The changes in semen quality,arginase activity and nitric oxide level in dexamethasone-treated rams

This study was made to investigate the effects of intramuscular administrations of dexamethasone on seminal plasma nitric oxide levels and arginase activity, and some spermatological parameters in rams. Ten Akkaraman rams weighing 50–60 kg and 2 years old were used as material in this study. The study was performed during the breeding season (September–November) for rams. The semen was collected by artificial vagina at 1st, 4th, 24th, 48th, 72nd and 96th hours for control group before dexamethasone administration. For treatment group, 0.25 mg/kg dexamethasone was administered and semen was collected at the time points described for control group. Spermatological characteristics of semen samples (semen volume, pH, sperm motility, density and abnormal sperm rate), seminal plasma arginase enzyme activities and nitric oxide levels were determined. It was determined that the administration of dexamethasone was detected to decrease seminal plasma arginase activity (p < .05 and .01) and nitric oxide level (p < .05), semen volume (p < .05 and .01), mass activity (p < .05 and .01), sperm density (p < .05) and sperm motility (p < .05 and .01), and to increase abnormal sperm rate (p < .05 and .01). In conclusion, dexamethasone is not recommended to be used during the breeding season as it damages the sperm quality of the rams.     

Cyclooxygenase 1 (COX1) as an indicator of sperm quality in humans

Sperm quality is important for in vitro fertilisation and embryo transfer and intracytoplasmic sperm injection in the treatment of human infertility. The purpose of this study was to screen for biomarkers that could evaluate sperm quality. We analysed semen samples in 172 fertile males; multivariate logistic regression analysis showed that the levels of COX1 (17.5 ng/ml) in seminal plasma may represent a useful biomarker for sperm quality (area under the curve: 0.745; sensitivity: 0.808; specificity: 0.722). Analysis indicated that the values of parameters related to sperm quality changed significantly (p < .05) between COX1 level ≥ 17.5 ng/ml group and COX1 level < 17.5 ng/ml group. Further analysis of two consecutive semen samples (1-hr interval) from 48 subjects revealed that the first semen samples (COX1 levels ≥ 17.5 ng/ml) had a higher sperm concentration and a larger proportion of spermatozoa showing progressive motility, a lower rate of sperm DNA fragmentation and a lower proportion of spermatozoa undergoing the acrosome reaction spontaneously (p < .05); identical results were observed for the second semen samples. These data indicated that COX1 could be used as an indicator for sperm status and may be helpful for identifying better quality spermatozoa for artificial insemination.     

Evaluation of the role of L-arginine on spermatological parameters,seminal plasma nitric oxide levels and arginase enzyme activities in rams

The purpose of this study is to investigate the effects of L-arginine on spermatological parameters, seminal plasma nitric oxide levels and arginase enzyme activities. Fertile rams that are 2–3 years old and weighing 50–60 kg were used as material. The semen was collected by artificial vagina at 1st, 4th, 24th, 48th, 72nd, 96th and 120th hours for the control group before L-arginine administration. For treatment groups, L-arginine was administered intraperitoneally at a dose of 5 mg kg⁻¹ bw⁻¹ and semen was collected at the time point described for the control group. Spermatological characteristics of semen samples (semen volume, pH, sperm motility, concentration and abnormal sperm rate), seminal plasma nitric oxide levels and arginase enzyme activities were determined. Increased seminal plasma nitric oxide level (p < .01), seminal plasma arginase activity (p < .01), semen volume (p < .05), semen mass activity (p < .05), sperm motility (p < .05) and concentration (p < .01) and decreased abnormal sperm rate (p < .05 and p < .01) were observed by L-arginine administration. In conclusion, it may be concluded that L-arginine application in rams during the breeding season may have positive effects on rams' reproductive performance.     

Chlorogenic acid improves the quality of boar semen subjected to cooled storage at 15°C

The aim of this study was to investigate the effect of chlorogenic acid (ChA) in boar semen stored at 15°C. Twelve ejaculates were processed into insemination doses at different concentrations of ChA (0.0, 1.5, 3.0, 4.0 and 6.0 mg/ml) or vitamin E (200 μl/ml) as positive control. Semen was analysed after 0, 24 and 72 hr of storage. ChA improved (p < .05) sperm motility, acrosome integrity and mitochondrial activity in all periods of storage. Furthermore, after 24 hr of storage, ChA above 1.5 mg/ml supported the sperm viability until 120 min after reheating (p < .05). Both ChA and vitamin E were similarly efficient in increasing the antioxidant capacity of semen, reducing the malondialdehyde levels before and after 72 hr of storage (p < .05). However, with 72 hr of storage, ChA at 3.0 mg/ml improved the mitochondrial activity over vitamin E (p < .05). In conclusion, results suggest that the concentration of 3.2 mg/ml of ChA is the best for semen stored for up to 24 hr. However, for semen stored for a longer period, 6.0 mg/ml or more should be used.     

Is addition or removal of seminal plasma able to compensate for the dilution effect of buffalo semen?

The present study aimed to compensate dilution effect using additional seminal plasma (SP) in conventional (80 million (M) spermatozoa/ml) dose and low spermatozoa/dose (8M spermatozoa/ml). We also attempted to confirm whether removal of SP before the extension of ejaculates affects post-thaw sperm quality of buffalo semen. For this, semen ejaculates (N = 15) were divided into four groups: control (CON), removal of SP by centrifugation (NSP), resuspension of the centrifuged semen pellet into SP (CEN) and extra supplementation of SP (ESP). All groups were diluted into two different semen doses to 20 and 2M spermatozoa/0.25 ml using tris egg yolk extender and subsequently cryopreserved. We found that neither addition nor removal of SP affected sperm motility, kinematics, longevity, mitochondrial superoxide production and high mitochondrial membrane potential (MMP). Further, the addition or removal of SP was not able to compensate dilution effect in 2M groups resulting in a significantly (p < .05) reduction in sperm motility, kinematics, sperm longevity, membrane integrity, MMP, and an increase production of mitochondrial superoxide. In conclusion, it appears that role of SP in the sperm cryopreservation process is insignificant.     

The presence of seminal plasma,especially derived from stallion semen,helps preserve chilled Asian elephant (Elephas maximus) sperm motility

The effects of seminal plasma (SP), derived from autologous, homologous and heterologous species (stallion, boar and dog) on chilled Asian elephant sperm quality, were determined. Semen was collected from eight males and samples with ≥30% motile spermatozoa were used in the study. Semen was diluted with Tris–glucose–egg yolk extender, supplemented with different SP types and preserved at 4°C for 48 hr. Experiment 1 (n = 31), showed that the presence of SP (autologous) helped to preserve sperm quality in terms of sperm motility and acrosome integrity (p < .05). Homologous SP did not result in better sperm quality than autologous SP. Heterologous SP from stallion provided higher sperm motility and velocities compared to autologous SP (p < .05). Experiment 2 (n = 14) determined the effect of different SP from four stallions. All stallion SP gave higher (p < .05) results for motile spermatozoa and sperm velocities than autologous SP. In conclusion, the presence of SP helps preserve Asian elephant sperm quality and stallion SP supports the motility of Asian elephant spermatozoa during cold storage.     

Prediction of reproductive function recovery after microsurgical varicocelectomy in men from infertile couples: Clinical and laboratory predictors

The objective was to reveal predictors for fertility recovery after varicocelectomy in subfertile men. This retrospective study recruited 93 men with clinical varicocele and pathozoospermia who underwent microsurgical varicocelectomy. Stepwise discriminant analysis was performed to identify predictors of spontaneous pregnancy (SP) after surgery. ‘Clinically significant improvement’ (CSI) following varicocelectomy was defined as an increase in total progressively motile sperm count (TPMSC) by at least 12.5 million (calculated from WHO-2010 reference values). 52% of patients showed CSI, and 28% reported SP after surgery. Patients who reported SP (group II), compared to that remained infertile (group I), were younger (27.3 ± 2.9 versus 30.2 ± 4.2 years; p < .01), had less infertility period (24.1 ± 14.0 versus 44.4 ± 32.9 months; p < .05) and had initially higher TPMSC (median (25% −75%) = 34 (11–67) versus 9.5 (0–33) mln; p < .05). The stepwise discriminant analysis showed that male age (coefficient value = −0.157), total sperm motility (0.024) and postoperative increase in TPMSC (0.010) were the significant predictors of SP. The predictive ability, sensitivity and specificity of the discriminant function were 84%, 87%, 76% respectively. This algorithm can be recommended after varicocelectomy in decision-making on natural conception or the ART protocols usage.     

A randomised trial comparing conventional semen parameters,sperm DNA fragmentation levels and satisfaction levels between semen collection at home and at the clinic

The aim of the randomised trial was to compare conventional semen parameters, sperm DNA fragmentation levels and satisfaction levels between semen samples collected at home and at the clinic. We recruited 110 men with a history of infertility for at least 1 year from the outpatient andrology clinic. Each man collected two semen samples, one at home and one at the clinic. Men were randomly assigned into the home first (n = 55) or clinic first (n = 55) groups. The primary outcome was sperm concentration. There was no significant difference in sperm concentration, sperm DNA fragmentation levels or other conventional semen parameters between home first and clinic first samples (p > .05), while satisfaction levels were significantly higher for home first samples (p < .01). Consistent results were obtained when comparing home-collected and clinic-collected samples within individuals. Men can be offered the option to collect semen samples at home for examination or assisted reproduction without compromising semen quality, especially for those with difficulty in producing semen samples at the clinic.     

Aqueous leaf extract of Moringa oleifera reduced intracellular ROS production,DNA fragmentation and acrosome reaction in Human spermatozoa in vitro

The effects of aqueous leaf extract of Moringa oleifera (MO) on human sperm functions and integrity was studied in vitro. Semen was obtained by masturbation after 3–5 days' abstinence from 34 healthy donors in Western Cape, South Africa. Liquefied semen was washed in human tubular fluid supplemented with 1% bovine serum albumin (HTF-BSA;1:5) with 10 min centrifugation at 300 g. Sperm suspensions were subsequently incubated with MO extract (0.625, 6.25, 62.5 and 625 µg/ml) for 1 hr, where HTF-BSA served as control. Sperm motility, vitality, DNA fragmentation, reactive oxygen species production, mitochondrial membrane potential, capacitation and acrosome reaction were assessed. Sperm motility, vitality, mitochondrial membrane potential and capacitation remained unchanged (p > .05). A dose-dependent decrease in sperm reactive oxygen species production (p < .0001), DNA fragmentation (p < .0001) and acrosome reaction (p < .001) was observed. An increase in the percentage of non-capacitated sperm (p < .01) was noted at 625 µg/ml. The antioxidant properties of MO actively maintained basic sperm functions, inhibited excess sperm free superoxide production and preserved acrosome reaction and DNA integrity. Further studies are needed to confirm the effect of aqueous MO leaf extract on fertility potential.     

Effects of temperature and storage time on the motility,viability, DNA integrity and apoptosis of processed human spermatozoa

The aim of this study was to evaluate motility, viability, DNA integrity and apoptosis of spermatozoa when washed semen samples were kept for up to 12 days at 4–6°C and 25°C. In this experimental study, 26 normozoospermic semen samples were washed twice in Modified Ham's F10 and resuspended in IVF fertilisation medium. Half of the specimens were stored at 4–6°C, and the other half was kept at 25°C for 12 days. The proportions of viable, motile, spermatozoa with double-stranded DNA and apoptotic spermatozoa were examined during storage time. Apoptosis was measured using annexin V-PI staining followed by flow cytometry. Results showed that sperm motility and viability decreased during 12 days of sample storage (p < .001). There was no significant difference between the two temperatures in terms of motility and viability for up to 2 days (p < .05). The percentage of spermatozoa with double-stranded DNA remained unchanged during the 12 days of storage at both temperatures (p > .05). Although there was no difference between the two temperatures in terms of motility, viability and apoptosis during the first two days of storage, storage of spermatozoa at 4–6°C is better than storage for a longer period than storage at 25°C. Sperm DNA resisted against denaturation during storage.     

Caffeine increased progressive motility of human spermatozoa in normozoospermic and asthenozoospermic semen samples and enhanced activity of seminal creatine kinase

Even though the effect of caffeine on humans' health has been revealed in various research studies, its effect on semen quality has yet to be well explained. Here, we measured the effect of caffeine at 1, 5, 10 and 20 mM on motility of human spermatozoa in normozoospermic and asthenozoospermic semen samples, level of seminal nitric oxide, chelation of seminal calcium ions and activity of seminal creatine kinase. Fifty-one normozoospermic and ten asthenozoospermic semen samples were recruited in this study. Sperm motility was evaluated by Makler counter, and seminal nitric oxide, seminal-free calcium and activity of seminal creatine kinase were measured spectrophotometrically. Caffeine at 10 mM significantly (p < .05) increased progressive motility of spermatozoa in both tested groups. Also, caffeine significantly increased (p < .05) activity of creatine kinase and insignificantly (p > .05) altered nitric oxide and free calcium levels in seminal plasma. In conclusion, progressive motility of human spermatozoa was found to be higher in the presence of caffeine at 10 mM in normozoospermic and asthenozoospermic semen samples; this increase, albeit partially, could be due to increased activity of seminal creatine kinase, but not to increased production of nitric oxide or chelation of free calcium ions.     

Novel additive for sperm cryopreservation media: Holotheria parva coelomic cavity extract protects human spermatozoa against oxidative stress—A pilot study

Cryopreservation is the most effective method for preserving semen for a long period of time. However, during the freeze–thaw process, production of reactive oxygen species (ROS) leads to a steep reduction in sperm fertility indices. In this study, we tested the effects of the extract of the coelomic cavity of five Holotheria parva, a marine organism rich in antioxidants, for its ROS-scavenging activity and cryoprotective effects on oxidative stress. Using a total of 50 semen samples, our results demonstrated that doses of 250 and 500 µg/ml of H. parva coelomic cavity extract significantly increased sperm vitality as compared to the control (p < .05). The addition of 250 µg/ml of the extract exerted a significant positive effect on sperm motility. Moreover, sperm DNA damage and ROS production were significantly reduced at extract concentrations of 250 and 500 µg/ml (p < .05). To the best of our knowledge, the results of this study represent the first demonstration of the possibility of improving sperm parameters and reducing ROS production and DNA damage by supplementing sperm freezing media with H. parva coelomic extract. Our results suggested that H. parva coelomic extract could be useful for improving the fertilising ability of frozen-thawed human semen.     

Relationship between visceral adiposity index and male infertility

The visceral adiposity index (VAI), which is calculated by using parameters such as body mass index (BMI), waist circumference, triglyceride, and high-density lipoprotein cholesterol, has been reported to be associated with visceral fat distribution and increased cardiometabolic risk. We aimed to investigate the relationship between the VAI and hormone profiles as well as semen analysis in male infertility. This retrospective study included 278 adult men that were admitted to the urology and endocrine outpatient clinics. Patient data were reviewed, and outcomes were recorded. BMI and VAI were calculated and noted separately. BMI was negatively correlated with serum testosterone and testosterone/ estradiol ratio (T/E2) (p < .05 for all). There was no significant relationship between BMI and semen parameters (p > .05 for all). However, there was a significant negative correlation between VAI and semen parameters, total serum testosterone and T/E2 (p < .05 for all). Furthermore, VAI was independently associated with the total motility score. The increase in visceral adiposity that can be demonstrated by VAI may adversely affect male fertility due to its metabolic and hormonal effects. Since infertility occurs as a result of different metabolic conditions, the underlying mechanisms can be better understood from a multidisciplinary perspective.     

Long-term stiripentol administration,an anticonvulsant drug,does not impair sperm parameters in rats

In this study, we investigated the influence of long-term administration of stiripentol on sex hormones and semen quality in young Wistar rats. Investigated animals received for 6 months either stiripentol or saline solution. After one month, stiripentol increased temporarily serum level of testosterone (p < 0.05) and FSH (p < 0.01). However, after 6 months levels of testosterone, FSH, LH, prolactin and SHBG were comparable in both groups. After 6 months, semen analysis did not reveal differences in sperm concentration, total sperm count and sperm motility between groups. However, stiripentol increased the rate of head defect (p < 0.001) and midpiece abnormalities (p < 0.05). Flow cytometry revealed higher percentage of live cells without lipid peroxidation (p < 0.00001) and higher percentage of live spermatozoa with intact acrosomes (p < 0.000001) in rats receiving stiripentol. There was no significant difference between groups in sperm mitochondrial activity and DNA fragmentation index. However, percentage of high DNA stainability cells was increased in stiripentol group (p < 0.001). The data showed that stiripentol does not cause obvious disturbances in young rat's semen. Detected changes in semen morphology and chromatin structure need further explanation, and their influence on rat's fertility should be evaluated.     

Apoptotic sperm biomarkers and the correlation between conventional sperm parameters and clinical characteristics

The principal aim of this retrospective study was to examine the relationship between sperm apoptotic biomarkers and the patient's biclinical characteristics, the conventional sperm parameters and the results of assisted reproductive technology. Sperm analysis, activated caspases, annexin V staining for phosphatidylserine (PS) externalisation and labelling assay for DNA fragmentation were assessed in 122 males of infertile couples. Fifty‐seven couples were allocated to the natural conception group, and 65 couples underwent IVF or ICSI. Semen of IVF/ICSI patients showed a higher proportion of apoptotic spermatozoa in their spermatozoa when compared with a natural conception group (p < .05). Sperm apoptotic biomarkers correlated with age, FSH, and conventional sperm parameters. DNA fragmentation correlated positively with the percentage of semen having externalised PS (r = .78, p = 0) and activated caspases (r = .71, p = 0). Patients without clinical pregnancy had higher frequency of DNA fragmentation, externalised PS and activated caspases compared to patients with clinical pregnancy (p < .001). The best specificity and greater sensitivity were obtained with the test of the DNA fragmentation compared to the other biomarkers. Among the apoptotic biomarkers, only DNA fragmentation was found to predict natural or assisted pregnancy better than conventional sperm parameters.     

Response to cooling of pony stallion semen selected by glass wool filtration

The aim of this study was to compare the sperm separation technique using filtration through glass wool compared with just diluted cooled semen. Eighteen ejaculates were collected from 6 pony stallions of the Brazilian pony breed. Evaluations were done on pH, osmolarity, total motility, membrane functionality (HOST), membrane integrity (CFDA/PI), morphology and mitochondrial viability (MTT) in fresh, 24 and 48 h of cooled semen at 5°C. After dilution, the half of the extended semen was cooled (control group). The other half was cooled after filtration trough glass wool (filtered group). Retained semen was considered the portion of cells that did not transpose glass wool barrier. Total motility from the control, filtered and retained groups after 24 h of cooling was 35.5%, 43.3% and 10% (p < .0001) respectively. Sperm membrane integrity percentage at the CFDA/PI test was 37.9%, 44.8% and 14.8% (p < .0001), on the control, filtered and retained groups respectively. The results confirmed that the passage of spermatozoa through glass wool increased the selection of spermatozoa from pony stallions with higher motility, mitochondrial viability and membrane integrity for cooling in milk extender up to 24 h. Moreover, it was not obtained higher sperm parameters to control after cooling 48 h under the conditions that the study was conducted.     

Nano-purification of raw semen minimises oxidative stress with improvement in post-thaw quality of buffalo spermatozoa

The study consisted of application of anti-ubiquitin antibodies (Abs)-coated iron oxide-nanoparticles (IONPs) for minimisation of oxidative stress to contemporary live spermatozoa from the raw semen. Round-shaped IONPs (12.09 ± 0.91 nm) after two-stage functionalisation (silanisation and pegylation) were conjugated with Abs. Four aliquots from each of the 24 ejaculates (4 buffalo bulls) formed Control (Group I) and treatment (II, III and IV) groups; each containing 150 ± 25 million dead/damaged spermatozoa. IONPs-Abs complex were added at ratio of 1:1 (0.5 µg/ml), 1:2 (1.0 µg/ml) and 1:4 (2.0 µg/ml), respectively, in Groups II, III and IV. The semen quality parameters showed improvement at lag-stage (post-nano-purification before processing for cryopreservation). The mean post-thaw motility (%) in Group IV was found to be greater (p < .05) than Group I. Moreover, the overall DNA integrity (%) at post-thaw stage was improved in the nano-purified semen samples. The value of malondialdehyde was greater (p < .001) in Group I than Groups II, III and IV. The mean total antioxidant capacity and superoxide dismutase (U/mg protein) activity values in Group IV was greater (p < .05) than Group I. The study results show that IONPs conjugated with anti-ubiquitin Abs at 2.0 µg/ml can be an effective dose for depletion of dead/damaged spermatozoa from buffalo ejaculates to minimise oxidative stress.     

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.     

The presence of SARS-CoV-2 virus in semen samples of patients with COVID-19 pneumonia

The SARS-CoV-2 set off a pandemic involving millions of people around the world. The topic of discussion is the possible viral detection in different body fluids than respiratory droplets. Therefore, we evaluated the possible presence of SARS-CoV-2 in semen and urine samples. Thirty patients were included in the study aged 35.67 ± 6.84 years. The day after the pharyngeal and/or nose swab of SARS-CoV-2 was positive, urine and semen samples were taken from patients, and the presence of SARS-CoV-2 was investigated. Laboratory tests and chest CT findings were evaluated simultaneously. SARS-CoV-2 was detected in four (13.3%) patients’ semen samples and in seven (23.3%) patients’ urine samples. White blood cell (WBC), neutrophil, C-reactive protein (CRP), ferritin, alanine transaminase (ALT), lactate dehydrogenase (LDH) and procalcitonin were significantly higher in patients with SARS-CoV-2 in semen (p < .05), though no statistical difference was found in urine (p > .05). Patients with severe pneumonia findings in Chest CT images are likely to be PCR positive in semen and urine samples (p = .005, p = .001). SARS-CoV-2 was not detected in urine and semen samples of patients after they had recovered (average duration 23 ± 4 days). SARS-CoV-2 can be detected in the urogenıtal fluıds of patients with severe clinical conditions and high viral load.     

Aspirin decreases human sperm motility and vitality,chelates seminal calcium,but insignificantly reduces seminal nitric oxide production

Lately, there is a systematic research consensus that reveals adverse effects of aspirin on semen quality characteristics; however, such consensus is lacking further confirmation by human studies. Therefore, here, we asked whether sperm motility and vitality are affected in the presence of aspirin at 0.1 and 1 mM in the ejaculated semen, and whether such effect may be due to an alteration in seminal calcium ions or seminal nitric oxide production. Forty-three semen samples from different normozoospermic men were recruited in this study. Sperm motility was measured by Makler counting chamber, and sperm vitality was measured by Eosin test. Calcium chelating effect of aspirin and seminal nitric oxide production was measured spectrophotometrically. Aspirin at both tested concentrations significantly (p < .05) reduced progressive grade-a motility and vitality of spermatozoa. Additionally, aspirin was found to have significant ability (p < .05) to bind seminal calcium ions, but insignificantly reduced the amount of seminal nitric oxide. In conclusion, sperm motility and vitality were reduced in the presence of aspirin at 0.1 and 1 mM in semen. Such reduction may be attributable to the ability of aspirin to chelate seminal calcium ions, but not to an alteration in the amount of nitric oxide produced.