Semen Extender Research Articles

β-Nicotinamide mononucleotide improves chilled ram sperm quality in vitro by reducing oxidative stress damage.

The present study aimed to investigate the effect of β-nicotinamide mononucleotide (NMN) supplementation on ram sperm quality during storage at 4°C in vitro. Tris-citric acid-glucose solution containing different doses of NMN (0, 30, 60, 90, and 120 μM) was used to dilute semen collected from rams and it was stored at 4°C. Sperm motility, plasma membrane integrity as well as acrosome integrity were evaluated at 0, 24, and 48 h time points after storage at 4°C. In addition, sperm mitochondrial activity, lipid peroxidation (LPO), malondialdehyde (MDA) content, reactive oxygen species (ROS) content, glutathione (GSH) content, superoxide dismutase (SOD) activity, and apoptosis were measured at 48 h time point after storage at 4°C. Results demonstrate that the values obtained for sperm motility, acrosome integrity, and plasma membrane integrity in the NMN treatments were significantly higher than control (p<0.05). The addition of 60 μM NMN significantly improved ram sperm mitochondrial activity and reduced LPO, MDA content, and ROS content compared to control (p<0.05). Interestingly, sperm GSH content and SOD activity for the 60 μM NMN treatment were much higher than those observed for control. NMN treatment also decreased the level of Cleaved-Caspase 3, Cleaved-Caspase 9, and Bax while increasing Bcl-2 level in sperm at 48 h time point after storage at 4°C. Ram sperm quality can be maintained during storage at 4°C with the addition of NMN at 60 μM to the semen extender. NMN also reduces oxidative stress and apoptosis. Overall, these findings suggest that NMN is efficient in improving the viability of ram sperm during storage at 4°C in vitro.

The Effects on Post-Thaw Sperm Quality and Nuclear DNA Integrity of Supplementation of Low-Density Lipoprotein to Freezing Extender in the Mouse

Mice are an important research tool for genetic and molecular biology, allowing researchers to explore a variety of human illness models. Egg yolk is a common component of semen extenders for domestic animals and low-density lipoproteins (LDL) from egg yolk have some cryoprotective properties. This study aimed to investigate sperm quality characteristics and nuclear DNA integrity after post-thawing in an extender (18% raffinose + 3% skim milk) supplemented with different concentrations of LDL (2.5%, 5.0%, 7.5%, or 10%) in mice. 18% Raffinose+3% skim milk extender was used as a control group without LDL. CD-1 mice were used in the study, and semen was collected from the cauda epididymis and diluted with the extender. The straws were then frozen and thawed to evaluate progressive motility, viability, plasma membrane (HOST), acrosome, and nuclear DNA integrity parameters. Fresh sperm had the highest progressive motility, viability, plasma membrane integrity, and longevity (endurance) of progressive motility for 4 h in HTF solution. The greatest spermatologic results, including nuclear DNA integrity, were determined in fresh sperm (p

Preparation, characterization, and ex-vivo evaluation of curcumin-loaded niosomal nanoparticles on the equine sperm quality during cooled storage

Oxidative damage to sperm during cooled storage is a significant issue, and curcumin, with its antioxidant properties, could be a solution. However, its low bioavailability presents a challenge that this study aims to address. The primary objective of this study was to investigate the potential of curcumin-loaded niosomal nanoparticles (Cur-LNN) to enhance the antioxidant properties of curcumin and its effect on sperm quality during 72 h cooled storage. The thin-film hydration procedure was applied to prepare Cur-LNN. The fabricated noisomal nanocarriers were characterized using dynamic light scattering (DLS), transmission electron microscopy (TEM), field emission scanning electron microscopy (FESEM), and Fourier-transform infrared (FT-IR) spectroscopy. Moreover, the encapsulation and loading efficiency, in vitro release study, and ex-vivo antioxidant functionality of Cur-LNN on the stallion sperm preserved under cooled storage conditions were assessed. The fabricated Cur-LNN was spherical in shape and had an average particle size of 163.1 ± 1.8 nm, a zeta potential of −34.1 ± 1.9 mV, a poly-dispersity index of 0.339 ± 0.045, an encapsulation efficiency of 92.34 ± 0.18 %, and a loading efficiency of 35.57 ± 1.36 %. Ex-vivo evaluation revealed that supplementation of the semen extender with Cur-LNN has the potential to enhance sperm quality by improving total and progressive motility, plasma membrane functionality, and lipid peroxidation. These results demonstrate that Cur-LNN exhibited stronger antioxidant and protective effects than curcumin. Although further in vivo investigations are warranted, our ex-vivo results suggest that Cur-LNN has the potential to attenuate oxidative damage and can be used to fortify the antioxidant capacity of equine semen under cooled storage conditions.

Morphokinetic Profiling Suggests That Rapid First Cleavage Division Accurately Predicts the Chances of Blastulation in Pig In Vitro Produced Embryos.

The study of pig preimplantation embryo development has several potential uses: from agriculture to the production of medically relevant genetically modified organisms and from rare breed conservation to acting as a physiologically relevant model for progressing human and other (e.g., endangered) species' in vitro fertilisation technology. Despite this, barriers to the widespread adoption of pig embryo in vitro production include lipid-laden cells that are hard to visualise, slow adoption of contemporary technologies such as the use of time-lapse incubators or artificial intelligence, poor blastulation and high polyspermy rates. Here, we employ a commercially available time-lapse incubator to provide a comprehensive overview of the morphokinetics of pig preimplantation development for the first time. We tested the hypotheses that (a) there are differences in developmental timings between blastulating and non-blastulating embryos and (b) embryo developmental morphokinetic features can be used to predict the likelihood of blastulation. The abattoir-derived oocytes fertilised by commercial extended semen produced presumptive zygotes were split into two groups: cavitating/blastulating 144 h post gamete co-incubation and those that were not. The blastulating group reached the 2-cell and morula stages significantly earlier, and the time taken to reach the 2-cell stage was identified to be a predictive marker for blastocyst formation. Reverse cleavage was also associated with poor blastulation. These data demonstrate the potential of morphokinetic analysis in automating and upscaling pig in vitro production through effective embryo selection.

Effect of asiatic acid supplementation in tris-extender on post-thaw functional competence, antioxidant enzyme activity and in vivo fertility of bull sperm.

Reactive oxygen species at supra-physiological levels trigger oxidative stress during cryopreservation, which can be neutralised by incorporating suitable antioxidants into the semen extender medium. This study was intended to explore the effect of asiatic acid (AA) as an antioxidant in semen extender on frozen-thawed sperm quality and in vivo fertility of bull sperm. Semen was collected from Holstein Friesian bulls for 10 consecutive weeks (total ejaculates = 60). Semen was cryopreserved with a Tris citric acid egg yolk-based extender supplemented with 0 (control), 20, 40, 60, and 100 μM AA. The supplementation of the extender with 40 and 60 μM AA improved (p < 0.05) post-thaw motility kinematics, plasma membrane integrity, acrosome integrity, sperm viability, and DNA integrity of bull sperm. Mitochondrial membrane potential was high (p < 0.05) with 60 μM of AA concentration in extender media. The catalase activity in seminal plasma was maintained (p < 0.05) when semen was added with 20, 40, and 60 μM of AA. The in vivo fertility was found to be significantly high with the semen extended with 60 μM AA. Conclusively, this study showed that AA supplementation in semen extender significantly improved sperm motility kinematics and cell integrity, conserved antioxidant enzyme activity, and improved in vivo fertility.

Some Semen Characteristics and Oxidant/Antioxidant Markers of Chilled Diluted Rabbit Semen in Tris-Based Extender Supplemented with Wheat Germ Extracts

Some Semen Characteristics and Oxidant/Antioxidant Markers of Chilled Diluted Rabbit Semen in Tris-Based Extender Supplemented with Wheat Germ Extracts

Synthesis of green zinc-oxide nanoparticles and its dose-dependent beneficial effect on spermatozoa during preservation: sperm functional integrity, fertility and antimicrobial activity.

Introduction: The development of an effective extender is important for semen preservation and the artificial insemination (AI) industry. This study demonstrates the beneficial effect of zinc oxide nanoparticles (ZnO-NPs) as an additive to semen extenders to improve semen quality, fertility, and antibacterial activity during liquid preservation in a boar model. Methods: Initially, to find out the safe concentration of ZnO-NPs in sperm cells, a wide range of ZnO-NP concentrations (0, 5, 10, 50, 100, 500, and 1,000μM) were co-incubated with sperm at 37°C for a cytotoxic study. These NP concentrations were compared to their salt control zinc acetate (ZA) at the same concentrations and to a control group. The effect of the different concentrations of ZnO-NPs on sperm motility, membrane integrity, mitochondrial membrane potential (MMP), and apoptosis was assessed. Accordingly, the non-toxic dose was selected and supplemented in MODENA extender to determine its beneficial effect on the boar semen parameters mentioned and the lipid peroxidation (LPO) levels during liquid preservation at 16°C for 6days. The non-cytotoxic dosage was subsequently chosen for AI, fertility investigations, and the evaluation of the antibacterial efficacy of ZnO-NPs during preservation hours. An antibacterial study of ZnO-NPs and its salt control at doses of 10μM and 50μM was carried out by the colony forming unit (CFU) method. Results and discussion: The cytotoxic study revealed that 5, 10, and 50μM of ZnO-NPs are safe. Consequently, semen preserved in the MODENA extender, incorporating the non-toxic dose, exhibited 10 and 50μM ZnO-NPs as the optimal concentrations for beneficial outcomes during liquid preservation at 16°C. ZnO-NPs of 10μM concentration resulted in a significantly (p < 0.05) improved conception rate of 86.95% compared to the control of 73.13%. ZnO-NPs of 10 and 50μM concentrations exhibit potent antimicrobial action by reducing the number of colonies formed with days of preservation in comparison to the negative control. The investigation concluded that the incorporation of 10μM ZnO-NPs led to enhancements in sperm motility, membrane integrity, and MMP, attributed to a reduction in the malondialdehyde (MDA) levels. This improvement was accompanied by a concurrent increase in fertility rates, including farrowing rate and litter size, during the liquid preservation process. Furthermore, ZnO-NPs exhibited an antimicrobial effect, resulting in decreased bacterial growth while preserving boar semen at 16°C for 6days. These findings suggest that ZnO-NPs could serve as a viable alternative to antibiotics, potentially mitigating antibiotic resistance concerns within the food chain.

In-Vivo Effects of Canary (Serinus Canary) Semen Diluted with Different Semen Extenders on Eggs in Artificial Insemination

In this study, the effect of Dulbecco's Phosphate Buffered Saline (DPBS) and Dulbecco's Modified Eagle's Medium (DMEM) extenders on artificial insemination was determined.. For this purpose, the number of fertile eggs, the total number of eggs, the number of hatched eggs, and the number of hatchlings from fertile eggs were determined in the study. From these values, the fertility rate (%), egg hatch rate (%) and fertile egg hatch rate (%) were calculated. Twelve male and female canaries (Gloster breed; 1-2 years old) were used in each group. In the study, three groups, one control (C) and the other two treatment groups (T-1, T-2) were formed. Female canaries in group C were directly inseminated without using any extenders. Females in the T-1 group were diluted 1:1 with DPBS; females in the T-2 group were inseminated with semen diluted 1:1 with DMEM. Although there was a significant difference between the groups in terms of fertility rate (P0.05). The highest fertility rate (61.81%) was in group C; the lowest was in the T-1 (29.17%) group. (P

Freezing Stallion Semen-What Do We Need to Focus on for the Future?

Artificial insemination (AI) is used frequently in the breeding of sport horses, apart from Thoroughbreds. Most AIs are carried out with cooled semen rather than frozen semen because of the difficulties in identifying a protocol that is suitable for freezing most ejaculates and the necessity to inseminate close to ovulation because of the short life of the thawed spermatozoa. More widespread use of frozen semen would improve biosecurity, allow greater choice of stallions, and offer more flexibility when managing deliveries of semen to the stud. It would even decrease the amount of antibiotics used in semen extenders, since the volume of frozen semen is smaller than when cooled semen is inseminated. However, there is considerable variability in the cryosurvival of spermatozoa from different stallions, leading to the classification of stallions as good or bad freezers. Improvements could be made at the level of stallion nutrition, the semen collection regimen, the extender, the removal of seminal plasma, and the cooling protocol, among others. Stallion sperm membranes are highly susceptible to lipid peroxidation, but research on antioxidants has failed to identify an additive that would benefit all stallions. In the future, biomarkers for sperm freezability could be used as an aid in identifying suitable ejaculates for cryopreservation.

Bacterial diversity in semen from stallions in three European countries evaluated by 16S sequencing

The microbiome plays a significant role in shaping the health and functioning of the systems it inhabits. The seminal microbiome of stallions has implications for the health of the reproductive tract, sperm quality during preservation and antibiotic use in semen extenders. Diverse bacteria are present on the external genital tract and a mix of commensal microorganisms populates various parts of the reproductive tract, influencing the seminal bacterial content. Other sources of bacteria include the environment, semen collection equipment, and personnel. The bacterial load can adversely affect sperm quality and fertility, particularly in artificial insemination, where semen is extended and stored before use. Antibiotics are frequently used to inhibit bacterial growth, but their effectiveness varies depending on the bacterial strains present. The aim of this study was to assess the bacterial diversity in semen from 37 healthy stallions across three European nations (Germany, Portugal, and Sweden) using 16S sequencing. Semen samples were collected from individual stallions at three AI centers; DNA extraction, sequencing, and bioinformatic analysis were performed. Differences in bacterial diversity among the stallions were seen; although bacterial phyla were shared across the regions, differences were observed at the genus level. Climate, husbandry practices, and individual variability likely contribute to these differences. These findings underscore the importance of tailoring antibiotic strategies for semen preservation based on regional bacterial profiles. The study presents a comprehensive approach to understanding the intricacies of the stallion seminal microbiome and its potential implications for reproductive technologies and animal health.

Effect of incorrect storage of bull semen samples on sperm morphology assessment

Thirteen pairs of bull semen samples were assessed. One of each pair of samples was inadvertently stored in phosphate buffered saline (PBS) that was used for analysis of motility immediately after collection. Other sample of the pair that was correctly stored in buffered formol saline (BFS) was assessed later. Proportion of morphologically normal sperm was different (p = 0.0021) between the 2 storage methods. Most of the abnormalities in the PBS samples were loose and detached heads. Closer examination of these sperm illustrated changes in the composition of the tail, with an apparent loss of the plasma membrane. This serendipitous error allowed documentation of the importance of correct storage of semen samples for morphological assessment of sperm, and the tertiary defects detected with the incorrect storage of semen samples in semen extender. Tertiary defects should always be considered a possibility during morphological assessment of sperm via spermiogram.

Effect of reduced glutathione on quality, lipid peroxidation and antioxidant potential of frozen‐thawed ring‐necked pheasant semen

AbstractReduced glutathione (GSH) is a natural redox force immediately consumed by cells to combat toxins. Today's world is shifting towards clean farming practices whence ring‐necked pheasants dependent on croplands are facing threats and sperm‐assisted reproductive techniques become indispensable in preserving this valuable species for game and meat purposes. This study aims at evaluating the cryosurvivability (motility, viability, plasma membrane and acrosomal integrity, DNA fragmentation) and antioxidant capacity (Free radical scavenging activity, LPO detection, Ferric reducing antioxidant power, mitochondrial status) of ring‐necked pheasant semen with GSH supplementation in the extender. Semen collected from ten males was pooled, divided and diluted with Beltsville poultry semen extender (1:5) containing 0.0, 0.2, 0.4, 0.6 and 0.8 mM GSH. Samples were cryopreserved, and sperm quality was evaluated post‐dilution, cooling, equilibration, and freeze–thaw. Free radical scavenging activity, total antioxidant potential, mitochondrial action and lipid peroxidation were assessed at post‐thawing. Sperm motility, plasma membrane and acrosomal integrity, viability and DNA integrity were recorded highest (p &lt; .05) at 0.4 mM GSH compared to other concentrations and control at all the stages of cryopreservation. Similarly, sperm mitochondrial action, antioxidant potential and free radical scavenging capacity were recorded higher (p &lt; .05) with 0.4 mM GSH at post‐thaw compared to other concentrations of GSH and control. Lipid peroxidation was recorded as highest (p &lt; .05) in the control compared to GSH‐supplemented groups at post‐thaw. Hence, it is concluded that GSH at 0.4 mM preserves the quality and ROS scavenging capacity of cryopreserved ring‐necked pheasant sperm as an antioxidant.

Astaxanthin Improved the Quality of Hu Ram Semen by Increasing the Antioxidant Capacity and Mitochondrial Potential and Mitigating Free Radicals-Induced Oxidative Damage.

The objective of this research was to investigate the effect of astaxanthin supplementations of semen extender on the quality of Hu ram semen after up to five days of preservation at 4 °C. Semen samples were collected from five healthy Hu rams using an artificial vagina during breeding season (April to August 2023) and diluted with a basic extender supplemented with control (0), 1 µM, 2 µM, 3.5 µM, or 4.5 µM of AXT. Overall, 170 semen ejaculate samples (34 repetitions) from five healthy Hu rams were used in our research study. The results revealed that the addition of AXT (3.5 µM) significantly (p ≤ 0.05) increased the sperm kinematic indexes (T.M%, P.M%, MAD%, STR%, and LIN %), sperm viability, plasma membrane integrity, acrosome integrity, total antioxidant content (T-AOC), and mitochondrial membrane potential (MMP) of the Hu rams spermatozoa after up to five days of preservation at 4 °C. Contrary to that, the addition of the best concentration of AXT (3.5 µM) to the semen extender significantly (p ≤ 0.05) reduced the reactive oxygen species (ROS) and malondialdehyde (MDA) concentration of Hu ram semen. In conclusion, the results of the current study indicate that the addition of a semen extender with AXT improves the quality of Hu ram spermatozoa by increasing the total antioxidant capacity (T-AOC) and mitochondrial membrane potential (MMP). On the other hand, reducing free radicals induced oxidative (ROS) and per oxidative (MDA) damage to Hu ram semen.

Strategies to Reduce the Use of Antibiotics in Fresh and Chilled Equine Semen.

The study assessed the impact of four equine semen processing techniques on sperm quality and microbial load immediately post-processing and after 48 h of refrigeration. The aim was to explore the potential reduction of prophylactic antibiotic usage in semen extenders. Semen from ten adult stallions was collected and processed under a strict hygiene protocol and divided into four aliquots: Simple Centrifugation with antibiotics (SC+), Simple Centrifugation (SC-), Single-Layer Colloidal Centrifugation (CC-), and Filtration (with SpermFilter®) (F-), all in extenders without antibiotics. Sperm motility, viability, and microbial load on three culture media were assessed. No significant differences were observed in the main in the sperm quality parameters among the four protocols post-processing and at 48 h (p < 0.05 or p < 0.1). Microbial loads in Columbia 5% Sheep Blood Agar and Schaedler vitamin K1 5% Sheep Blood Agar mediums were significantly higher (p < 0.10) for raw semen than for CS+, CC-, and F- post-processing. For Sabouraud Dextrose Agar medium, the microbial load was significantly higher (p < 0.10) in raw semen compared to CS+ and F-. No significant differences (p < 0.10) were found in 48 h chilled samples. Regardless of antibiotic presence, the evaluated processing methods, when combined with rigorous hygiene measures, maintained semen quality and reduced microbial load to the same extent as a traditional protocol using antibiotics.

Exploring Myrciaria dubia liquid extract as a potential semen extender for breeding roosters.

The current investigation aimed to explore the effects of Myrciaria dubia liquid extract (MDLE) as the primary component of an extender for breeder rooster semen over different periods at room temperature. Fifteen breeder roosters (40 weeks of age, average body weight of 2.05±0.12) with confirmed fertility were used. Employing a factorial design (3x4), the treatments consisted of semen in natura and two semen extenders (an experimental based on MDLE and a commercial) subjected to four periods at room temperature post-collection (5, 10, 15 and 20 minutes) with four replicates (tubes) each. All variables evaluated in this study yielding significant results (p<0.05). Analyzed individually, the experimental extender based on MDLE exhibited a linear reduction (p<0.05) in motility and vigor results, while it caused an increase in pH values and percentages of sperm defects evaluated. When compared with semen in natura and commercial extender, the efficiency of MDLE as a semen extender was inferior to that observed with the commercial extender and similar to the results observed with semen in natura. Nonetheless, the experimental extender based on MDLE yielded satisfactory results for up to 15 minutes of storage time. In conclusion, MDLE can be considered as an alternative for composing a roosters' semen extender, maintaining sperm characteristics within acceptable limits for up to 15 minutes at room temperature. However, this experimental extender demonstrated lower efficiency than the commercial extender in maintaining the sperm quality at room temperature across all periods tested.

Assessing the efficacy of different semen extenders on post-thaw quality of Bangladeshi buffalo sperm under a manual cryopreservation protocol

Cryopreservation has been used extensively for cattle in Bangladesh, albeit no study was conducted on the cryopreservation of buffalo sperm in Bangladesh. This study aimed to evaluate the suitability of different semen extenders for improving the post-thaw quality of Bangladeshi buffalo sperm under a manual cryopreservation protocol. The manual cryopreservation protocol was compared and optimized with a commercial biofreezer protocol. Then, the efficacy of different diluters was evaluated using the optimized manual cryopreservation protocol. Meanwhile, the post-thaw sperm quality in terms of motility and morphology was evaluated by CASA during the optimization process. During manual cryopreservation, the first cooling from 37 °C to 5 °C was done in a BLRI-developed equilibration chamber and the second cooling from 5 °C to -120 °C in a Styrofoam box using liquid nitrogen vapor from different distances (0.5, 1.5, 1.6, 2 and 3 inches). Simultaneously, another batch of sperm was cryopreserved using a programmable freezer. The highest number of motile sperms (62.67±1.12; P&lt;0.01) and progressive motility (38.97±1.10; P&lt; 0.001) was observed at 1.6 inches above liquid nitrogen, which are similar to the results obtained from automated biofreezer protocol (65.94±4.65 and 45.54 ± 3.64, respectively). To evaluate the semen extenders’ efficacy, one locally developed Tris-fructose-egg yolk-based diluter and two commercial diluters (Andromed, Triladyl) were used in the freezing of buffalo sperm. The highest recovery and conception rates were observed in sperm diluted with TFE (82.4% and 80%, respectively). Therefore, it is clear that this manual cryopreservation protocol and the TFE diluter could be a suitable and inexpensive alternative for Bangladeshi buffalo sperm considering post-thaw sperm quality and fertility.

AN OVERVIEW OF SEMEN EXTENDERS AND FUTURE CHALLENGES TO DEVELOPMENT OF ARTIFICIAL INSEMINATION IN CAMEL

Artificial insemination is an emerging technique to ensure productivity and rapid genetic improvement in camels. This technique is not well developed due to involving different challenging factors because of semen collection, viscosity, nature of ejaculates, dilution in stabilized semen extenders, processing and cryopreservation, challenging related to artificial insemination and low conception rate. Numerous bovine commercial and non-commercial extenders with slight modification have been utilized for camel semen cryopreservation with little success. The gelatinous nature of the camel semen is one of the major challenges for proper evaluation, dosage calculation and cryopreservation. Various techniques have been used to liquefy the semen within a short spin of time without compromising the quality of sperm cells. This review provides precise data that appears in the peer reviews journals collected most of information to highlight the different semen extenders, addition of different supplementation and essential additives in semen extenders, challenges factors regarding camel bull and camel cow involved in success of artificial insemination.

Exploring enzymatic antioxidant activities in Asian Elephant (Elephas maximus) sperm and seminal plasma

This study explored the relationships between sperm quality and the activities of antioxidant enzymes (superoxide dismutase, SOD, and glutathione peroxidase, GPx) in sperm (S) and seminal plasma (SP) of Asian elephants, as well as the potential protective role of glutamine in semen extenders during chilled storage. In experiment 1, twenty ejaculates from 11 bulls were analyzed for percentages (%) of sperm motility (%MOT), viability (%VIA), and activities of SOD and GPx. A significant negative correlation was found between %VIA and SP-SOD (r = − 0.71), and positive correlations between %MOT and both S-GPx (r = 0.49) and SP-GPx (r = 0.57). Following these findings, glutamine, a precursor of glutathione and semen antioxidant was introduced in Experiment 2. Each ejaculate (in a total of 8 ejaculates obtained from four bulls) was diluted in Tris-fructose- (EYT) and Tris-glucose-egg yolk extender (TCG), respectively, supplemented with 0 (control), 20 and 40 mM glutamine and stored at 4 °C. Sperm parameters including %MOT, %VIA, functional membrane integrity (%sHOST), and DNA integrity (%DNA) were assessed after 24 and 48 h of storage. Results demonstrated that glutamine did not significantly affect %VIA or %DNA. A significant reduction in %MOT was observed at 24 h, but 20 mM glutamine added in extenders reversed this decline at 48 h. A 40 mM addition of glutamine in TCG markedly reduced %sHOST at 24 h. In conclusion, SOD and GPx activities in the sperm and seminal plasma are indicative of sperm quality. During chilling, using a higher level of glutamine may damage the spermatozoa of Asian elephants.

Antioxidant Glutathione Supplementation in Semen Extenders and its Effect on the Quality of Cryopreserved Bull Sperm: A Meta-Analysis

Antioxidant Glutathione Supplementation in Semen Extenders and its Effect on the Quality of Cryopreserved Bull Sperm: A Meta-Analysis

Sperm hyperactivation in the uterus and oviduct: a double-edged sword for sperm and maternal innate immunity toward fertility.

In cattle, artificial insemination (AI) is a technique that allows breeding by depositing frozen-thawed and extended semen into the female reproductive tract. The semen contains sperm with various motility patterns including dead, progressive and hyperactivated. Sperm hyperactivation is high amplitude, asymmetrical beating of sperm tail which usually occurs in the oviduct as part of the capacitation process, but it can also be induced by cryopreservation. After insemination, sperm enter the uterine glands and trigger a pro-inflammatory response in the uterus. Hyperactivated sperm, stimulated by sperm-Toll-like receptor 2 (TLR2), penetrates the mucus and uterine glands more efficiently and enhances the immune response. This facilitates the clearance of excess and dead sperm from the uterus. Some sperm escape the immune response and reach the oviduct either before or after the immune response is initiated. In the oviduct, sperm bind to the epithelium and form a reservoir. This triggers an anti-inflammatory response and preserves the fertilization potential of sperm. Hyperactivation facilitates sperm detaching from the epithelium, swimming through the viscous mucus and cumulus cells, and penetrating the egg's zona pellucida. Sperm-TLR2 activation enhances Ca2+-influx and acrosome reaction, which enables sperm to penetrate and fertilize oocytes during in vitro fertilization. Altogether, post-AI in cattle, sperm and maternal immunity interact differentially depending upon the site of sperm hyperactivation - whether it occurs within the uterus or oviduct. Specifically, hyperactivated sperm that enter the uterus after AI or are triggered via sperm-TLR2 activation or other stimuli contribute to sperm-induced uterine inflammation. Such hyperactivated sperm may impede their capacity to ascend to the oviduct. Conversely, sperm that become hyperactivated within the oviduct modulate their interactions with the oviduct and oocytes, which is pivotal during fertilization process. Indeed, the location and timing of sperm hyperactivation partially via TLR2 activation are critical determinants of their different influence on fertility.