In vitro reprotoxicity of carboxyl‐functionalised single‐ and multi‐walled carbon nanotubes on human spermatozoa

Reproductive toxicity of carboxyl‐functionalised carbon nanotubes (CNT‐COOH), as the most commonly used form of water‐soluble CNTs, is not clearly studied. The aim of this study was to investigate in vitro toxicity of carboxylated single‐walled and multi‐walled CNTs (SWCNT‐COOH and MWCNT‐COOH) against human spermatozoa. Sperm cells from healthy donors were incubated with 0.1–100 μg/ml of SWCNT‐COOH or MWCNT‐COOH at 37°C for up to 5 hr. Viability of sperm cells was assessed using MTT test, and sperm motility was evaluated following World Health Organization guideline. Production of reactive oxygen species (ROS) and nitric oxide (NO) in sperm was also assessed. We showed that both MWCNT‐COOH and SWCNT‐COOH following incubation in vitro with human spermatozoa did not exert negative effect on viability while motility was significantly (p < .05) dropped in a dose‐dependent manner. Moreover, there was no significant effect of the type, dose and exposure time of the CNT‐COOH on NO production. Exposure of sperm cells to both examined types of CNTs at concentrations as low as 0.1 μg/ml caused a significant increase in ROS levels. In conclusion, carboxylated forms of CNTs seem to be harmful for human spermatozoa. Further studies, especially using in vivo models, are needed to decide about reprotoxicity of carboxylated forms of CNTs.     

Protective effect of the superoxide dismutase mimetic MnTBAP during sperm vitrification process

Sperm cryopreservation is widely used in assisted reproduction and male infertility therapy; however, it induces oxidative stress affecting sperm quality. This work evaluated the effect of the antioxidant MnTBAP during vitrification steps in human spermatozoa. First, the effect of MnTBAP on viability and ROS production was evaluated. Then, the spermatozoa were vitrified in straws with the vitrification, warming and post-warming incubation media separately supplemented with MnTBAP. An untreated control was included. The sperm viability, ROS production, total and progressive motility were evaluated. The results showed that the direct exposure of spermatozoa to MnTBAP significantly decreases the ROS levels in comparison with the untreated control without affecting the viability. The supplementation of the vitrification medium with MnTBAP did not affect the parameters analysed. However, the supplementation of the warming and incubation post-warming media resulted in a decrease in ROS production and maintained viability and motility for 4 hr after warming with concentrations up to 100 μM of MnTBAP. Higher concentrations of MnTBAP caused a decrease in total motility. In conclusion, the use of MnTBAP during the warming or post-warming incubation media has beneficial effect decreasing ROS levels and maintaining the viability and motility during the vitrification procedure.     

The effect of vitamin E on sperm motility and viability in asthenoteratozoospermic men: In vitro study

Induction of oxidative stress during the sperm preparation process for assisted reproductive techniques (ART) in men can weaken sperm parameters. Vitamin E (VE) is considered a factor in boosting male fertility. This experimental study (in vitro) aimed to assess the impact of VE supplementation on sperm quality and lipid peroxidation during sperm sampling at different times. For this mention, semen samples were collected from 50 asthenoteratozoospermic men. Samples were divided into control and test groups for 2, 4 and 6 hr that the test group was incubated with VE (2 mM). In two groups, total motility, progressive motility and viability based on the WHO 2010 criteria were assessed. Moreover, malondialdehyde (MDA) levels were evaluated in each group. In the control group, total and progressive motility and sperm viability were decreased significantly after 2 hr; however, MDA levels were increased significantly after 6 hr. Also, in the test group, sperm parameters were increased significantly after 2 hr, and MDA levels were decreased significantly after 6 hr compared to the control group. In outcome, in vitro VE supplementation may protect spermatozoa from the adverse effect of oxidative stress during sperm preparation via preservation antioxidant processes in normal condition.     

LED‐based red light photostimulation improves short‐term response of cooled boar semen exposed to thermal stress at 37°C

Pre‐treatment of boar semen with a red light photostimulation procedure increases its “in vivo” fertilising ability. However, “in vitro” conducted studies shown contradictory results regarding the ability of photostimulated spermatozoa to react against strong stress and to achieve the capacitation status. The aim here was to determine the effect of photostimulation on the response to short‐term moderate thermal stress of boar semen. Boar semen was exposed to red LED light regime emitting a 620–630 nm during 10 min of light, 10 min of rest and 10 min of light after 3 hr since semen was collected. An aliquot without photostimulation was included as a control. After the photostimulation, the sperm cells were incubated for 15 min at 37°C. Afterwards, motility, viability, intracellular Ca²⁺ level and production of reactive oxygen species (ROS) and peroxynitrite were analysed. The results showed that the photostimulated group maintained total motility throughout the time, whereas a significant decrease in total motility was observed in the nonphotostimulated control group. Furthermore, for kinetic parameters of motility, a significant increase was observed in LIN, STR and WOB in photostimulated spermatozoa. Peroxynitrite production was significantly increased in the photostimulated spermatozoa, whereas viability, ROS production and intracellular Ca²⁺ levels were not affected by photostimulation. In conclusion, photostimulation of commercial boar semen has a positive effect on motility of spermatozoa subjected to a short‐term moderate thermal stress, which was concomitant with an increase in peroxynitrite production.     

Impact of biological and artificial seminal fluids on sperm parameters and DNA status in asthenozoospermic ejaculates

The chemical composition and physiological properties of seminal fluid (SF) affect sperm quality. The objective was to investigate the effects of in vitro exposure of artificial seminal fluid (ASF) and biological seminal fluid (SF) on sperm quality. Asthenozoospermic ejaculates (n = 20) were divided into two aliquots. The first aliquot was centrifuged for obtaining asthenozoospermic SF. The second aliquot was processed with density gradient centrifugation (DGC), and the pellet was diluted separately with following media: (a) ASF; (b) Ham's F10; (c) normozoospermic SF; and (d) asthenozoospermic SF. Sperm parameters and DNA status were assessed after DGC, as well as 2 and 24 hr after incubation. The data showed that sperm progressive motility, viability and DNA integrity were significantly higher in ASF than Ham's F10 medium immediately after DGC. At 2 and 24 hr, the progressive motility was significantly decreased in biological SF compared with ASF and Ham's F10. DNA fragmentation index (DFI) was significantly lower in normozoospermic SF than asthenozoospermic SF and Ham's F10 at time 2 hr. In conclusion, normal SF showed the protective role on sperm DNA structure. Moreover, ASF preserved sperm motility better than biological SF during 24 hr, despite being similar to normal SF regarding DNA integrity preservation in short time.     

Antioxidant effect of hydroxytyrosol on human sperm quality during in vitro incubation

The aim of the study was to evaluate the antioxidant potential of hydroxytyrosol (HT) on human sperm quality during incubation in vitro. Semen samples collected from men attending the Laboratory of Histology‐Embryology of Sfax Faculty of Medicine (Tunisia) for infertility investigations were evaluated for initial sperm parameters. Only normal selected ejaculates (n = 15) were centrifuged and incubated further with or without HT (200ug ml^?1) at room temperature for 45 min. After incubation, sperm motility and viability, DNA oxidation and reactive oxygen species (ROS) production were assessed. The results showed that centrifugation significantly influenced sperm motility and viability. The supplementation of HT in incubating media improved (P = 0.01) significantly sperm viability and decreased sperm DNA oxidation (P < 0.001) and ROS levels (P = 0.03) following centrifugation. It can be concluded that supplementation of HT might be helpful to maintain the human spermatozoon after centrifugation.     

Deoxynivalenol reduces quality parameters and increases DNA damage in mice spermatozoa

This study was performed to investigate in vitro effects of deoxynivalenol (DON) on mice sperm quality parameters including viability, motility and DNA damages at various concentrations and exposure times. Mice spermatozoa were exposed to DON at 0, 2.5, 5 and 10 µM for 1, 3 and 6 hr, motility parameters were evaluated by computer‐assisted analysis and viability was examined by colorimetric metabolic activity assay and HOS test. DNA damage was examined by acridine orange staining, and sperm damages via lipid peroxidation pathway were determined by malondialdehyde (MDA) content measurement. DON affected sperm parameters in a concentration‐ and time‐dependent manner. In all test groups, the average path velocity and progressive motile spermatozoa were remarkably reduced. In comparison with the controls, after 1, 3 and 6 hr exposure to DON, viability of spermatozoa was reduced 25, 30 and 49% respectively. DON exposure at 10 µM for 6 hr resulted in 15% DNA damage and 2.5‐fold more MDA generation, when compared with nonexposed spermatozoa. Our data suggest that DON causes sperm quality parameters decline in concentration‐ and time‐dependent fashion, which attribute to the reduction in sperm metabolic activity and membrane integrity and equally to increase in lipid peroxidation rate and DNA damage.     

Effect of tertiary‐butyl hydroperoxide (TBHP)‐induced oxidative stress on mice sperm quality and testis histopathology

Male infertility is responsible for approximately 50% of infertility worldwide. Reactive oxygen species are one of the major causes of male infertility. In this study, the effects of oxidative stress induced by tertiary‐butyl hydroperoxide (TBHP) on sperm quality and testis tissue are investigated. After determination of LD₅₀, TBHP with a concentration of 1 : 10 LD₅₀ was injected in adult male mice strains Balb/c for two consecutive weeks. Their testis tissues were used for cell viability, histopathology analysis and ROS assay. The epididymis was also surveyed for sperm analysis by CASA system. The sperm motility, count and viability decreased in the TBHP‐treated mice compared to the control mice. The flow cytometry analysis showed a significant increase in H₂O₂ and levels in both testis and sperm within 2 weeks after intraperitoneal injection. Body weights revealed no treatment‐related effects, but atrophy of testis and a decrease of testis cells viability were observed. The results showed that exposure to TBHP could lead to morphological changes in seminiferous tubules. TBHP‐induced oxidative stress caused a decrease in sperm parameters and testis cells viability. That is due to an increase level of ROS in the testis and their deleterious effects on genomic levels.     

Famotidine does not affect human sperm fertility characteristics (motility,viability and DNA integrity) in vitro

Famotidine, a histamine‐2 receptor antagonist, is commonly used to relieve the acid‐related gastrointestinal diseases; however, its effect on human sperm parameters, and hence on sperm function, is still undetermined. Here, we intended to measure human sperm motility, viability, and DNA integrity of ejaculated human sperm in the presence of famotidine at 0, 0.1, 1 and 10 mM concentrations in vitro. Forty‐nine semen samples of normal count, motility, and morphology were included in this study. Sperm motility was assessed using Makler counting chamber and a phase contrast optics (200× magnification), whereas sperm viability was assessed using eosin–nigrosin staining procedure. The effect of famotidine on sperm DNA integrity was measured using flow cytometry. Famotidine at 0.1, 1 or 10 mM had insignificant effect on human sperm motility (progressive, p = .9594; and total, p = .8420), sperm viability (p = .6471), and content of DNA breaks in sperm (p > .05) compared with the control. In conclusion, famotidine at 0.1, 1 or 10 mM did not alter human sperm motility, viability or DNA integrity in vitro. Although, these findings indicate safety of famotidine in human sperm, further in vivo studies are required to establish the drug's safety.     

N‐acetyl‐L‐cysteine pre‐treatment protects cryopreserved bovine spermatozoa from reactive oxygen species without compromising the in vitro developmental potential of intracytoplasmic sperm injection embryos

Excess of reactive oxygen species (ROS) on in vitro embryo production systems negatively affects the quality and developmental potential of embryos, as result of a decreased sperm quality and increased DNA fragmentation. This issue is of major importance in assisted fertilisation procedures such as intracytoplasmic sperm injection (ICSI), because this technique does not allow the natural selection of competent spermatozoa, and therefore, DNA‐damaged spermatozoa might be used to fertilise an egg. The aim of this study was to investigate a new strategy to prevent the potential deleterious effect of ROS on cryopreserved bovine spermatozoa. We evaluated the effect of a sperm pre‐treatment with different concentrations of N‐acetyl‐L‐cysteine (NAC) on ROS production, viability and DNA fragmentation and assessed the effect of this treatment on the in vitro developmental potential and quality of embryos generated by ICSI. The results show a strong scavenging effect of 1 and 10 mm NAC after exposure of spermatozoa to a ROS inducer, without compromising the viability and DNA integrity. Importantly, in vitro developmental potential and quality of embryos generated by ICSI with spermatozoa treated with NAC were not affected, confirming the feasibility of using this treatment before an ICSI cycle.     

Comparison of two cooling protocols for llama semen: with and without collagenase and seminal plasma in the medium

Seminal plasma (SP) of South American Camelids could interfere with the interaction of spermatozoa with the extenders; therefore it becomes necessary to improve semen management using enzymatic treatment. Our objective was to compare two cooling protocols for llama semen. Twelve ejaculates were incubated in 0.1% collagenase and then were divided into two aliquots. One was extended in lactose and egg yolk (LEY) (Protocol A: collagenase and SP present). The other aliquot was centrifuged, and the pellet was resuspended in LEY (Protocol B: collagenase and SP absent). Both samples were maintained at 5°C during 24 hr. Routine and DNA evaluations were carried out in raw and cooled semen. Both cooling protocols maintained sperm viability, membrane function and DNA fragmentation, with Protocol A showing a significantly lowered total and progressive motility (p < .05) and Protocol B showing a significant increase in chromatin decondensation (p < .05). Protocol A avoids centrifugation, reducing processing times and making application in the field simpler. However, as neither protocol showed a significant superiority over the other, studies should be carried out in vivo to evaluate the effect on pregnancy rates of the presence of collagenase and SP in semen samples prior to either cooling or freeze‐thawing.     

DNA integrity and methylation changes of mouse spermatozoa following prolonged incubation

Sperm quality can be affected by different factors including the length of incubation time between sperm preparation and intracytoplasmic sperm injection. Here, we have evaluated the level of DNA methylation and expressions of related genes in mice spermatozoa. The spermatozoa were divided into three groups: fresh, spermatozoa incubated at room temperature (RT) and 37°C for 24 hr. The sperm chromatin structure assay was used to determine the DNA fragmentation index (DFI), and DNA methylation was analysed by flow cytometry. The expression levels of DNA methylation‐related genes were determined by quantitative real‐time PCR (qRT‐PCR). According to the results, we observed significantly higher sperm progressive motility and viability in the group incubated at RT compared to the spermatozoa incubated at 37°C (p < 0.05). Spermatozoa incubated at 37°C had a higher DFI compared to the other groups (p < 0.05), but the DNA methylation level significantly decreased (p < 0.05). qRT‐PCR analysis showed increased Dnmt‐1 expression in spermatozoa after 24‐hr incubation at 37°C. However, there were significantly higher expression levels of Dnmt‐3l, Dnmt‐3a and Dnmt‐3b after incubation at both RT and 37°C compared to the fresh group (p < 0.05). The 24‐hr incubation period affected both sperm DNA methylation and integrity. This study indicated that incubation at RT resulted in better sperm quality.     

Survival of buffalo bull spermatozoa: effect on structure and function due to alpha‐lipoic acid and cholesterol‐loaded cyclodextrin

Sperm survival depending upon integral membranes and function is imperative for fertilization. This study was designed to augment survival of buffalo spermatozoa using alpha‐lipoic acid (ALA) and cholesterol‐loaded cyclodextrin (CLC) during cryopreservation. Semen was frozen using 0, 0.5, 1, 1.5, 2 and 2.5 mmol L^?1 ALA (experiment 1) and ALA or CLC separately or together (experiment 2). Semen was assessed for post‐thaw motility, plasma membrane integrity (PMI), intact acrosome and plasma membrane (IACR‐IPM) and DNA integrity at 0, 1.5, 3 and 4.5 hr of incubation. In experiment 1, use of 0.5 mmol L^?1 ALA enhanced the sperm cryosurvival and post‐thaw longevity than other groups up to 4.5 hr of incubation, and this concentration of ALA was used in second experiment with CLC. The results revealed higher (p < .05) sperm survival function and time of sperm attributes due to use of ALA than CLC and control. However, the sperm quality did not improve (p > .05) when ALA was combined with CLC. In conclusion, survival of buffalo bull spermatozoa during freeze‐thawing and post‐thaw incubation can be enhanced more with ALA than CLC or control, followed by CLC than control. However, there is no synergistic effect on survival of buffalo bull spermatozoa due to ALA and CLC.     

Effect of mitochondrial calcium uniporter blocking on human spermatozoa

Calcium (Ca²⁺) regulates a number of essential processes in spermatozoa. Ca²⁺ is taken up by mitochondria via the mitochondrial calcium uniporter (mCU). Oxygen‐bridged dinuclear ruthenium amine complex (Ru360) has been used to study mCU because it is a potent and specific inhibitor of this channel. In bovine spermatozoa, it has been demonstrated that mitochondrial calcium uptake inhibition adversely affects the capacitation process. It has been demonstrated in human spermatozoa that mCU blocking, through Ru360, prevents apoptosis; however, the contribution of the mCU to normal human sperm function has not been studied. Therefore, the aim of this study was to evaluate the effect of mCU blocking on human sperm function. Spermatozoa obtained from apparently healthy donors were incubated with 5 and 10 μm Ru360 for 4 h at 37 °C. Viability was assessed using propidium iodide staining; motility was determined by computer‐aided sperm analysis, adenosine triphosphate (ATP) levels using a luminescence‐based method, mitochondrial membrane potential (ΔΨm) using JC‐1 staining and reactive oxygen species (ROS) production using dihydroethidium dye. Our results show that mCU blocking significantly reduced total sperm motility and ATP levels without affecting sperm viability, ΔΨm and ROS production. In conclusion, mCU contributes to the maintenance of sperm motility and ATP levels in human spermatozoa.     

Cadmium inhibits motility,activities of plasma membrane Ca2+‐ATPase and axonemal dynein‐ATPase of human spermatozoa

Cd²⁺ has been associated with decreased sperm motility in individuals exposed to this element, such as smokers. Among other factors, this lowered motility could be the result of inhibition exerted by Cd²⁺ on the activity of the sperm ATPases associated with sperm motility. In this study, we evaluated the plasma membrane Ca^{2 +} ‐ATPase and the axonemal dynein‐ATPase activities as well as sperm motility, in the presence of different free Cd²⁺ concentrations in the assay media. It was found that spermatozoa incubated for 5 h in a medium containing 25 nm free Cd²⁺ showed a significant inhibition of progressive motility, reaching values even lower at higher Cd²⁺ concentrations. In addition, it was found that the activity of the plasma membrane Ca²⁺‐ATPase reached maximal inhibition at 50 nm free Cd²⁺, with a K_{50% inhibition} of 18.3 nm free Cd²⁺. The dynein‐ATPase activity was maximally inhibited by 25 nm free Cd²⁺ in the assay medium, with a K_{50% inhibition} of 11.3 nm Cd²⁺. Our results indicate that the decreased activity of the sperm ATPases might have a critical importance in the biochemical mechanisms underlying the decreased sperm motility of individuals exposed to Cd²⁺.     

The effect of reactive oxygen species on equine sperm motility, viability, acrosomal integrity, mitochondrial membrane potential, and membrane lipid peroxidation

The objective of this study was to examine the influence of reactive oxygen species (ROS), generated through the use of the xanthine (X)-xanthine oxidase (XO) system, on equine sperm motility, viability, acrosomal integrity, mitochondrial membrane potential, and membrane lipid peroxidation. Equine spermatozoa were separated from seminal plasma on a discontinuous Percoll gradient, and spermatozoa were incubated with 0.6 mM X and 0.05 U/mL XO for 30 minutes. Catalase (150 U/mL), superoxide dismutase (SOD, 150 U/mL), or glutathione (GSH, 1.5 mM) were evaluated for their ability to preserve sperm function in the presence of the induced oxidative stress. At the end of the 30-minute incubation, sperm motility was determined by computer-assisted semen analysis. Viability and acrosomal integrity were determined by Hoechst-Pisum sativum staining, and mitochondrial membrane potential was determined by staining with JC-1. Incubation with the X-XO system led to a significant (P < .01) increase in hydrogen peroxide production and an associated decrease (P < .01) in motility parameters. Total motility was significantly (P < .01) lower in the presence of X-XO compared with the case of the control (29%+/-9% vs 73%+/-1%, respectively). Catalase, but not SOD, prevented a decline in motility secondary to oxidative stress (71%+/-4% vs 30%+/-3%, respectively). The addition of glutathione had an intermediate effect in preserving sperm motility at the end of the 30-minute incubation (53%+/-3%). No influence of X-XO could be determined on viability, acrosomal integrity, or mitochondrial membrane potential. In order to promote lipid peroxidation, samples were incubated with ferrous sulfate (0.64 mM) and sodium ascorbate (20 mM) for 2 hours after the X-XO incubation. No increase in membrane lipid peroxidation was detected. This study indicates that hydrogen peroxide is the major ROS responsible for damage to equine spermatozoa. The decrease in sperm motility associated with ROS occurs in the absence of any detectable decrease in viability, acrosomal integrity, or mitochondrial membrane potential or of any detectable increase in lipid peroxidation.     

Trehalose sustains a higher post‐thaw sperm motility than sucrose in vitrified human sperm

One of the cryopreservation methods that best preserves sperm function is vitrification. However, comparative studies have not been performed to evaluate the effect of nonpermeable cryoprotectors on sperm function for prolonged periods of time post‐devitrification. These times are necessary, especially in in vitro fertilisation and intrauterine insemination, for gamete interaction and then fertilisation to occur, while maintaining motility to arrive at the fertilisation site. In this study, sucrose (.25 m ) and trehalose (.1 and .05 m ) were compared in essential parameters like motility and plasma membrane integrity for 12 hr. Post‐devitrification sperm motility using .1 m trehalose was 68.9%, higher than that obtained with .05 m trehalose (59.9%, p < .0081) and .25 m sucrose (57.9%, p < .0002). Similar results were obtained at 6 and 12 hr with .1 m trehalose (58.0% and 42.3% respectively) compared to .05 m trehalose (p < .0184 and p < .033) and .25 m sucrose (p < .0001 and p < .0012).There was no difference between .25 m sucrose and .05 m trehalose. Membrane integrity was best preserved at time 0 by .1 m trehalose (p < .05), but there was no significance at 6 and 12 hr compared to sucrose. Our results suggest that for assisted reproduction techniques that require motile spermatozoa for a longer period of time, use of .1 m trehalose is recommended in the sperm vitrification technique.     

Reactive oxygen species generation by seminal cells during cryopreservation

Objectives. To test the hypothesis that conventionally used procedures for semen cryopreservation may cause an increase in the production of reactive oxygen species (ROS) by sperm or by seminal leukocytes, which may contribute to poor sperm function following cryopreservation.Methods. Eighteen semen specimens with normal parameters from healthy male donors 22 to 40 years of age were each divided into two portions. The first portion was combined 1:1 with Test Yolk Buffer-Glycerol Freezing Medium and was frozen by gradual cooling into liquid nitrogen (−196°C). The second portion was washed and the cells were resuspended in Sperm Washing Medium (SWM) and incubated at room temperature to serve as controls. After a period of treatment, frozen samples were thawed and semen cells were washed and resuspended in SWM. ROS generation by semen cells from each treatment group was measured on a luminometer. Sperm motility, sperm viability, and sperm membrane integrity were also measured in both control and freeze-thaw samples. To further assess ROS generation by semen cells during the cooling process, aliquots of washed semen cells and purified polymorphonuclear leukocytes (PMNs) were incubated separately at different temperature conditions (37°C, 22°C, 4°C, and −20°C). ROS activity in each treatment group was measured and compared with each other.Results. In both semen cells and PMNs, ROS activity increased significantly during the cooling process. The highest ROS levels were recorded in both groups when cooled to 4°C. The ROS levels were extremely low in samples cooled to −20°C and in freeze-thaw samples, probably due to marked loss of cell viability.Conclusions. Gradual reduction of temperature during the process of semen cryopreservation can cause a significant ROS generation by semen cells. ROS is particularly elevated during cooling if the semen sample is contaminated by more than 0.5 × 10⁶ leukocytes. Removal of leukocytes from semen samples or treatment with antioxidants prior to cryopreservation may improve sperm viability and function.     

Omeprazole does not alter human sperm motility,viability or DNA integrity in vitro

The effect of omeprazole, a commonly used drug belongs to proton–‐pump inhibitor class, on human sperm function is still undetermined. Here, we hypothesised that addition of omeprazole to the ejaculated human semen may affect sperm parameters, and hence sperm function. Therefore, we assessed the in vitro effect of omeprazole on human sperm motility, viability and DNA integrity. Sixty‐six normozoospermic semen samples were collected randomly from men who attended the andrology laboratory at King Abdullah University Hospital. Sperm motility, viability and DNA breaks were assessed in the presence (1‐hr incubation at 37°C) of omeprazole at 5, 10, 20 and 50 µM compared to control (0 µM). None of the examined sperm parameters, at any tested omeprazole concentration, showed significant difference (p > 0.05) compared with the control. In conclusion, omeprazole at 5, 10, 20 and 50 µM does not alter human sperm motility, viability or DNA integrity in vitro.     

The effect of cysteine and glutamine on human sperm functional parameters during vitrification

Assuming the adverse effects of reactive oxygen species (ROS) on sperm function, this study was conducted to assess the effects of cysteine and glutamine as effective antioxidants on human sperm parameters under vitrification. Twenty normozoospermic samples were used. The samples were subjected to a vitrification process and cysteine (5 and 10 mM) and glutamine (10 and 15 mM). The sperm motility parameters, mitochondrial membrane potential (MMP), plasma membrane integrity (PMI), DNA damage and intracellular ROS damage were assessed for each sample. Statistical analyses showed that motility, mitochondrial membrane potential and DNA damage decreased in the vitrified groups with cysteine 5, 10 mM and glutamine 10, 15 mM separately. Also intracellular ROS increased significantly compared to the fresh group (p < .05). No significant differences were observed for PMI compared with the fresh group (p > .05). Supplementation of cysteine and glutamine in both concentrations separately decreased intracellular ROS and DNA damage of spermatozoa with significant increase in PMI, MMP and progressive motility compared to vitrified control group (p < .05). The results showed no significant effect of a specific concentration in cysteine and glutamine on sperm parameters compared to other concentrations. Both amino acids have the potential to improve the harmful effects of freezing on sperm parameters.