Increased Sperm Motility Research Articles

Temporal Trends in Semen Quality, Hormone Levels, and Substance Use Among Infertile Men in Pre- and Post-Cannabis Legalization Eras in Canada.

Background: The Cannabis Act (Bill C-45) was enacted in 2018, to legalize and regulate the use, production, and sale of nonmedical cannabis in Canada. While public health and safety implications of cannabis legalization have yet to be elucidated, the wide availability of cannabis necessitates health care providers to be knowledgeable about therapeutic potential and side effects of use. This study aimed to examine the temporal trends over two decades and the impact of the Cannabis Act in Canada, implemented in October 2018, on substance use, semen parameters, and testosterone levels of infertile men. Methods: We conducted a retrospective cohort study from a prospectively maintained database of a single infertility clinic. Demographic, fertility, and substance use history were correlated with semen and hormone assessments. Temporal trends in cannabis use and semen quality between 2001 and 2021 were investigated and compared between pre-cannabis legalization eras (PRCL) and post-cannabis legalization eras (POCL). Results: Our cohort included 11,630 patients (9411 PRCL and 2230 POCL). Cannabis use increased by 8.4% per year (p<0.001), while alcohol and tobacco consumption declined (0.8% and 1.5% per year, p<0.05 and p=0.004, respectively). Similar trends were noticed in the POCL, with higher rates of cannabis use (22.4% vs. 12.9%, p<0.001) and decreased tobacco and alcohol intake (15.2% vs. 17.7%, p=0.005 and 50.5% vs. 55.2%, p<0.001, respectively) compared to the PRCL group. Semen concentration was lower in the POCL group (24.8±44.8 vs. 28.7±48.3 million/mL, p=0.03). Testosterone did not differ between the cohorts. Comparison between cannabis users (n=1715) and nonusers (n=9924) demonstrated a slight increase in sperm motility (25.9%±15.3% vs. 23.9%±15.0%, p=0.002) and decreased sperm concentration among users (27.6±53.5 vs. 23.9±15.0 million/mL, p=0.03). Conclusion: A nearly 10% rise in cannabis use in the POCL era was observed among men being investigated for infertility. Our data suggest cannabis use may be associated with an increase in testosterone, slightly improved sperm motility, and decreased sperm concentration.

Roles of Y-27632 on sheep sperm metabolism.

To investigate the effect of Y-27632 on low-temperature metabolism of sheep sperm, different concentrations of Y-27632 were added to sheep semen at 4°C in this experiment to detect indicators such as sperm motility, plasma membrane, acrosome, antioxidant performance, mitochondrial membrane potential (MMP), and metabolomics. The results showed that the addition of 20µM Y-27632 significantly increased sperm motility, plasma membrane integrity rate, acrosome integrity rate, antioxidant capacity, MMP level, significantly increased sperm adenosine triphosphate (ATP) and total cholesterol content, and significantly reduced sperm Ca2+ content. In metabolomics analysis, compared with the control group, the 20µM Y-27632 group screened 20 differential metabolites, mainly involved in five metabolic pathways, with the most significant difference in Histidine metabolism (P = 0.001). The results confirmed that Y-27632 significantly improved the quality of sheep sperm preservation under low-temperature conditions.

In vitro impact of ethanolic extract of Bryonia laciniosa seed on Gir bull spermatozoa: a comprehensive evaluation through transcriptome profiling.

This study examines the in vitro impact of an ethanolic extract derived from Bryonia laciniosa seeds on the Gir bull (Bos indicus) spermatozoa. The objective is to thoroughly assess the effects of the seed extract on the physiological parameters of bull spermatozoa, followed by evaluating its effects on X and Y-bearing spermatozoa and its impact on gene expression through transcriptome profiling. For this study, one Gir bull was selected, and 12 ejaculates were collected at one-week time intervals. Sperm cells were isolated from each ejaculate and incubated with varying concentrations of the ethanolic extract. The physiological parameters of the spermatozoa were assessed using Computer Assisted Semen Analysis (CASA) and compared with control groups to evaluate the extract's effects on sperm quality and motility. At a concentration of 18 mg/mL B. laciniosa extract, we noticed a statistically significant 16.4% increase in sperm motility (p = 0.0065). In order to understand the specific effect on X and Y-bearing spermatozoa, motile and non-motile sperm separated by glass wool column method and further evaluated for quantification of X and Y-bearing sperm in all samples by ddPCR. To understand the effect of B. laciniosa extract on spermatozoa at the molecular level, whole transcriptome profiling was carried out using Illumina MiSeq. Transcriptome profiling revealed 81 genes that were expressed differently between the group treated with the extract and the control group. The current investigation revealed an increase in the expression of TLX1, CRYGB, KLF13, and ZAR1 transcripts, which play a role in embryonic development. In addition, several genes have been identified that are involved in sperm motility, such GSK3B, LAPRS, MAPK1, CAMK2B, and AQP7. The findings exhibited the therapeutic effectiveness of B. laciniosa seeds in augmenting fertility through a synergistic blend of activities, including enhanced sperm motility and positive influence on embryogenesis.

Tepung Labu Kuning (Cucurbita moschata), Tepung Porang (Amorphophallus muelleri Blume), dan Kombinasi Kedua Tepung sebagai Penurun Gula Darah dan Peningkat Motilitas Spermatozoa Mencit (Mus musculus)

Diabetes Mellitus (DM) is a global epidemic with high morbidity and mortality. Diabetes Mellitus (DM) is thought to have a negative impact on sperm quality. Yellow pumpkin contains antioxidants that can neutralize free radicals which can improve fertility. Glucomannan is a substance contained in porang which functions as a soluble fiber and plays a role in blood glucose balance. This study aims to examine pumpkin flour, porang flour and a combination of both flours on sperm motility and body weight of alloxan-induced diabetic mice. This study used 25 mice aged 3 months, body weight 25gr-30 gr. This study used a completely randomized plan (CRD) with 5 replications and 5 treatments. The treatments were negative control (diabetic mice), positive control (administration of glimepiride at a dose of 0.0078 mg/mice per day), D1 (pumpkin flour 134.4 mg/30g BW of mice), D2 (porang flour 63mg/30g BW mice), D3 (combination 50% : 50% = 67.2 mg : 31.5 mg/30 grBW mice). Diabetic mice were made by intraperitoneal induction of alloxan dose of 200 mg/Kg for 3 days. The parameters observed were spermatozoa motility, body weight and blood sugar levels of the mice. The research data were analyzed using the ANOVA test followed by the BNT test. The results showed that the administration of glymeride, pumpkin flour, porang flour and their combination had a significant effect on reducing blood sugar (p=0.000) and sperm motility (p=0.000), but had no effect on body weight (p=0.317). Conclusion: pumpkin flour, porang flour or a combination of both flours can be used as herbs to lower blood sugar and increase sperm motility in diabetes sufferers.

Supplementing semen extenders with a combination of phosphorus and vitamin B12 Improves post-thawed cryopreserved rooster semen quality.

Semen cryopreservation is an important technique for preserving the genetic material of numerous species. However, frozen semen is highly susceptible to sperm DNA damage and reduced motility, resulting in decreased fertility. The standard method for cryopreservation and several approaches have not been elucidated. This study aimed to determine the effects of supplementing rooster semen extender with a combination of phosphorus and vitamin B12 on cryopreserved semen quality. Semen was collected weekly via dorso-abdominal massage from 57 Burmese × Vietnam-crossbred Thai native roosters aged 1-3 years. In total, 139 semen samples were collected, pooled, and diluted to 200 million sperm per dose. The pooled sample was divided into six experimental groups: a control group (0.00%) diluted with modified Beltville Poultry Semen Extender (BPSE) and five treatment groups diluted with modified BPSE supplemented with phosphorus and vitamin B12 at concentrations 0.02, 0.04, 0.06, 0.08, and 0.10%, respectively. The semen samples were frozen and evaluated at 0, 15, and 30 min after thawing. Sperm kinematic parameters were determined using a computer-assisted sperm analysis system. Sperm quality was evaluated by measuring sperm viability, mitochondrial activity, acrosome integrity, and plasma membrane integrity. Statistical analyses were performed using a general linear mixed model (MIXED) in SAS. Factors in the statistical model were experimental groups, time after thawing, and interaction between experimental groups and time after thawing. Total and progressive motilities were greater in semen supplemented with 0.04% phosphorus and vitamin B12 compared with those in the control (p < 0.05). At 15 min post-thawing, VCL, VAP, and HPA in the 0.04% phosphorus and vitamin B12 supplementation group was greater than that in the control (p < 0.05). Phosphorus and vitamin B12 supplementation did not affect sperm kinematics at 0 and 30 min after thawing (p > 0.05). All the sperm parameters that were tested for the 0.04% phosphorus and vitamin B12 supplementation group in modified BPSE were the highest at all the timepoints after thawing. Thus, supplementing frozen semen extender with 0.04% phosphorus and vitamin B12 increased sperm motility, sperm kinematic parameters, and sperm quality.

Optimum Doses and Forms of Selenium Maintaining Reproductive Health via Regulating Homeostasis of Gut Microbiota and Testicular Redox, Inflammation, Cell Proliferation, and Apoptosis in Roosters

BackgroundThere is a U-shaped relationship between dietary selenium (Se) ingestion and optimal sperm quality. ObjectivesThis study aimed to investigate the optimal dietary dose and forms of Se for sperm quality of breeder roosters and the relevant mechanisms. MethodsIn experiment 1, 18-wk-old Jingbai laying breeder roosters were fed a Se-deficient base diet (BD, 0.06 mg Se/kg), or the BD + 0.1, 0.2, 0.3, 0.4, 0.5, or 1.0 mg Se/kg for 9 wk. In experiment 2, the roosters were fed the BD or the BD + sodium selenite (SeNa), seleno-yeast (SeY), or Se-nanoparticles (SeNPs) at 0.2 mg Se/kg for 9 wk. ResultsIn experiment 1, added dietary 0.2 and 0.3 mg Se/kg led to higher sperm motility and lower sperm mortality than the other groups at weeks 5, 7, and/or 9. Furthermore, added dietary 0.2–0.4 mg Se/kg produced better testicular histology and/or lower testicular 8-hydroxy-deoxyguanosine than the other groups. Moreover, integrated testicular transcriptomic and cecal microbiomic analysis revealed that inflammation, cell proliferation, and apoptosis-related genes and bacteria were dysregulated by Se deficiency or excess. In experiment 2, compared with SeNa, SeNPs slightly increased sperm motility throughout the experiment, whereas SeNPs slightly reduced sperm mortality compared with SeY at week 9. Both SeY and SeNPs decreased malondialdehyde in the serum than those of SeNa, and SeNPs led to higher glutathione peroxidase (GPX) and thioredoxin reductase activities and GPX1 and B-cell lymphoma 2 protein concentrations in the testis compared with SeY and SeNa. ConclusionsThe optimal dietary Se dose for reproductive health of breeder roosters is 0.25–0.35 mg Se/kg, and SeNPs displayed better effects on reproductive health than SeNa and SeY in laying breeder roosters. The optimal doses and forms of Se maintain reproductive health of roosters associated with regulation intestinal microbiota homeostasis and/or testicular redox balance, inflammation, cell proliferation, and apoptosis.

Low-Fat, High-Carbohydrate Diets Reduce Body Weight and Sperm Count but Increase Sperm Motility in Mice

BackgroundMale reproduction is impacted by both over- and under-nutrition, demonstrated by animal studies using high-fat and low-protein dietary interventions. Little is known about the impacts of low-fat, high-carb diets and types of dietary carbohydrates on sperm traits. ObjectivesUsing a nutritional geometry approach, we investigated the effects of partially or completely substituting glucose for fructose in isocaloric diets containing either 10%, 20%, or 30% fat (by energy) on sperm traits in mice. MethodsMale C57BL/6J mice were fed 1 of 15 experimental diets for 18 wk starting from 8 wk of age. Reproductive organs were then harvested, and sperm concentration, motility, and velocity were measured using Computer-Assisted Sperm Analysis. ResultsIncreasing dietary fat from 10% to 30% while maintaining energy density at 14.3 kJ/g and protein content at 20% resulted in increased body weight and sperm production but reduced the percentage of motile sperm. Body weight and seminal vesicle weight were maximized on diets containing a 50:50 mix of fructose and glucose, but carbohydrate type had few significant impacts on epididymal sperm traits. ConclusionsThe opposing impacts of dietary fat on mouse sperm quantity and quality observed suggest that male fertility may not be optimized by a single diet; rather, context-specific dietary guidelines targeted to specific concerns in semen quality may prove useful in treating male infertility.

Effect of Micronutrients and L-Carnitine as Antioxidant on Sperm Parameters, Genome Integrity, and ICSI Outcomes: Randomized, Double-Blind, and Placebo-Controlled Clinical Trial.

The evaluation of sperm DNA integrity is recommended in the sixth edition of the 2021 World Health Organization guidelines. Oxidative stress has been identified as a crucial factor leading to genome decay, lipid peroxidation, and nucleoprotein oxidation. This double-blind, placebo-controlled clinical trial aimed to assess the effect of oral antioxidant treatment (Fertilis), which contains L-carnitine and some micronutrients, in the improvement of conventional sperm parameters, sperm DNA integrity and in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) outcomes. A total of 263 participants were enrolled and randomly divided into two groups: 131 participants received the antioxidant treatment, while 132 participants received a placebo. The male partners in both groups underwent the antioxidant treatment or the placebo for a duration of three months. For each participant, we performed a hormonal test, an infectious test, a spermogram, a TUNEL assay for sperm DNA fragmentation, a toluidine blue staining for sperm DNA decondensation, and an IVF/ICSI procedure. Sperm characteristics analysis (volume, count, motility, and vitality), sperm DNA fragmentation, and sperm DNA decondensation were assessed and compared to the results preceding the antioxidant treatment. The study outcome revealed a significant decrease in the DNA fragmentation index and a significant increase in sperm motility after 3 months of treatment (p = 0.01 and p = 0.02, respectively). Additionally, a significant improvement in clinical pregnancy rate (p = 0.01) and life birth rate (p = 0.031) was observed. No significant changes were observed in conventional sperm parameters (volume, count, and vitality) or sperm DNA decondensation (SDI). Antioxidant therapy has a beneficial impact on achieving pregnancy, whether through spontaneous conception or assisted reproductive procedures (ART).

A comparative study of canine epididymal sperm collection techniques and cryopreservation

IntroductionAn optimized collection method and freezing protocol for preservation of epididymal spermatozoa remains a topic of interest to many scientists. The current study focused on the collection and preservation of canine epididymal spermatozoa. During the process of collection of canine epididymal spermatozoa, blood content can occur, which may affect sperm cryopreservation in a negative way. Here, we compared first two epididymal sperm collection techniques [epididymal mincing (EM) and single incision epididymal sperm aspiration (SESA)]; and next we tried to solve the issue of blood content using an erythrocyte lysis buffer (ELB).MethodsHence spermatozoa were collected after weighing the epididymides, either by EM or SESA, and sperm quality assessed prior to and post freezing (concentration, total sperm output (TSO), motility, viability and morphology). Next, new sperm samples were collected from eight epididymides by EM and subjected either to a standard freezing protocol or to an ELB treatment freezing protocol. Post-thaw sperm parameters (concentration, TSO, motility, viability and morphology), including intracellular reactive oxygen species (ROS) and lipid peroxidation were assessed. The correlation between the weight of the epididymis and the TSO was evaluated based on the collection technique, and differences in sperm parameters were detected both within different collection techniques and between different pre-freezing treatment protocols.ResultsThere was a very strong correlation between the weight of the epididymis and the TSO for the EM technique (p = 0.002, R2 = 0.6), along with an increased sperm motility with EM compared to SESA (median 80%, inter-quartile range (IQR) 88–65 and median 67.5%, IQR 72.5–52.5, respectively; (p = 0.002). Post-thaw samples subjected to ELB treatment freezing protocol had lower motility and higher intracellular ROS compared to the standard freezing protocol (motility: median 56.25%, IQR 60–48.75 and median 70%, IQR 72.5–63, respectively; p = 0.01; ROS: median 78.5%, IQR 81.25–75.5 and median 70%, IQR 70.5–68.75, respectively; (p = 0.04).DiscussionThe results indicated that EM is a better technique to harvest epididymal spermatozoa despite the presence of some blood content. Furthermore, the ELB treatment should not be implemented to remove those red blood cells prior to cryopreservation of epididymal spermatozoa in dogs.

Granulocyte-Macrophage Colony-Stimulating Factor Cytokine Addition After the Freeze-Thawing Process Improves Human Sperm Motility and Vitality in Asthenoteratozoospermia Patients.

The cryopreservation-thawing process of spermatozoa cells has negative impacts on their structure, function, and fertility parameters, which are known as cryoinjury. Asthenozoospermia patients are more susceptible to cryoinjury. Granulocyte-macrophage colony-stimulating factor (GM-CSF) increases sperm glucose uptake via the induction of glucose transporters, resulting in increased sperm motility. This study aimed to investigate the efficiency of GM-CSF supplementation of the cryopreservation media for semen samples of asthenoteratozoospermia patients. The study was carried out on 20 semen samples from infertile men referred to diagnosing semen analysis. To avoid subjective bias, two main sperm motility parameters, including velocity along the curvilinear path and velocity along the straight-line path were considered by the computer-assisted sperm analysis system. Afterward, each semen sample was divided into three equal aliquots and randomly assigned to one of the following groups: group I (control, freezing media only), group II (+GM-CSF, freezing medium supplemented with 2 μL/mL GM-CSF), or group III (GM-CSF added after thawing and washing). Following semen thawing, standard parameters, mitochondrial membrane potential (MMP), and the DNA Fragmentation Index were analyzed. Total sperm motility (progressive and non-progressive) improved significantly in group III samples after a 30-minute incubation with GM-CSF compared with the control group (26.5% ± 3.1% vs. 17.51% ± 2.59%). However, no differences in progressive motility or sperm morphology were found among the three thawed samples. The percentage of vitality was significantly higher in group III compared with the other two groups (28.38% ± 3.4% vs. 22.4% ± 3.08% and 22.14% ± 2.77%, respectively) (p < 0.05). JC-1 levels (a marker of MMP) were not significantly different between the examined groups (44.95% ± 8.26% vs. 36.61% ± 6.95% vs. 46.67% ± 7.7%, for control, group II, and group III, respectively) (p > 0.05). GM-CSF may be advantageous as an additive after freezing, improving total motility and viability after 30 minutes of post-thaw incubation; however, when supplied to the freezing media before cryopreservation, it is unable to protect against cryoinjury.

Grape Seed Proanthocyanidins Improve the Quality of Fresh and Cryopreserved Semen in Bulls.

Oxidative stress leads to a decrease in semen quality during semen cryopreservation and fresh semen production. Grape seed proanthocyanidins (GSPs) are endowed with well-recognized antioxidant, anti-inflammatory, anti-cancer, and anti-aging activities. Therefore, the objective of this experiment was to explore the effects of GSPs on the quality of fresh and cryopreserved semen to provide a basis for GSPs as a new dietary additive and semen diluent additive for males' reproduction. Fresh semen from three healthy bulls aged 3 to 5 years old were gathered and mixed with semen diluents dissolved with 0 µg/mL, 30 µg/mL, 40 µg/mL, 50 µg/mL, and 60 µg/mL GSPs respectively. The motility, physiological structures (acrosome integrity, membrane integrity, mitochondrial activity), and antioxidant capacity of frozen-thawed sperm were measured after storage in liquid nitrogen for 7 days (d). Bulls were fed with 20 mg/kg body weight (BW) GSPs in their diet for 60 days; the weight of the bull is about 600 kg. Then, the reproductive performance and antioxidant indexes of bulls were measured before and after feeding. The results demonstrated that GSPs supplementation significantly increased sperm motility, physiological structures, GSH-Px, and CAT enzyme activities and significantly decreased MDA content in sperm during semen cryopreservation. The optimal concentration of GSPs was 40 µg/mL (p < 0.05). After 20 mg/kg (body weight) GSP supplementation, sperm motility was significantly heightened (p < 0.05), the sperm deformity rate was significantly reduced (p < 0.05), and antioxidant enzyme activities (such as SOD, CAT, and GSH-Px) were significantly enhanced (p < 0.05), and the production of MDA was significantly suppressed (p < 0.05) in serum compared with that before feeding. In conclusion, these results reveal that a certain concentration of GSPs has a good protective effect on sperm damage caused by semen cryopreservation and the reproductive performance reduction caused by stress in bulls, which may be attributed to the antioxidant function of GSPs. In summary, GSPs are a useful cryoprotective adjuvant and dietary additive for bull sperm quality.

P-073 Comparison of human preimplantation embryo development after sperm selection for ICSI using the microfluidic ZyMōt device versus density gradient centrifugation

Abstract Study question Does microfluidic sperm selection using ZyMōt device has a significant effect on blastocyst development, ploidy, and improves pregnancy outcomes when compared with density-gradient centrifugation. Summary answer The study demonstrated that sperm selection using a microfluidic device had no impact on euploid blastocyst rates or clinical pregnancy outcomes. What is known already Selecting competent spermatozoa with the highest genomic integrity for ICSI is a key to achieving normal embryo development and a healthy live birth. Several sperm preparation techniques have long been scrutinized with varied success. The novel microfluidic ZyMōt device appears promising for improving the quality of sperm used in Assisted Reproductive Technologies. Preliminary tests have shown that ZyMōt may increase sperm motility and considerably decrease DNA fragmentation, which could potentially enhance euploid blastocyst development rates. However, the impact of this sperm selection method on clinical outcomes has not been fully investigated. Study design, size, duration This is a retrospective cohort study performed in a single academic IVF centre (IRB # 16367). Data were collected from 1286 ICSI cycles in 2020-2021 (730 with ZyMōt and 556 with DGC), including 867 cycles with PGT-A outcomes. Morphological grading of 6640 blastocysts cultured in a time lapse imaging incubator (Embryoscope™) and ploidy of 4783 blastocysts using high resolution NGS (Illumina platform and Blue Gnome) were compared between the two study groups. Participants/materials, setting, methods All male partners were normo-spermic according to standard semen analysis. ICSI results for a total of 7277 mature oocytes injected with spermatozoa processed with ZyMōt device and 5977 oocytes injected with spermatozoa processed by gradient-density centrifugation were compared. For the analysis of 1478 euploid embryo transfer outcomes, all cases included a single blastocyst transferred in a frozen-thaw cycle. Associated patient characteristics, including age and fertility diagnosis were collected. Main results and the role of chance A significantly higher fertilization rate was observed with ZyMōt compared to the gradient prepared sperm (83.4% vs 81.0%, P = 0.0004, respectively). The blastocyst rates (61.3%, vs 60.4%, P = 0.34) as well as the proportion of high-quality blastocysts were similar in both groups (P = 0.20). Euploidy rates were not different between all female age groups, including young egg donors (≤ 30y of age) to patients with advanced maternal age (&amp;gt;40 y of age). Microfluidic sperm selection did not improve clinical pregnancy rates after euploid embryo transfer (42.9% vs 41.4%; P = 0.58). Limitations, reasons for caution Although this is the largest study to date evaluating application of the ZyMōt device, the retrospective nature and cohort design limited the interpretation of our results. The patient population did not include male factor cases and sperm DFI data was not available. Embryo transfer outcomes were limited to clinical pregnancies. Wider implications of the findings The ZyMōt device offers a convenient alternative to standard sperm processing by density gradient method. Further studies are needed to fully determined the impact of ZyMōt on embryo quality and clinical outcomes in different age and diagnosis groups of patients; particularly for those with high DFI and poor IVF/ICSI outcomes. Trial registration number not applicable

P-074 In-vitro vitamin D treatment on human sperm improves sperm motility, capacitation, and oocyte activation

Abstract Study question Does in-vitro vitamin D treatment on fertile and infertile human sperm improves sperm motility and alter protein markers involved in capacitation and oocyte-activation. Summary answer In-vitro vitamin D treatment enhance functionality of sperm by increase intracellular calcium ion, thus increasing sperm motility and capacitation status. What is known already Many studies have been done to investigate the relationship between vitamin D and male infertility. It is known that low serum vitamin D in human contribute to male infertility. However, the effect of in-vitro vitamin D treatment on sperm remains unclear, particularly on the effect of vitamin D following ejaculation into female reproductive tract. This includes processes involved in fertilisation such as capacitation, hyperpolarization, oocyte activation and sperm-oocyte fusion. Therefore, we examined whether in-vitro vitamin D treatment on sperm can improve the sperm parameters, hence will then improve fertilisation and pregnancy. Study design, size, duration Randomised controlled trial with prospective interventions (in-vitro vitamin D treatment). Based on power analysis, 40 subjects both fertile and infertile men who came for semen analysis were recruited at University Malaya Medical Centre from January 2022 to January 2023. All men were consented to participate in this study. Each washed semen sample were aliquoted to serve as control and receive in-vitro vitamin D treatment. Participants/materials, setting, methods Total of 40 samples were included for final analysis. All semen samples were washed and treated with 1nM vitamin D in-vitro for 45 minutes at 37ºC. Motility, intracellular calcium released, capacitation status, mitochondria membrane potential and other basic semen parameters were measured after in-vitro vitamin D treatment for fertile and infertile samples. Furthermore, immunofluorescence of sperm origin oocyte activation factor, Phospholipase C Zeta (PLCζ), was done to investigate fluorescence intensity per cell using Image J. Main results and the role of chance In both fertile and infertile sample group, total motility is increased after in-vitro vitamin D treatment (MD = 8.727%, P &amp;lt; 0.001). Fluo-4 AM stain shows there are increase of intracellular calcium release after in-vitro vitamin D, with mean difference of Corrected Total Cell Fluorescent (CTCF) for Fluo-4 AM (MD = 25043, P = 0.0228. Chlortetracyclin (CTC) test for capacitation status and immunofluorescence of phosphotyrosine shows there are increase CTCF (MD = 0.02595, P = 0.0142) after in-vitro vitamin D treatment. Staining of JC-1 shows there are more hyperpolarised mitochondria membrane in in-vitro vitamin D treatment group compare to the control group. Ratio of red to green fluorescent is 0.639 in control group and 2.22 in in-vitro vitamin D treatment group. Moreover, immunofluorescence of sperm origin oocyte activation factor, Phospholipase C Zeta (PLCζ) show in-vitro vitamin D improve the expression of these protein. Mean difference of CTCF between control and vitamin D treatment group for PLCζ is 8.013x 10-3, P = 0.0226. Thus, in-vitro vitamin D enhance functionality of sperm by increase intracellular calcium ion, thus increasing sperm capacitation status and sperm membrane hyperpolarization. Limitations, reasons for caution Although power analysis is used to calculate sample size, however bigger sample size always needed to increase the accuracy. Moreover, patients’ sample are examined and treated after 3 hours due to limited collection room in the facility. Wider implications of the findings Vitamin D can be used as a tool in in-vitro fertilisation (IVF) procedure and can be added to sperm preparation medium due to its implication in increasing capacitation and hyperpolarised sperm membrane to increase chance of successful IVF. Trial registration number not applicable

Lactate as the sole energy substrate induces spontaneous acrosome reaction in viable stallion spermatozoa.

Equine spermatozoa appear to differ from spermatozoa of other species in using oxidative phosphorylation preferentially over glycolysis. However, there is little information regarding effects of different energy sources on measured parameters in equine spermatozoa. To determine the effect of three individual energy substrates, glucose, pyruvate, and lactate, on motion characteristics, membrane integrity, and acrosomal status of stallion spermatozoa. Freshly ejaculated stallion spermatozoa were incubated with combinations of glucose (5mm), pyruvate (10mm), and lactate (10mm) for 0.5 to 4h. Response to calcium ionophore A23187 (5μm) was used to evaluate capacitation status. Motility was evaluated using computer-assisted sperm analysis, and plasma membrane and acrosomal integrity were evaluated by flow cytometry. Incubation with lactate alone for 2h increased acrosomal sensitivity to A23187. Notably, incubation with lactate alone for 4h induced a significant spontaneous increase in acrosome-reacted, membrane-intact (viable) spermatozoa, to approximately 50% of the live population, whereas no increase was seen with incubation in glucose or pyruvate alone. This acrosomal effect was observed in spermatozoa incubated at physiological pH as well as under alkaline conditions (medium pH approximately 8.5). Sperm motility declined concomitantly with the increase in acrosome-reacted spermatozoa. Sperm motility was significantly higher in pyruvate-only medium than in glucose or lactate. The addition of pyruvate to lactate-containing medium increased sperm motility but reduced the proportion of live acrosome-reacted spermatozoa in a dose-dependent fashion. This is the first study to demonstrate that incubation with a specific energy substrate, lactate, is associated with spontaneous acrosome reaction in spermatozoa. The proportion of live, acrosome-reacted spermatozoa obtained is among the highest reported for equine spermatozoa. These findings highlight the delicate control of key sperm functions, and may serve as a basis to increase our understanding of stallion sperm physiology.

The quality of ram spermatozoa after thawing with the addition of Mn2+, Zn2+ and Cu2+ nanocitrate to cryopreservation diluent

The aim of the study was to find out the effect of adding nanocitrate of Mn, Zn and Cu to the diluent for ram spermatozoa cryopreservation on its quality and ability for fertilizing. The experiment was carried out on six clinically healthy breeder 2–4-year-old rams of the Texel breed. The received ejaculates of the rams were evaluated for the volume, sperm concentration and motility and then divided into control and experimental groups. Control sperm samples were diluted with lactose-yolk-tris-citrate-glycerin medium (LYTCGM). Nanocitrates of microelements were added to the medium in experimental samples of ram sperm in the following doses: Zn2+ and Mn2+ — 2.5, 5.0 and 7.5 μg/l, Cu2+ — 1.25, 2.5 and 3.75 μg/l. The diluted sperm was packaged in straws, equilibrated for 2.5 h and frozen. After thawing of sperm we determined motility, survival of sperm, activity of succinate dehydrogenase (SDH) and cytochrome oxidase (CO), activity of antioxidant protection enzymes superoxide dismutase (SOD), glutathione peroxidase (HPO) and catalase (CAT). A dose- dependent effect of Mn, Zn, and Cu nanocitrates upon their addition to LYTCGM was established. Addition of nanocitrates of Mn, Zn to LYTCGM at a dose of 5.0 μg/l increased sperm motility by 22.2% (P&lt;0.05) and 26.0% (P&lt;0.01), and sperm survival, respectively, by 12.6% on (P&lt;0.01) and 5.9% (P&lt;0.05) compared to the control. Nanocitrates of Mn, Zn at a dose of 5.0 μg/l as part of LYTCGM caused a probable increase in SDH (P&lt;0.001) and CO (P&lt;0.05–0.01), which indicates a high fertilizing ability of ram spermatozoa. Similarly, when Mn, Zn nanocitrates were added to LYTCGM at a dose of 5.0 μg/l, SOD activity decreased by 29.6% (P&lt;0.01) and 38.8% (P&lt;0.01) and HPO activity increased by 43.5% (P&lt;0.01) and 39.1% (P&lt;0.01), and CAT — by 40.0% (P&lt;0.05) and 37.5% (P&lt;0.05), respectively. At the same time, the addition of Cu nanocitrate to LYTCGM with an increase in the dose significantly reduces the activity, survival and fertilizing capacity of thawed ram spermatozoa, and also worsens their antioxidant protection.

Conditioned medium and secretome from epididymal epithelial cell cultures improve sperm kinetics and capacitation.

Sperm maturation occurs in the epididymis through interactions with existing molecules inside the lumen. However, the mechanism of epididymis molecular transfer is currently unclear. This study was aimed to determine the necessity of the epididymal epithelial cells (EECs) in the process of sperm maturation in terms of sperm kinetics and tyrosine phosphorylation. A true experimental research design was used in this study. The medium tested was a primary culture of mice caput epididymal cells (cells and culture medium), conditioned medium (CM) (supernatant of EECs), and secretome (CM filtered at 0.22 μm). Sperm was cocultured in EEC culture, CM, and secretome for 1, 2, 3, or 4 h. The original culture medium was used as the control. Sperm kinetic analysis was performed after the indicated times using computer-assisted sperm analysis, and tyrosine phosphorylation was detected using the Western blot technique. A primary culture of caput EECs was successfully generated. The results showed increased sperm motility and progressive movement after 3 h of incubation (p < 0.05). There was a significant decrease in the average path velocity (VAP) values after 4 h of incubation (p < 0.05), but there was no significant change in the 1, 2, and 3 h incubation groups. The EEC culture-CM and secretome groups showed a significant increased progressivity and VAP percentage values compared with the control medium (p < 0.05). In terms of percentage motility, the culture and CM groups were significantly different from the control medium, but the secretome group was not. The sperm kinetics of sperm cultured in CM, secretome, and EEC were significantly increased after 3 h of incubation, suggesting that CM and secretome can be used to replace EECs, especially when analyzing molecules secreted by the epididymal epithelium during sperm maturation. The results of this study highlight the potential of CM and secretome as therapy mediums for sperm kinetic abnormalities.

Understanding the impact of varicocele on sperm capacitation

To study the relationship between the seminal sample quality of men with varicocele and sperm capacitation. Cross-sectional observational study. Academic hospital. Seventy-six men (19 control and 57 with varicocele) were analyzed. Semen samples were submitted to a discontinuous density gradient for sperm selection. Sperm capacitation was induced using a human tubal fluid medium supplemented with bovine serum albumin. After capacitation induction, the sperm were assessed by capacitation state, computer-assisted sperm motility, mitochondrial activity, membrane integrity, acrosome reaction, and intracellular oxidative stress. The capacitation period increased sperm motility, showing an increase in the average path velocity and a decrease in the straightness compared with sperm before capacitation (paired analysis). After capacitation, the rate of capacitated sperm, motility, and mitochondrial activity showed differences between groups (control and varicocele). The varicocele group showed lower mitochondrial activity and capacitation than the control group. On the other hand, no significant differences were observed in the other variables evaluated. Varicocele men showed less viable sperm and mitochondrial activity than control men after capacitation sperm. The induction of capacitation altered motility by increasing path velocity and decreasing straightness in all of the studied groups, evidencing the occurrence of hyperactivation.

Effect of ovarian fluid on sperm performance in teleost with internal and external fertilization strategies

Ovarian fluid is essential for successful fertilization by maintaining the viability, motility, and velocity of sperm. The organic compounds and inorganic ions in ovarian fluid significantly influence spermatozoa's motility, velocity, and longevity. However, the effect of ovarian fluid on sperm performance is limited in teleost fish. In this study, the effect of ovarian fluid on sperm performance and its components in external fertilization species (Scophthalmus maximus, turbot) and internal fertilization species (Sebastes schlegelii, black rockfish) was investigated using computer-assisted sperm analysis, high-performance liquid chromatography, and metabolome analysis. The ovarian fluid had a distinct and species-specific effect on both species. In the black rockfish, the ovarian fluid from turbot significantly increased sperm motility (74.07% ± 4.09%), as well as VCL (45 ± 1.67 μm/s), VAP (40.17 ± 1.6 μm/s), and VSL (36.67 ± 1.86 μm/s), and longevity (352 ± 11.31 min) (P < 0.05). In the turbot, only the longevity (71.33 ± 5.69 min) and fertilization rate (65.27% ± 11.59%) showed significantly improvement (P < 0.05). The ovarian fluid was rich in organic compounds, suggesting enrichment in the glycolysis/gluconeogenesis pathways. The results suggest that glycometabolism plays a crucial role in improving sperm performance in teleost with internal fertilization. Thus, incorporating ovarian fluid into the sperm activation medium can enhance artificial fertilization in fish breeding.

Antifertility Effect of Polyaniline with Iron α-Fe2O3 on Epididymal Sperm of Mice

Chemical polymerization produces polyaniline in two concentrations (0.02,0.05g) at room temperature. The reaction of polymerization was exothermal in nature. The characterization peak was observed in the FTIR range between 1470 and 1560cm-1which defined benzenoid and quinoid ring. To find out the effects Fe2O3, this investigation was carried out with two weights of Fe2O3 (0.05 and 0.02 g). In the experiment part, 32 mice were separated into eight groups, as indicated. The experiment lasted for 35 days. The results showed a significant (P &lt; 0.05) decrease in sperm motility, concentration, percentage of live sperm and testosterone concentration with a significant (P &lt; 0.05) increase in the percentage of abnormal sperm in group treated with PANI-Fe2O3-Cys without magnetic field in comparison with control group. The results also showed a significant (P &lt; 0.05) increase in sperm motility, Relative sperm count of live sperm and testosterone concentration, with a significant (P &lt; 0.05) decrease in the percentage of abnormal sperm in group treated with PANI-Cys and PANI-Fe2O3-Cys exposed to magnetic field in comparison with the samples without magnetic field. The group treated with PANI alone showed an improvement in all parameters studied except the percentage of abnormal sperms which showed a significant (P &lt; 0.05) decrease. Using the alternating magnetic field, it was found that it has a obvious effect on the work of the material PANI- Fe2O3-Cys and PAN-Cys. Thus, it affects sperm motility

Antifreeze protein type III addition to freezing extender comprehensively improves post-thaw sperm properties in Okinawan native Agu pig

Sperm cryopreservation often leads to physical cell damage through ice crystal formation. This study evaluates the improvements to freezing extender cryoprotective activity due to antifreeze protein (AFP) addition, which primarily acts on ice crystal formation, through investigating the post-thaw sperm properties of Okinawan native Agu pig. Six individual boar sperm samples were diluted with the freezing extender supplemented with 1 μg/mL of AFP I or AFP III and then subjected to cryopreservation. Treatment with AFP I during the freezing procedure had no improvement for any characteristics after thawing compared to untreated sperm. In contrast, the addition of AFP III to the freezing extender strongly increased sperm motility, mitochondria and cell membrane integrity, and the acrosomal proteolytic activity of frozen-thawed sperm in 5 of 6 individuals (P < 0.05). Furthermore, cryoinjury prevention by AFP III significantly enhanced sperm viability (by ATP content), and maintained DNA quality and in vitro sperm penetrability compared with AFP I treatment (P < 0.05). These findings demonstrate that AFP III addition to the freezing extender of boar sperm is more effective in maintaining sperm characteristics than the extender without AFP III or AFP I, despite individual differences in response.