Percentage Of Total Motility Research Articles

Semen quality and sperm DNA integrity in city policemen exposed to polluted air in an urban industrial agglomeration.

We examined sperm quality in a cohort of city policemen in Ostrava at the end of a period with high concentrations of air pollutants (winter) and in the same cohort at the end of a relatively low exposure period (summer). The study group was comprised of 54 nonsmoking city policemen living and working in Ostrava, Czech Republic. Average daily air-pollutant concentrations recorded by stationary monitoring for 90 days preceding the collection of semen samples were evaluated for different city districts of Ostrava. Standard semen parameters were assessed according to the guidelines of the World Health Organization (2010). The parameters were semen volume, sperm concentration, sperm morphology, sperm motility, acrosome reaction and sperm plasma membrane integrity. Sperm DNA damage was analysed by the sperm chromatin structure assay (SCSA). Sperm motion characteristics were determined by Computer Assisted Semen Analysis (CASA). The concentrations of all monitored pollutants (particulate matter, sulphur dioxide, nitrogen oxides, carbon monoxide, benzo[a]pyrene, benzene) were significantly increased during winter (p<0.001), except for ozone, the concentration of which was significantly higher during summer. Sperm volume, concentration, % vitality, % sperm morphology (normal form) and % acrosome-intact sperm did not differ significantly between the monitoring periods. The percentages of total motility and progressive motility were significantly higher in March, i.e. at the end of winter (p=0.001). However, CASA testing showed differences in sperm motion kinetics between spring and autumn samples. In the spring samples, we found a significantly lower % of straightness (p=0.044) and the length of straight-line path (p=0.01), while linearity and straight-line velocity were near the borderline value (p=0.064; p=0.054, respectively). As compared to summer, high exposure to air pollution during winter significantly increased the extent of sperm chromatin integrity damage (median 22.6 vs. 18.6%) (p=0.003) and the proportion of immature spermatozoa (median 11.2 vs. 9.9%, p=0.001). Sperm DNA damage negatively correlated with total motility and progressive motility (r=-0.611, -0.299; p<0.001). The negative correlation with vitality, normal morphology and acrosome-intact sperm (r=-0.522, -0.550 and -0.511, respectively) was also significant (p<0.001). The examination of the same cohort of city policemen at the end of a period of high air pollution and at the end of relatively low exposure reduced the effects of age, different lifestyles, different occupational exposures, localities and genetic polymorphism on sperm quality impairment associated with air pollution. This study did not demonstrate impaired standard semen parameters in association with exposure. It was shown that sperm chromatin damage and the percentage of immature sperm were highly sensitive to air pollution.

Proteins involved in mitochondrial metabolic functions and fertilization predominate in stallions with better motility

Even in stallions with sperm quality within normal reference ranges at ejaculation, subtle differences in sperm quality exist that in many cases lead to reduced time frames for conservation of the ejaculate and/or reduced fertility. The spermatozoon is a cell highly suitable for proteomics studies, and the use of this technique is allowing rapid advances in the understanding of sperm biology. The aim of the present study was to investigate differences among stallions of variable sperm quality (based on motility and sperm velocities), although all horses had sperm characteristics within normal ranges. The proteome was studied using UHPLC/MS/MS and posterior bioinformatic and enrichment analysis; data are available via ProteomeXchange with identifier PXD025807. Sperm motility, linear motility and circular, straight line and average velocities (VCL, VSL, VAP) were measured using computer assisted sperm analysis (CASA). In stallions showing better percentages of motility, circular and average velocity predominated mitochondrial proteins with roles in the Citric acid cycle, pyruvate metabolism and oxidative phosphorylation. Interestingly, in stallions with better percentages of total motility, sperm proteins were also enriched in proteins within the gene ontology (G0) terms, single fertilization (G0: 0007338), fertilization (G0: 0009566), and zona pellucida receptor complex (GO:0002199). The enrichment of this proteins in samples with better percentages of total motility may offer a molecular explanation for the link between this parameter and fertility. SignificanceProteomic analysis identified a high degree of specificity of stallion sperm proteins with discriminant power for motility, linear motility, and sperm velocities (VCL, VAP and VSL). These findings may represent an interesting outcome in relation to the molecular biology regulating the movement of the spermatozoa, and the biological meaning of the measurements that computer assisted sperm analysis (CASA) provide. Of a total of 903 proteins identified in stallion spermatozoa, 24 were related to the percentage of total motility in the sample; interestingly, gene ontology (G0) analysis revealed that these proteins were enriched in terms like single fertilization and fertilization, providing a molecular link between motility and fertility. Field studies indicate that the percentage of total motility is the CASA derived parameter with the best correlation with fertility in stallions.

Does COVID-19 Worsen the Semen Parameters? Early Results of a Tertiary Healthcare Center

Introduction: The coronavirus disease 2019 (COVID-19) is a global pandemic which may affect multiple organs and systems including testes and disrupt the gonadal functions. The current study aimed to evaluate the effect of COVID-19 on the semen parameters and sex-related hormone levels in infertile men. Methods: The study included 21 patients who were evaluated in Ankara City Hospital, Andrology Clinic, for male infertility and have had the diagnosis of COVID-19. All the patients were evaluated in terms of semen parameters. The follicle-stimulating hormone, luteinizing hormone, and testosterone (T) levels were also evaluated in 8 of the patients. The results were presented through 2 dependent group analyses, based on the data of the patients collected before and after the diagnosis of COVID-19. Results: None of the patients needed to be hospitalized at any time through the course of COVID-19. There was a significant decrease in semen volume, percentage of total motility, percentage of progressive motility, and normal sperm morphology after COVID-19 (3 [1–8] vs. 2.5 [1.5–5], p = 0.005; 48.6 ± 22.1 vs. 34.7 ± 20.7, p = 0.001; 35.1 ± 21.7 vs. 21.8 ± 15.9, p < 0.001; 6 [3–24] vs. 5 [3–18], p = 0.015; respectively). There was also a significant decline in T level of the patients after the diagnosis of COVID-19 (350.1 ± 115.5 vs. 289.8 ± 103.3, p = 0.009). Conclusion: COVID-19 may have unfavorable effects on the gonadal functions and may lead to further deterioration of the semen parameters in infertile men, which should be considered through the evaluation for infertility.

The effect of Quercetin on the quality of sperm parameters in frozen-thawed semen of patients with Asthenospermia.

The study has aimed to investigate the effect of Quercetin, as a potent antioxidant, on preventing the negative effects of freezing process on sperm quality of patients with Asthenospermia. Semen sample from 25 patients was randomly divided into three groups; fresh, control and Quercetin (50μM). Seven days after freezing, samples were thawed at ambient temperature. Total motility, progressive sperm motility, normal morphology, viability and DNA integrity were evaluated according to WHO criteria, Papanicolaou, eosin- nigrosine and acridine orange staining respectively. In addition, the health of sperm membrane and mitochondrial membrane potential was assessed by HOS test and rhodamine staining. MDA and antioxidant enzyme activity were also evaluated using ELISA method. In contrast to the fresh group, the mean level of MDA and DNA damage had significant increase in the control group and decreased significantly sperm quality (p≤0.001). The mean percentage of total motility and progressive motility, normal morphology, viability and antioxidant enzyme activity had significant increase in the Quercetin group than the control group. In the Quercetin group, the MDA level and DNA damage also had significant reduction in comparison with the control group (p≤0.001). Therefore, the Quercetin supplementation improves the quality of cryopreserved human semen.

Seasonal effect on semen availability and quality of racing pigeon in Thailand.

Background and Aim:Seasonal variations among geographical regions could influence pigeon semen quality differently. This study aimed to determine the seasonal effect on semen availability and quality of racing pigeons in Thailand to understand and improve breeding management in the country.Materials and Methods:Semen was collected from six fertile captive pigeons once a week during summer (March-June), monsoon (July-October), and winter (November-February) during 2019-2020. The success rate of semen collection and semen quality was determined in each season – by which changes in average temperature, humidity, and photoperiod were obtained.Results:Comparable success rates of semen collection were acquired among different seasons, while varied semen qualities were revealed. The percentages of total motility and progressive motility score of sperm were significantly lowest in summer (66.35±3.40 and 3.88±0.15, respectively) compared to monsoon (85.45±2.91 and 4.67±0.10, respectively) and winter (79.29±1.96 and 4.37±0.10, respectively), while its concentration (×109 sperm/mL) and outputs (×106 sperm) were significantly highest in winter (7.62±0.54 and 91.44±10.83, respectively) compared to summer (4.23±0.41 and 48.45±6.35, respectively) and monsoon (3.57±0.30 and 51.45±7.21, respectively). Besides, semen samples collected from birds housing at an average temperature of <29.5°C demonstrated better sperm motility sperm concentration and total sperm counts than those from at a higher temperature.Conclusion:Winter was regarded as the best season contributing the best semen quality, while summer was the worst. Due to the fluctuation of temperature during summer and winter, the seasonal temperature was implied as the major factor contributing to changes in sperm quality of racing pigeons in Thailand.

Zika RNA and Flavivirus-Like Antigens in the Sperm Cells of Symptomatic and Asymptomatic Subjects.

Zika virus (ZIKV) RNA has been found to remain in human semen for up to one year after infection, but the presence of Flavivirus antigens in the different compartments of semen has been largely unexplored. Following the introduction of ZIKV in Nicaragua (2016), a prospective study of patients with clinical symptoms consistent with ZIKV was conducted in León to investigate virus shedding in different fluids. ZIKV infection was confirmed in 16 male subjects (≥18 years of age) by RT-qPCR in either blood, saliva or urine. Of these, three provided semen samples at 7, 14, 21, 28, 60 and 180 days postsymptom onset (DPSO) for Flavivirus antigens and RNA studies. These cases were compared with 19 asymptomatic controls. Flavivirus antigens were examined by immunofluorescence (IF) using the 4G2 Mabs, and confocal microscopy was used to explore fluorescence patterns. The three (100%) symptomatic subjects and 3 (16%) of the 19 asymptomatic subjects had Flavivirus antigens and viral RNA in the spermatozoa fraction. The percentage of IF Flavivirus-positive spermatozoa cells ranged from 1.9% to 25% in specimens from symptomatic subjects, as compared with 0.8% to 3.8% in specimens from asymptomatic controls. A marked IF-pattern in the cytoplasmic droplets and tail of the spermatozoa was observed. The sperm concentrations (45 × 106/mL vs. 63.5 × 106/mL, p = 0.041) and the total motility percentage (54% vs. 75%, p = 0.009) were significantly lower in specimens from ZIKV-positive than in those of ZIKV-negative. In conclusion, this study demonstrated the presence of Flavivirus antigens and RNA within a time frame of 28 DPSO in sperm cells of symptomatic and asymptomatic subjects during the ZIKV epidemic. These findings have implications for public health, in terms of nonarthropod-born, silent transmission facilitated by sperm cells and potential transmission from asymptomatic males to pregnant women, with consequences to the fetus.

Effects of NGF Addition on Llama (Lama glama) Sperm Traits After Cooling.

To provide new insights into the mechanisms through which seminal plasma proteins can protect sperm from damage caused during refrigeration, we evaluate the possibility that β-NGF can contribute to the improvement of sperm quality after cooling. First, β-NGF was detected in refrigerated sperm and compared with unrefrigerated sperm by western blotting of the proteins adsorbed by sperm, showing that native β-NGF is still present even 24 h after cooling only as an active form. Then, the effect of exogenous β-NGF on sperm quality after cooling was evaluated. A total of 12 ejaculates from male llamas (three ejaculates per male), were obtained by electro-ejaculation, diluted 4:1 with buffer Hepes-balanced salt solution and centrifuged at 800 × g for 8 min to remove the seminal plasma. Sperm were suspended in Tris-citrate-fructose-egg yolk diluent for a final concentration of 30 ×106/ml and cooled at 5°C for 24 h. After refrigeration, the extended sperm were equilibrated for 5 min at 37°C and divided into the following subgroups: sperm samples without treatment (control) and sperm samples supplemented with exogenous human β-NGF (10, 100, and 500 ng/ml). At 5, 30, and 60 min of incubation sperm were evaluated for sperm viability (using eosin/nigrosin stain), sperm motility and vigor (observed under light microscopy), and mitochondrial activity (using the JC-1 fluorescent marker). Vigor data were analyzed with the nonparametric Kruskal-Wallis test. The rest of the variables were analyzed with a mixed models approach. Mean comparisons were performed using Fisher's LSD test with a confidence level of 95%. A principal components analysis was performed to analyze the relationships between variables. Treatment of 24 h cooled sperm with 10 or 100 ng/ml of human β-NGF increased the percentage of total motility and vigor (p < 0.05). Besides, an incubation time of 60 min would be adequate to improve sperm quality, since all variables are positively related. The significant improvement observed in the motility and vigor of post-refrigerated sperm suggests that supplementation with exogenous β-NGF may be profitable for the improvement of cooled llama sperm.

Effect of Repeated Freezing and Thawing on Nguni Sperm Parameters Evaluated by Computer Assisted Sperm Analyzer System

The aims of the current study were to assess the effect of double freezing and thawing on Nguni semen parameters. Semen was collected from Nguni bulls. The Sperm Class Analyzer® system was used to evaluate sperm motility rates prior and post freezing, repeatedly. Semen samples were diluted with egg yolk citrate extender. Cooled semen samples were loaded into straws and placed inside the programmable freezer for freezing. The frozen semen straws were stored inside liquid nitrogen tank. The frozen semen straws were then thawed at 15°C (1st and 2nd freezing and thawing followed the same process) for a duration of 5 min and further assessed post-thawed at 0 and 15 min during in vitro liquid storage at 15°C. The treatment means were analyzed using Fisher’s protected t-test least. There was no significant differences recorded on raw semen sperm total motility (93.2%) and 1st frozen-thawed at 0 min (82.6%), with recovery rate of 88.6% of the sperm total motility. There was a drastic decline observed in sperm total motility during the 1st frozen-thaw at 15 min (77.6%), 2nd frozen-thaw at 0 min (31.3%) and 2nd frozen-thaw at 15 min (30.1%; P<0.05). In conclusion, the freezing and thawing process did not decrease the bull’s sperm total motility percentage during the 1st freezing and thawing process, as compared with raw semen. However, a drastic decline was observed during the 2nd freezing-thawing process, as compared with raw semen.

Protective effects of Tribulus terrestris and Cinnamomum zeylanicum extracts and trehalose added to diluents on goat epididymal sperm freezability

This study investigates the effect of adding Tribulus terrestris ethanol extract (TEE) and Cinnamomum zeylanicum ethanol extract (CEE) and trehalose on freezability of goat epididymal spermatozoa. In Experiment 1, the treatments consist of basic extender containing 25, 50 or 100 μg/ml of TEE or CEE. The control contained no additives. Experiment 2 was carried out to compare the effect of best concentrations resulted in the first experiment with 150 mM trehalose added to basic extender. The results of experiment 1 showed that supplementation of 50 μg/ml TEE and 50 μg/ml CEE increased significantly the percentages of motility, progressive motility and viability of cryopreserved spermatozoa, while the level of malondialdehyde concentration was decreased. Moreover, the 50 μg/ml TEE treatment indicate significantly) P < 0.05) the lowest DNA fragmentation among the other treatments. The data obtained from experiment 2 show that all treatments increased significantly) P < 0.05) the percentages of total motility, viability and membrane integrity, and concurrently decreased the rate of MDA compared to control. In addition, the rates of viability and progressive motility were significantly (P < 0.05) higher in diluents contained herb extracts and trehalose. Regarding DNA fragmentation, the results demonstrate that using the extracts and trehalose in diluents decreased the DNA damages and thereby improved the rate of intact sperm heads. In conclusion, the results of this study indicate that 50 μg/ml of Tribulus terrestris and Cinnamomum zeylanicum ethanolic extracts alone and plus trehalose improved the spermatozoa quality and could be used for cryopreservation.

Evaluation of used Purslane extracts in Tris extenders on cryopreserved goat sperm

This study aimed to evaluate the comparative effects of Purslane aqueous extract (PAE), Purslane methanolic extract (PME) and Purslane ethanolic extract (PEE on the quality of frozen-thawed goat spermatozoa. Collected semen with motility >75% and sperm concentration >1.0 × 109 sperm/ml was pooled and divided into 10 equal aliquots and supplemented by basic extender containing 25, 50 or 100 μg/ml of Purslane aqueous extract (PAE25μg/ml, PAE50μg/ml, PAE100μg/ml, respectively), basic extender containing 25, 50 or 100 μg/ml of Purslane methanolic extract (PME25μg/ml, PME50μg/ml, PME100μg/ml, respectively), basic extender containing 25, 50 or 100 μg/ml of Purslane ethanolic extract (PEE25μg/ml, PEE50μg/ml, PEE100μg/ml, respectively). Control diluent contained no additives. For the determination of sperm quality, frozen straws were thawed and then the sperm characteristics were assessed. Results indicated that higher (P < 0.05) percentages of total motility, viability, mitochondrial activity and lower percentages of malondialdehyde (MDA) for PAE50μg/ml, PME50μg/ml and PEE50μg/ml than those of the control. In addition, PME50μg/ml resulted in the highest) P < 0.05) total motility and the lowest (P < 0.05) MDA levels compared to other treatments. Compared to the control group, PME50μg/ml resulted in higher integrity (P < 0.05) of plasma membranes and in lower amounts of apoptotic and dead spermatozoa. PME50μg/ml and PAE50μg/ml showed higher (P < 0.05) percentages of progressive motility, DNA integrity and live post-thawed spermatozoa than those of the control. No significant differences in the motility, viability, mitochondrial activity and number of live sperms were observed between PME50μg/ml and PAE50μg/ml treatments. In conclusion, the results of this study indicated that 50 μg/ml purslane extracts could be used for the cryopreservation. However, the results of methanolic extract was more beneficial compared to other extracts.

Single-Layer Colloid Centrifugation as a Method to Process Urine-Contaminated Stallion Semen After Freezing-Thawing

Urospermia is a major ejaculatory dysfunction affecting stallions. It has been thought that urine-contaminated semen should not be cryopreserved; however, on select cases, urine contamination of semen cannot be avoided. A recent study suggested that urospermic semen can be cryopreserved after cushion centrifugation and extension. Thus, this study aimed to assess the use of single-layer colloid centrifugation (SLC) to process frozen-thawed urine-contaminated stallion semen. Raw ejaculates (n = 55) from eight stallions were split into three groups: no urine, low (20%), or high (50%) urine contamination. Semen was extended 1:1, cushion-centrifuged, and resuspended at 200 million sperm/mL in BotuCrio. Resuspended semen was loaded in 0.5 mL straws and cryopreserved in liquid nitrogen. Samples were thawed (37°C for 30 seconds) and processed by SLC (400 g/30 minutes). Percentages of total motility (TM) and progressive motility (PM) were assessed with computer-assisted semen analyzer. Sperm viability (%VIAB) and yield were assessed with a NucleoCounter before and after gradient centrifugation. Data were analyzed with two-way ANOVA and Tukey’s test. The motility parameters TM before SLC (control: 35 ± 2; low: 33 ± 0.7; high: 22 ± 1.8) after SLC (control: 51 ± 3.6; low: 42 ± 2.2; high: 25 ± 2.8) and PM before SLC (control: 24 ± 1.8; low: 21 ± 1.14; high: 12 ± 1.5) and after SLC (control: 40.3 ± 3.2; low: 31 ± 3.9; high: 14 ± 2) significantly decreased with increasing urine contamination. Urine contamination marginally reduced (P < .05) sperm viability after cryopreservation before SLC (control: 45 ± 0.7; low: 27 ± 0.2; high: 27 ± 0.3) and after SLC (control: 54 ± 0.5; low: 49 ± 0.7; high: 38 ± 0.6). Recovery rates of sperm after centrifugation were not significantly different between groups. In conclusion, urine contamination affects sperm motility parameters in a dose-dependent manner. Post-thaw SLC selected sperm with higher motility and viability in control and low groups but only selected sperm with higher viability in the high group.

Association of seminal angiotensinogen with sperm motility and morphology in male infertility.

Many researchers have shown that renin-angiotensin system (RAS) is involved in various important aspects of male reproduction. In this study, we assessed whether abnormal levels of seminal angiotensinogen (AGT) may be associated with semen parameters in infertile males. A total of 115 male patients were recruited, and semen parameters, seminal AGT and the electrolytes including K+ , Na+ , Cl- , P and Ca were evaluated. According to the World Health Organization (WHO) 2010 criteria, the patients were divided into two groups: G1 group with normal semen parameters (n=42) and G2 group with subnormal semen parameters (n=73). The level of seminal AGT was significantly higher in G2 group compared with G1 group. Moreover, the level of AGT was negatively correlated with the percentage of total motility (r=-.322, p=.000), progressive motility (PR) (r=-.339, p=.000) and morphologically normal forms (r=-.263, p=.004). This study suggests that elevated seminal AGT level is associated with increased risk of asthenospermia and teratozoospermia.

Electromagnetic field in human sperm cryopreservation improves fertilizing potential of thawed sperm through physicochemical modification of water molecules in freezing medium.

Physicochemical properties of water molecules as the main compositions of the freezing media can be affected by the electromagnetic fled. The purpose of this study was to apply extremely low repetition rate electromagnetic fields (ELEFs) to change the molecular network of water molecules existing in freezing media used for human sperm cryopreservation. First, different time periods and pulsed electromagnetic fields were used to evaluate the physiochemical properties of water. The lowest rate of cluster size, surface tension, viscosity, and density was observed for water samples exposed to 1000 Hz ELEF for 60 min (P < 0.05) that could be results in small ice crystal formation. Therefore, this treatment was selected for further evaluations in human sperm freezing because there was minimal probability of amorphous ice crystallization in this group. To assess fertilizing potential, human semen samples were subjected to ELEF (1000 Hz) water-made freezing medium and cryopreserved. The highest percentage of total motility, progressive motility, viability, membrane integrity, mitochondrial membrane potential, DNA integrity, and TAC were obtained in frozen ELEF as compared to other groups. The percentage of viable spermatozoa (Annexin V-/PI-) in frozen ELEF was significantly higher than in frozen control. The level of ROS was significantly lower in frozen ELEF when compared to frozen control. It can be concluded that the modification of physicochemical properties of water existing in cryopreservation media by ELEF is a suitable strategy to improve the outcome of cryopreservation.

Effect of Mare Colostrum in Extenders for Freezing Stallion Semen

This study aimed to evaluate the addition of mare colostrum in stallion freezing extenders to improve sperm quality. First, colostrum samples were collected from four mares after the foal's birth and their composition was determined. Ejaculates were collected from nine fertile stallions. Sperm samples were pooled, diluted, and cryopreserved into three experimental extender groups: Lactose-based extender supplemented with mare colostrum (20%), lactose-based extender supplemented with egg yolk (20%), and BotuCrio. The quality of the post-thaw semen samples were evaluated assessing sperm motility by means of computer-assisted analysis, viability by SYBR-14 and propidium iodine (PI) stain, acrosome integrity by fluorescein isothiocyanate and peanut agglutinine (FITC-PNA) and PI stain, plasma membrane functionality by hypo-osmotic swelling (HOS) test, and DNA denaturation by acridine orange (AO) test. There were no significant differences in the percentages of total motility, acrosome integrity, and DNA fragmentation among the extenders after thawing. Kinematics parameters showed significantly higher values in BotuCrio than in lactose extenders (P < .05). BotuCrio and lactose colostrum extender yielded significantly better rates for HOS-test, linearity, straightness, and wobble than egg-yolk extender (P < .05). However, in relation to sperm viability, lactose egg yolk extender showed significantly better results in comparison to the others seminal experimental media (P < .05). In conclusion, the incorporation of mare colostrum into cryopreservation media protected the sperm against cold-shock; therefore, it may be a good cryoprotectant agent alternative in extenders for freezing stallion semen.

Improvement on the efficiency of doses per conception by using a semen extender in timed artificial insemination

This study evaluated the efficiency of a post-thaw diluent on the conception rate of timed artificial insemination (TAI), on the in vitro embryo production (IVEP), the fertilization rate and the computerized semen analysis. In the execution of all the techniques, semen straws of a single bull with previously known fertility were evaluated both as control and extender groups. The conception rate was similar (P = 0.517) between the extender (52.11%) and the control group (55.48%), but the extender group showed a reduction (P < 0.001) in the number of doses per conception (0.95 for the extender vs. 1.80 for the control). We evaluated the rates of cleavage (81.56 vs. 78.52%), blastocyst formation (36.10 vs. 39.48%) and hatching (47.28 vs. 41.97%) on the IVEP and assessed the fertilization rate by counting pronuclei (61.76 vs. 65.22%). Importantly, we found that these measures did not differ (P > 0.05) between the extender and control group, respectively. The total (58.1 vs. 58.3%) and progressive (42.4 vs. 40.0%) motility post-thawing did not differ (P > 0.05) between the extender vs. the control group, respectively. However, when comparing different moments post-thawing (0 h, 10 min, 3 h, 6 h, 8 h e 12 h), initially (3 h) the extender group had a higher (P < 0.05) percentage of total motility (58.7 vs. 48.0%). Subsequently (8 h), the motility decreased (P < 0.05) in comparison to the control group (7.3 vs. 22.6%). In conclusion, the use of extender post-thawing was effective in TAI, reducing the number of doses per conception but not influencing the IVEP. Therefore, this extender allowed for the more efficient use of semen of high genetic value, reducing the cost per pregnancy.

Evaluation of a portable device for assessment of motility in stallion semen.

ContentsIn horse breeding, quality assessment of semen before insemination is often requested. Non‐laboratory‐based techniques for objective analysis of sperm motility are thus of interest. The aim of this study was evaluating a portable device for semen analysis (Ongo sperm test) and its comparison with computer‐assisted semen analysis (CASA). Semen was collected from 10 stallions, diluted to 100, 50 and 25 × 106 sperm/ml and analysed for total (TM) and progressive motility (PM). The final sperm concentration influenced total motility analysed by Ongo (p < 0.05) which was higher at 100 × 106 sperm/ml when compared to 25 × 106 sperm/ml (p < 0.05) but not when compared to 50 × 106 sperm/ml (n.s.). Sperm concentration did not influence total motility when assessed by SpermVision (n.s.). Agreement between methods was evaluated by correlation analysis and Bland–Altman plot. Intra‐assay variation of Ongo was 5.2% ± 3.0 for TM and 6.9% ± 3.4 for PM. Correlation between Ongo and CASA was r = 0.79, 0.88 and 0.83 for 100, 50 and 25 × 106 sperm/ml for TM, and r = 0.87, 0.89 and 0.87 for PM, respectively (all p < 0.001). At the 100 and 25 mio/ml dilutions, the difference between the two systems deviated significantly from 0, while no such bias existed at the 50 mio/ml dilution (TM Ongo 85.0%, CASA 82.3%; PM Ongo 64.1%, CASA 66.1%). The 95% confidence interval was 19.9%, 18.9% and 19.2% ± mean for TM and 20.7%, 17.4% and 20.3% ± mean for 100, 50 and 25 × 106 sperm/ml, respectively. In conclusion, Ongo sperm test sperm motility data were strongly correlated with data obtained by CASA. In addition, at a concentration of 50 × 106 sperm/ml values measured with both systems were close to identical. At this concentration, which is recommended in equine AI, Ongo and CASA can be used interchangeably.

Increased cryo-survival rate in ejaculated human sperm from infertile men following pre-freeze in vitro myo-inositol supplementation.

ObjectiveTo investigate the effects of in vitro myo-inositol (Myo-Ins) supplementation of cryopreserved human semen on the cryo-survival rate (CSR).MethodsSemen samples were obtained from 41 infertile men. Following routine semen analysis, each sample was divided into two equal aliquots (0.5 mL each). One aliquot was treated with 1 mg of Myo-Ins dissolved in 10 µL of sperm preparation medium. The second aliquot was treated with 10 µL of the same medium (control). Both aliquots were incubated for 20 minutes prior to freezing to slow the freezing process. The frozen samples were examined for post-thaw percentages of total motility (TM), progressive motility (PM), and the CSR, defined as the percentage of post-thaw TM divided by the percentage of pre-freeze TM and multiplied in 100. The results were expressed as median and interquartile range (25th and 75th percentiles).ResultsThe pre-freeze TM (50% [30%–50%]) and PM (35% [20%–35%]) were significantly higher than the post-thaw TM and PM in the Myo-Ins group (15% [10%–35%] and 10% [5%–20%]; p<0.001 and p<0.001, respectively) and the control group (10% [6%–30%] and 5% [3%–15%]; p<0.001 and p<0.001, respectively). The CSR of the 41 semen aliquots supplemented with Myo-Ins (40% [25%–70%]) was significantly higher than that of the control samples (30% [13%–58%], p=0.041). The CSR of the 26 abnormal semen samples that were supplemented with Myo-Ins (38% [20%–50%]) was significantly higher than that of the control samples (23% [12%–30%], p=0.031).ConclusionIn vitro Myo-Ins supplementation of ejaculated human sperm from infertile men resulted in a significant increase in the CSR in samples with abnormal pre-freeze sperm parameters.

Effects of pre-freeze Nigella sativa oil supplementation on cryosurvival of ovine spermatozoa.

The study was designed with three experiments to evaluate the effects of pre-freeze supplementation of Nigella sativa oil (NSO) and thymoquinone (TQ) on total motility, progressive motility, biokinetic characteristics, acrosomal integrity and DNA integrity of cryopreserved ovine spermatozoa. Semen samples collected from three proven fertile Merino rams were diluted with a Tris-based cryomedia containing different levels of NSO (Experiment I: 0, 10, 100 and 1,000g/ml), TQ (Experiment II: 0, 1, 10 and 20g/ml) and their optimum levels (Experiment III: 100g/ml of NSO, 10g/ml of TQ and 1mM of α-tocopherol and cryopreserved as pellet (200µL) and subsequently evaluated at different post-thaw incubation periods (0, 2 and 4hr). The results revealed that the percentage of total motility, progressive motility and biokinetic characteristics such as average path velocity, curvilinear velocity and straight-line velocity were higher (p<0.05) in the sperm aliquots cryopreserved with 100g/ml NSO or 10g/ml TQ than in the sperm aliquots cryopreserved without supplementation just after thawing and 2hr of post-thaw incubation. Among the supplements, NSO (100g/ml) showed higher values of the total motility, progressive motility, biokinetic characteristics specially, average path velocity, curvilinear velocity and straight-line velocity, acrosome integrity and DNA integrity compared with the spermatozoa frozen without supplementation. Therefore, the results suggest that NSO may be added to the cryomedium to improve the cryosurvival of ovine spermatozoa.

Dietary fish oil supplemented with vitamin E improves quality indicators of rooster cold-stored semen through reducing lipid peroxidation

Cockerel semen is sensitive to cooling, which limits chilled storage of semen for more than 24 h. Results of artificial insemination with cold-stored semen are not desirable. This study was conducted to evaluate the effects of dietary fish oil and vitamin E (vitE) for cold-storage of rooster semen and its effects on parameters of semen during 48 h cooling preservation. Roosters were assigned into four dietary treatments; 1) control group received a basal diet, 2) vitE group received a basal diet supplemented with 200 mg/kg vitE, 3) fish oil group (FO) received a basal diet supplemented with 2% fish oil and 4) fish oil and vitE group received a basal diet supplemented with 2% fish oil and 200 mg/kg vitE (FO + vitE). Semen samples were collected after 40 days of feeding and then diluted and cooled to 5 °C for preservation up to 2 days. Several quality indicators of sperm such as motion characteristics, membrane integrity, and viability, and abnormal morphology, activity of mitochondria, lipid peroxidation and acrosome integrity of the sperm were assessed at different times of storage (0, 24 and 48 h). None of sperm were significantly affected by the diets at the start of storage (0 h, p > 0.05). FO and FO + vitE improved the percentage of total motility, viability, and mitochondria activity at 24 h (P ≤ 0.05). After 48 h, only FO + vitE group produced the higher percentage of total motility, viability and membrane integrity (P ≤ 0.05). Lipid peroxidation was significantly reduced in sperm obtained from roosters fed diets of FO + vitE and vitE compared to FO and control (P ≤ 0.05) at times of 24 and 48 h. There was no significant difference between control and vitE groups in none of parameters (P > 0.05). Integrity of acrosome and abnormal morphology were not significantly affected by the diets (P > 0.05). Supplementation of roosters' diet with 2% fish oil and 200 mg/kg vitamin E improved the quality of cold-stored semen by supporting several indicators of sperm quality through reducing lipid peroxidation.

Induction of Sublethal Oxidative Stress on Human Sperm before Cryopreservation: A Time-Dependent Response in Post-Thawed Sperm Parameters.

Objective A recent innovative approach, based on induction of sublethal oxidative stress to enhance sperm cryosurvival, has been applied before sperm cryopreservation. The purpose of this study was to investigate the effects of different induction times of sublethal oxidative stress before cryopreservation on human post-thawed sperm quality. Materials and Methods In this experimental study, we selected semen samples (n=20) from normozoospermic men according to 2010 World Health Organization (WHO) guidelines. After processing the samples by the density gradient method, we divided each sample into 5 experimental groups: fresh, control freezing, and 3 groups exposed to 0.01 μM sodium nitroprusside (SNP) [nitric oxide (NO) donor] for 30 (T30), 60 (T60), or 90 minutes (T90) at 37˚C and 5% CO2 before cryopreservation. Motion characteristics [computer-assisted sperm analyser], viability, apoptosis [annexin V/propidium iodide (PI) assay], DNA fragmentation [sperm chromatin structure assay (SCSA)], and caspase 3 activity (FLICA Caspase Detection Kit) were assessed after thawing. The results were analysed by using one-way ANOVA and Tukey’s test. The means were significantly different at P<0.05. Results Cryopreservation significantly decreased sperm viability and motility parameters, and increased the percentage of apoptosis, caspase 3 activity, and DNA fragmentation (P<0.01) compared to the fresh group. The T60 group had a higher significant percentage of total motility (TM) and progressive motility compared with other cryopreserved groups (P<0.05). We observed a significantly lower percentage of apoptotic rate and caspase 3 activity in the T60 group compared to the other cryopreserved groups (P<0.05). DNA integrity was not significantly affected by this time of sublethal stress induction (P>0.05). ConclusionOur results have demonstrated that the application of sublethal oxidative stress by using 0.01 μM NO for 60 minutes before the freezing process can be a beneficial approach to improve post-thawed human sperm quality.