for detection of DNA damage in canine sperm.

Sperm DNA integrity is a fundamental prerequisite in fertilization and embryo development. Among DNA integrity tests, the Comet assay is an accurate and sensitive test for the detection of sperm oxidative damage. The aim of this work was to evaluate sperm oxidative damage using the Comet assay and to study the correlation between Comet and routine assays for the evaluation of semen quality. Dogs were divided in two groups: group A (n=6), comprising dogs with abnormal spermiogram, that is astheno-, terato- or oligoasthenoteratozoospermic (OAT); and group B (n=8), comprising normospermic dogs. The distribution of sperm oxidative damage was significantly different between the two groups (p=.001): group A-median: 31.55%, interquartile range (IQR): 30.18-38.01; group B-median: 0.90%, IQR: 0.65-1.96. The correlation between oxidative damage and abnormal morphology was high (r=.846; p<.001). There was a negative correlation between progressive motility and oxidative damage (r=-.792; p=.001). Basal and oxidative DNA damage of spermatozoa are increased in dogs with non-normospermic semen. In conclusion, and considering the elevated correlation with classical tests of sperm quality, the Comet assay has ample potential for clinical and research purposes in dogs.

Modification of sperm quality after sexual abstinence in Seba's short-tailed bat, Carollia perspicillata.

In polygynous mating systems, few males have stable access to sexual mates. With an expected higher copulation rate, harem males may deplete seminal fluids or increase epididymal sperm maturation, generating poor sperm quality. In a first study, we reported a higher sperm quality in sneaker males of Carollia perspicillata To test whether the lower sperm quality observed in harem males was generated by an elevated copulation rate, we temporarily removed males of both social statuses from the colony. We thus assessed status-related changes of sperm quality resulting from sexual abstinence. Moreover, released from territory and female guarding, harem males were expected to show a reduction in somatic costs. On the basis of sperm competition models, we predicted a higher resource investment in the ejaculate with the reduction of pre-copulatory efforts. In line with our predictions, sperm quality of harem males improved significantly in contrast to sneaker males, whose sperm quality did not change. Without an increase in ejaculate lipid peroxidation, our results also provide evidence that the duration of sexual abstinence was not sufficient to generate sperm oxidative damage through senescence. Harem males did not show a reduction in blood lipid peroxidation or in the ratio of oxidized to reduced glutathione. In line with the maintenance of these somatic costs, harem males did not invest more superoxide dismutase to the ejaculate to maintain sperm quality. Our results suggest that a difference in copulation rate rather than an adaptation to sperm competition provides sneaker males with higher sperm quality in C. perspicillata.

Oxidative stress at different stages of two-step semen cryopreservation procedures in dogs

Sperm cryopreservation generates sperm damage and reduced fertilization capacity as a consequence of reactive oxygen species formation. Identifying the critical points of the process will contribute to the development of strategies for oxidative stress prevention. Therefore, the aim of this experiment was to verify the occurrence of oxidative stress during the two-step cryopreservation process in dogs. Six healthy mature dogs were used and submitted to the two-step sperm cryopreservation protocol. The sperm analysis was done at three time points: after refrigeration, after glycerolization, and after thawing by sperm motility, measurement of spontaneous and induced oxidative stress, sperm mitochondrial activity, plasma membrane integrity, flow cytometric evaluation of plasma and acrosome membrane integrity, mitochondrial membrane potential, and sperm chromatin structure assay. There was an increase in free radical production after glycerolization (87.4 ± 15.5 ng/mL of spontaneous thiobarbituric acid reactive substances (TBARS) after refrigeration and 1226.3 ± 256.0 ng/mL after glycerolization; P < 0.05), in association with loss of sperm mitochondrial activity. However, frozen-thawed samples had lower sperm motility, lower resistance to oxidative stress (448.7 ± 23.6 ng/mL of induced TBARS after glycerolization and 609.4 ± 35.9 ng/mL after thawing; P < 0.05) and increased lipid peroxidation (4815.2 ± 335.4 ng/mL of spontaneous TBARS after thawing; P < 0.05) as well as increased damage to plasma and acrosomal membranes, compared with refrigeration and glycerolization. In conclusion, the production of free radicals by sperm cells begins during glycerolization. However, sperm oxidative damage intensifies after thawing. Despite intracellular ice formation during cryopreservation, the increased production of reactive oxygen species can be the explanation of the decrease in sperm motility, reduced mitochondrial activity, and sperm plasma membrane and acrosomal damage.

Sperm functional parameters and erythrocytes oxidant–antioxidant imbalance during municipal landfill leachate treatment withdrawal in rats

Adequate information on how leachates affect hematological and reproductive functions is necessary to help in linking causality with predictable response. The present study investigated the effects of Olushosun municipal landfill leachate (OMLL) exposure and withdrawal on sperm characteristics and erythrocytes oxidant–antioxidant balance in rats. Adult male Wistar rats were exposed to 0%, 12.5% and 25% OMLL in drinking water for 28 days. One-half of the rats in each group were sacrificed on day 29 while the remaining one-half stayed an additional 28 days without treatment. OMLL exposure significantly decreased sperm functional parameters, disrupted antioxidant systems with concomitant elevation in hydrogen peroxide and malondialdehyde levels in erythrocytes and sperm. Following withdrawal of treatment, OMLL-mediated decrease in sperm count and daily sperm production were reversed to near control. However, erythrocytes and sperm oxidative damage, increased sperm abnormalities, decreased epididymis weight, sperm progressive motility and testicular sperm number persisted and were consistent with results obtained from rats sacrificed immediately after OMLL treatment. Collectively, OMLL-induced irreversible oxidative damage to erythrocytes and sperm in rats within the time course of investigation. These findings highlight potential adverse effects of OMLL on individuals unduly exposed to leachates contaminated substances.

Females become infertile as the stored sperm's oxygen radicals increase

Predicting infertility is central to reproductive biology, medicine and evolutionary biology. In-vitro studies suggest that oxidative sperm damage causes infertility. Oxidative sperm damage can be reduced via two fundamental pathways: the removal of oxygen radicals by antioxidants, or the interference with cell metabolism to reduce the formation of oxygen radicals. Oxidative damage protection of spermatozoa should evolve frequently, especially during female sperm storage. However, in-vivo evidence linking oxidative protection and fertility is rare. We show that the intra-sperm production rate of oxygen radicals and the sperm metabolic rate were reduced in female bedbugs, Cimex lectularius, compared to males, and females laid fertile eggs. Females became infertile when sperm oxygen radicals and sperm metabolic rate increased to male levels. Our results link female fitness to sublethal sperm damage, imply adaptive benefits of interfering with sperm metabolism and offer the hypothesis that polyandry may serve to replace low-quality sperm.

Effects of reactive oxygen species from activated leucocytes on human sperm motility, viability and morphology

The accumulated data suggest that inflammation can increase the level of reactive oxygen species (ROS), which contribute to impaired sperm function and male infertility. Therefore, we propose that inflammation-mediated production of ROS in male and female reproductive tracts hinder sperm fertilisation. To test this hypothesis, phorbol myristate acetate (PMA) with polymorphonuclear leucocytes (PMNs) was applied to generate endogenous ROS. We evaluated the time-dependent effects of ROS on human sperm motility, viability and mitochondrial membrane potential (MMP). The results showed that after treatment with PMA and PMNs, the motility of human spermatozoa significantly decreased to 50% on Day 1 and 15% on Day 4 compared with that of the, respectively, negative controls (P = 0.012). The viability of human spermatozoa decreased on Day 4 of PMA + PMNs treatment (P = 0.028). The MMP of human spermatozoa significantly decreased from Day 2 to Day 4 in the PMA + PMN group compared with that of the controls (P = 0.019). Taken together, the 4-day cultivation approach provided an accurate evaluation of sperm quality, especially sperm motility and MMP. Our findings indicated that endogenous inflammation increased ROS levels, which might induce sperm oxidative damage. Additionally, sperm motility might be one of the earliest and most sensitive indicators of this damage.

Immune activation reduces sperm quality in the great tit.

Mounting an immune response against pathogens incurs costs to organisms by its effects on important life-history traits, such as reproductive investment and survival. As shown recently, immune activation produces large amounts of reactive species and is suggested to induce oxidative stress. Sperm are highly susceptible to oxidative stress, which can negatively impact sperm function and ultimately male fertilizing efficiency. Here we address the question as to whether mounting an immune response affects sperm quality through the damaging effects of oxidative stress. It has been demonstrated recently in birds that carotenoid-based ornaments can be reliable signals of a male's ability to protect sperm from oxidative damage. In a full-factorial design, we immune-challenged great tit males while simultaneously increasing their vitamin E availability, and assessed the effect on sperm quality and oxidative damage. We conducted this experiment in a natural population and tested the males' response to the experimental treatment in relation to their carotenoid-based breast coloration, a condition-dependent trait. Immune activation induced a steeper decline in sperm swimming velocity, thus highlighting the potential costs of an induced immune response on sperm competitive ability and fertilizing efficiency. We found sperm oxidative damage to be negatively correlated with sperm swimming velocity. However, blood resistance to a free-radical attack (a measure of somatic antioxidant capacity) as well as plasma and sperm levels of oxidative damage (lipid peroxidation) remained unaffected, thus suggesting that the observed effect did not arise through oxidative stress. Towards the end of their breeding cycle, swimming velocity of sperm of more intensely colored males was higher, which has important implications for the evolution of mate choice and multiple mating in females because females may accrue both direct and indirect benefits by mating with males having better quality sperm.

Protective effect of melatonin on sperm motility parameters on liquid storage of ram semen at 5°C

Oxidative damage to sperm resulting from reactive oxygen species generated by the cellular components of semen during liquid storage is possibly one of the main causes of the decline in motility during storage. The aim of this study was to determine the effects of melatonin as an antioxidant on sperm motility parameters in liquid storage of ram semen at 5°C. A total number of 24 ejaculates were collected using the artificial vagina from four chal rams and three replicates of the ejaculates were diluted with a Tris-based extender with melatonin at the doses of 0.1, 1 and 3 mM and an extender containing no melatonin (control). Diluted semen samples were cooled from 37 to 5°C. A computerassisted sperm motility analysis was used to analyze sperm motion characteristics. The sperm motility parameters such as progressive motility, straight linear velocity (VSL), curvilinear velocity (VCL), average path velocity (VAP), head displacement (ALH), sperm track straightness, (STR) and linearity (LIN) were determined in the storage of semen at 5°C for a period of 48 h. The extender supplemented with 1 mM melatonin led to higher percentages of progressive motility and LIN (46.4 and 58.9, respectively) and higher value of VSL and VAP (78.4 and 91.1 µm/s, respectively) in comparison to the control (P 0.05) effect in ALH and STR of the whole semen compared to the control sample. The results indicated that supplementation of melatonin to the semen extender can improve the semen motility.