Sperm Survival Rate Research Articles

α/β-Hydrolase D16B Truncation Results in Premature Sperm Capacitation in Cattle.

Recently it was shown that a specific form of male infertility in Holstein cattle was caused by a nonsense variant in the α/β-hydrolase domain-containing 16B (ABHD16B) gene resulting in a protein truncation at amino acid position 218 (p.218Q*) and loss of function. Lipidomics showed that the absence of ABHD16B influenced the content of phosphatidylcholine (PC), ceramide (Cer), diacylglycerol (DAG), and sphingomyelin (SM) in variant carrier sperm membranes. However, the exact cause of infertility in affected sires has remained unclear until now. To elucidate the cause of infertility, we analyzed (i) standard sperm parameters (i.e., total sperm number, morphological intact sperm, total sperm motility), (ii) in vitro fertilizability and effects on early embryonic development, and (iii) sperm survival rates (i.e., capacitation time). The affected spermatozoa showed no changes in the usual sperm parameters and were also capable of fertilization in vitro. Furthermore, the absence of ABHD16B did not affect early embryonic development. Based on these results, it was concluded that the affected spermatozoa appeared to be fertilizable per se. Consequently, the actual cause of the inability to fertilize could only be due to a time- and/or place-dependent process after artificial insemination and before fertilization. A process fundamental to the ability to fertilize after insemination is capacitation. Capacitation is a biochemical maturation process that spermatozoa undergo in the female genital tract and is inevitable for the successful fertilization of the oocyte. It is known that the presence and concentration of certain sperm membrane lipids are essential for the correct course of capacitation. However, precisely these lipids are absent in the membrane of spermatozoa affected by the ABHD16B truncation. Since all other causes of fertilization inability were excluded in the previous experiments, consequently, the only remaining hypothesis was that the loss of function of ABHD16B leads to a capacitation disruption. We were able to show that heterozygous and homozygous affected spermatozoa exhibit premature capacitation and therefore decay before fertilization. This effect of the loss of function of ABHD16B has not been described before and our studies now revealed why sires harboring the variant in the ABHD16B gene are infertile.

RhoA improves cryopreservation of rooster sperm through the Rho/RhoA-associated kinase/cofilin pathway

Cryopreservation of rooster sperm leads to relatively low semen quality due to cytoskeletal damage during the freeze-thawing process. This study aimed to explore how the addition of RhoA recombinant protein affected the viability and subcellular structure of rooster sperm after freeze-thawing and elucidated the molecular mechanisms of sperm cryopreservation. Semen quality and acrosome integrity testing revealed that the addition of 0.5 μg/mL RhoA recombinant protein to the cryoprotectant fluid significantly increased sperm motility, survival rate, linearity, straight-line velocity, and acrosome integrity after freeze-thawing (P < 0.05). Ultrastructure analysis of cryopreserved sperm showed structural damage to the sperm plasma membrane, nuclear membrane, and tail. However, compared to the control, these structural changes were reduced upon the addition of RhoA recombinant protein to the cryoprotective fluid (P < 0.05). Western blotting revealed that the expression of Rho/RhoA-associated kinase and p-cofilin was increased, and cofilin expression was decreased after sperm cryopreservation with recombinant RhoA protein. Treatment with Y-27632, a ROCK antagonist, suppressed ROCK and p-cofilin expression and decreased semen quality, acrosome integrity, and ultrastructure integrity. In summary, we have demonstrated a cryoprotective effect in spermatozoa involving the Rho/ROCK pathway during freeze-thawing. Furthermore, the addition of 0.5 μg/mL RhoA recombinant protein to the cryoprotective fluid improved rooster semen quality and subcellular structural homeostasis after freeze-thawing via the Rho/ROCK pathway. This pathway may regulate the dynamic reorganization of the actin cytoskeleton by regulating the cofilin phosphorylation.

A Sperm Quality Detection System Based on Microfluidic Chip and Micro-Imaging System

Sperm quality assessment is the main method to predict the reproductive ability of livestock. The detection of sperm quality of livestock is of great significance to the application of artificial insemination and in vitro fertilization. In order to comprehensively evaluate sperm quality and improve the real-time and portability of sperm quality detection, a portable microscopic imaging system based on microfluidic chip is developed in this paper. The system can realize the comprehensive evaluation of sperm quality by detecting sperm vitality and survival rate. On the hardware side, a microfluidic chip is designed, which can automatically mix samples. A set of optical system with a magnification of 400 times was developed for microscopic observation of sperm. In the aspect of software, aiming at the comprehensive evaluation of sperm quality based on OpenCV, a set of algorithms for identifying sperm motility and survival rate is proposed. The accuracy of the system in detecting sperm survival rate is 94.0%, and the error rate is 0.6%. The evaluation results of sperm motility are consistent with those of computer-aided sperm analysis (CASA). The system's identification time is 9 s. Therefore, the system is absolutely suitable for sperm quality detection.

EFFECT OF CONCENTRATION OF PHOSPHOLIPIDS&#x0D; AND HUMIC COMPOUNDS IN DILUENT COMPOSITION ON BIOLOGICAL PARAMETERS OF BULL AND HUMAN SPERM

In the course of the experimental work, it was found that the concentration of phospholipids and an aqueous solution of activated potassium humic substances affects the viability of bovine and human sperm.&#x0D; Phospholipids included in the composition of the diluent up to 10.0 and 20.0 mg, after 5 hours of incubation at a temperature of (37±1) °C, provide the survival rate of bull and human sperm at 34.4±0.86–35.5±0.86 % and 36.2±0.86–32.4±0.86 % germ cells, respectively.&#x0D; Diluents containing 0.0025 % aqueous solution of activated potassium humate and 10.0 and 20.0 mg of phospholipids provide the highest survival rate relative to the baseline in bovine and human spermatozoa, respectively, up to 72.9–78.8% and 73. 4–76.2 %.

Inhibition of human sperm function by an antibody against apolipoprotein A1: A protein located in human spermatozoa

Apolipoproteins A1 (ApoA1), an important protein in initiating sperm capacitation, has been found in the oviduct liquid, whilst whether it expresses in human spermatozoa has not yet been defined. ApoA1 expression was detected by reverse transcription-polymerase chain reaction, Western blot and immunofluorescence in sperm cells. Sperms were incubated with different concentration of ApoA1 antibody to observe the changes of sperm motility and apoptosis including annexin V binding to the cell surface, mitochondrial membrane potential and ultrastructural changes. In addition, cholesterol levels of human spermatozoa and fertilization in vitro were completed to explore the role of ApoA1 in fertilization process. We identify ApoA1mRNA is presented in spermatozoa, and the ApoA1 protein, the molecular weights of 31kD, is located at the postacrosomal region and neck of human spermatozoa. ApoA1 antibody affects cholesterol efflux of human spermatozoa and inhibits fertilization in vitro. Moreover, ApoA1 antibody decreases sperm motility and increases sperm apoptosis. This work shows ApoA1 plays an important role in maintaining cholesterol stability and sperm motility, survival rate and fertilization process. Furthermore, ApoA1 implications in the clinical work and other reproductive areas are needed to be studied.

Characteristics of diluted-stored and post-thawed semen of Hutsul stallions.

The use of frozen semen lowers the risk of disease transmission, eliminates geographical limitations and supports the implementation of genetic resource protection programs. However, due to the very rare use of frozen semen from Hutsul stallions, their genetic material is not secured in sperm banks, and very little information is available about their semen, including its suitability for cryopreservation, and sperm survival rates after thawing. The aim of this study was to analyse basic parameters such as sperm motility, vitality and morphology in diluted-stored and post-thawed Hutsul semen, using a CASA system. There were no differences in sperm motility (P = 0.3372) or morphology between the groups, although the progressive motility was higher in thawed semen (P = 0.0151), while the sperm vitality was higher in diluted-stored semen (P = 0.00517). This study demonstrates that semen from Hutsul horses is suitable for cryopreservation, thus supporting the creation of a sperm bank as a genetic reserve for representatives of this breed.

Sperm and testicular dysfunction during cecal ligation and puncture-induced sepsis in male rats and effects of tannic acid through reducing testicular oxidative stress and inflammation.

Objective(s):One of the problems caused by infectious diseases is the decrease in sperm count and motility. Tannic acid is known as an anti-oxidant and anti-inflammatory agent. In this study, Cecal Ligation and Puncture (CLP) sepsis model was induced to investigate the effect of tannic acid on oxidative stress and inflammation in testicular and sperm structure and function.Materials and Methods:Twenty-four male Wistar rats (250–300 g) were randomly divided into 3 groups of 8: 1) sham, 2) sepsis, and 3) sepsis + tannic acid (20 mg/kg at 6, 12, and 24 hr after sepsis induction). Thirty hours after induction of sepsis, testicular samples were collected to measure SOD activity and MDA, IL-6, and TNF-α levels. Another part of the testis was fixed in 10% formalin for histological examinations. Results:In the sepsis group, testicular MDA, TNF-α, and IL-6 levels increased and SOD activity decreased compared with the sham group. In addition, the percentage of motile sperm and the survival rate of sperm decreased significantly in the sepsis group. Administration of tannic acid significantly decreased inflammatory markers (TNF-α and IL-6) and MDA levels and increased SOD activity. Furthermore tannic acid significantly improved sperm parameters and increased sperm and animal survival rates.Conclusion:The results of this study showed that the reproductive system may be strongly affected by the conditions created during sepsis. Tannic acid improved reproductive dysfunction in sepsis by reducing oxidative stress and inflammation.

The Abscopal Effects of Cranial Irradiation Induce Testicular Damage in Mice.

To investigate whether the abscopal effects of cranial irradiation (C-irradiation) cause testicular damage in mice, male C57BL/6 mice (9weeks of age) were randomly divided into a sham irradiation group, a shielded group and a C-irradiation group and administered sham/shielded irradiation or C-irradiation at a dose rate of 2.33Gy/min (5Gy/d for 4 d consecutively). All mice were sacrificed at 4weeks after C-irradiation. We calculated the testis index, observed testicular histology by haematoxylin-eosin (HE) staining and observed testicular ultrastructure by transmission electron microscopy. Western blotting was used to determine the protein levels of Bax, Bcl-2, Cleaved caspase 3, glial cell line-derived neurotrophic factor (GDNF) and stem cell factor (SCF) in the testes of mice. Immunofluorescence staining was performed to detect the expression of Cleaved caspase 3 and 3β hydroxysteroid dehydrogenase (3βHSD), and a TUNEL assay was used to confirm the location of apoptotic cells. The levels of testosterone (T), GDNF and SCF were measured by ELISA. We also evaluated the sperm quality in the cauda epididymides by measuring the sperm count, abnormality, survival rate and apoptosis rate. The results showed that there was no significant difference in testicular histology, ultrastructure or sperm quality between the shielded group and sham group. Compared with the sham/shielded group, the C-irradiation group exhibited a lower testis index and severely damaged testicular histology and ultrastructure at 4weeks after C-irradiation. The levels of apoptosis in the testes increased markedly in the C-irradiation group, especially in spermatogonial stem cells. The levels of serum T and testicular 3βHSD did not obviously differ between the sham group and the C-irradiation group, but the levels of GDNF and SCF in the testes increased in the C-irradiation group, compared with the sham group. In addition, the sperm count and survival rate decreased in the C-irradiation group, while the abnormality and apoptosis rate increased. Under these experimental conditions, the abscopal effects of C-irradiation induced testicular damage with regard to both structure and function and ultimately decreased sperm quality in mice. These findings provide novel insights into prevention and treatment targets for male reproductive damage induced by C-irradiation.

Establishment of a Mouse Asthenospermia Model through Triggering D-Galactose Mediated Oxidative Stress Injury.

Asthenospermia is defined as the forward motility of sperm less than 32%. This study aimed to establish a mouse model of asthenospermia through triggering D-galactose mediated oxidative stress. A total of 40 Kunming male mice were randomly divided into control group, low-dose group (administrating D-galactose at 60 mg/kg), high-dose group (administrating D-galactose at 120 mg/kg), and high-dose+feed addition group (administrating D-galactose at 120 mg/kg together with oral D-galactose). The testicular weight, testicular organ coefficient, sperm viability, sperm concentration, and survival rate of the tail of epididymis were measured. Oxidative damage of D-- galactose to the reproductive system of mice was evaluated by measuring superoxide dismutase (SOD) and malondialdehyde (MDA) in the testicular homogenate of mice. The sperm motility, motility rate, concentration, and survival rate of low-dose, high-dose and high-dose+feed addition group were decreased, compared to that in the control group. However, there background:was a significant difference between high-dose group/high dose+feed group and the control group (p<0.05): the forward motile sperm motility rate and total motility rate are accorded with critical criteria of asthenospermia. As compared with the control group, the activity of SOD of model group mice significantly decreased, and MDA concentration significantly increased (p<0.05), except for low-dose versus control group for SOD activity. This suggests that testicular tissues suffered from oxidative damage. This study successfully established a mouse asthenospermia model through D-galactose mediated oxidative stress injury. The establishment of asthenospermia model in this study would provide new promising insight and act as a potential approach for studying asthenospermia in vivo levels.

INNOVATIVE ULTRA-RAPID VITRIFICATION METHOD FOR FIVE OR FEWER TESTICULAR SPERMATOZOA FROM NON-OBSTRUCTIVE AZOOSPERMIC MEN AFTER MICROSURGICAL TESTICULAR SPERM EXTRACTION (MICRO-TESE)

Clinical outcomes of ICSI when using only five or fewer testicular spermatozoa after cryopreservation have been unsuccessful and are considered to be inferior to those using testicular fresh spermatozoa from Micro-TESE. A possible cause of these poor results has been the lack of a successful freezing technique. In these cases, repeated Micro-TESE and simultaneous oocyte pick up has been the only available treatment. We developed an innovative ultra-rapid vitrification method for five or fewer testicular spermatozoa from non-obstructive azoospermic men after Micro-TESE. This study included 113 non-obstructive azoospermic men using Micro-TESE, it was conducted retrospectively at St. Mother Clinic between 2011 and 2018. Each motile spermatozoon was carefully aspirated tail first into the pipette, put into a 2-μl microdroplet media of the vitrification medium near the tip of the Cryotop (Kitazato Corporation, Tokyo, Japan) submerged in liquid nitrogen vapor for 2 min and then immediately plunged into liquid nitrogen. The vitrified spermatozoa were warmed by dipping them into a droplet media. Successfully recovered motile sperm were selected and used for ICSI. Number of patients, transfer cycles and collected sperms were 113, 192 and 560. Mean age of patients and their wives were 32.0±3.7y and 28.4±5.8y. Clinical pregnancy rate, miscarriage rate, live birth rate and number of live offspring were 24.0% (46/192), 19.6% (9/46), 19.3% (37/192) and 37 (Male: Female = 17: 20). Sperm recovery rate and survival rate were 90.3% (506/560) and 70.4% (356/506). Fertilization rate and mean number of transferred embryos were 51.6% (99/192) and 1.73 (1-2). Mean gestational weeks and mean body weight at birth were 39.23±5.27w and 2852.31±314.28g. No congenital anomalies were observed in any of the babies. This method for cryopreserving five or fewer spermatozoa from non-obstructive azoospermic men showed a recovery rate above 90% and a survival rate of about 70% which is the highest data ever been reported. The clinical pregnancy rate was 24.0% and 37 healthy babies were born.

P–023 Innovative ultra-rapid vitrification method for five or fewer testicular spermatozoa from non-obstructive azoospermic men after microsurgical testicular sperm extraction (Micro-TESE)

Abstract Study question What technique can be used to successfully cryopreserve five or fewer testicular spermatozoa from non-obstructive azoospermic men? Summary answer This method for cryopreserving five or fewer spermatozoa from non-obstructive azoospermic men showed a recovery rate above 90% and a survival rate of about 70%. What is known already Clinical outcomes of ICSI when using only five or fewer testicular spermatozoa after cryopreservation have been unsuccessful and are considered to be inferior to those using testicular fresh spermatozoa from Micro-TESE. A possible cause of these poor results has been the lack of a successful freezing technique. In these cases, repeated Micro-TESE and simultaneous oocyte pick up has been the only available treatment. Study design, size, duration Evaluation of the efficiency of cryopreservation by modified permeable cryoprotectant-free vitrification method (HTF supplemented with 0.1M sucrose and 10% SPS) for five or fewer testicular spermatozoa from 113 non-obstructive azoospermic men using Micro-TESE was conducted retrospectively at St. Mother Clinic between 2011 and 2018. Participants/materials, setting, methods This study included 113 non-obstructive azoospermic men. Each motile spermatozoon was carefully aspirated tail first into the pipette, put into a 2-μl microdroplet media of the vitrification medium near the tip of the Cryotop (Kitazato Corporation, Tokyo, Japan) submerged in liquid nitrogen vapor for 2 min and then immediately plunged in liquid nitrogen. The vitrified spermatozoa were warmed by dipping them into a droplet media. Successfully recovered motile sperm were selected and used for ICSI. Main results and the role of chance Number of patients, transfer cycles and collected sperms were 113, 192 and 560. Mean age of patients and their wives were 32.0±3.7y and 28.4±5.8y. Clinical pregnancy rate, miscarriage rate, live birth rate and number of live offspring were 24.0% (46/192), 19.6% (9/46), 19.3% (37/192) and 37 (Male: Female = 17: 20). Sperm recovery rate and survival rate were 90.3% (506/560) and 70.4% (356/506). Fertilization rate and mean number of transferred embryos were 51.6% (99/192) and 1.73 (1–2). Mean gestational weeks and mean body weight at birth were 39.23±5.27w and 2852.31±314.28g. No congenital anomalies were observed in any of the babies. Limitations, reasons for caution The maximum number of spermatozoa to which this method can be applied successfully is about 10. When the number of aspirated spermatozoa is over 10, some of them change direction and reach the mineral oil, and once this happens, they cannot be expelled out of the pipette. Wider implications of the findings: This technique is very useful for the cryopreservation of very small numbers of testicular spermatozoa (fewer than 10) in order to avoid or reduce Micro-TESE interventions. Trial registration number N/A

Effect of Semen Freezing and Thawing on Sperm Survival And Motility Rate : A Comparative Analysis

Objective : To compare the rate of sperm survival and motility through semen freezing and thawing during infertility treatment. Methodology: In this bidirectional observational study, we enrolled 31 patients who underwent semen analysis for infertility treatment at the Institute of Reproduction and Child Cares &amp; IRCC IVF Centre, Panchkula, Haryana, from June 2020 to December 2020. Out of these patients, 21 (67.74 %) were considered for semen freezing and thawing. For the rest of the ten patients (32.25 %), sperm count and motility were not good, and we excluded them from this study. Semen freezing based upon sperm count and motility were done. We did semen thawing after two weeks of semen freezing and recorded the sperm survival and motility. Results: Post thaw sperm survival rate and motility was 37.66% compare to pre-cryopreservation (61.82%). The observed rate of sperm motility declining was 24.16 % after cryopreservation/freezing. Conclusion: The present study results concluded that sperm's cryopreservation results in a decrease in sperm motility. There is a need of finding more accurate and reliable methods to freeze and thaw semen.

A systematic review on the effect of Ginger (Zingiber officinale) on improvement of biological and fertility indices of sperm in laboratory animals, poultry and humans.

There is an evidence that ginger enhance semen quality via improving different sperm parameters mainly count, viability, motility, morphology and DNA integrity. According to research results in various species, ginger seems to have strong antioxidant properties (due to the presence of active phenolic compounds) and androgenic activity. Ginger improves semen quality and increases fertility of sperm by disrupting the production of free radicals, dissolving oxidative chain reactions, reducing oxidative stress and altering the levels of gonadotropin hormones (LH, FSH) and sex hormones (such as testosterone). The antioxidant and androgenic properties of ginger give a sperm with normal morphological structure (head, middle and tail) and more integrated chromatin. The rate of DNA failure and damage to the mitochondrial genome in these cells is minimal and they have the most progressive motility, the highest viability and the best fertility. Therefore, the use of the ginger significantly improves the biological parameters of sperm (number, total motility, survival rate and normal morphology) and also increases all specialized fertility indicators of sperm. Tacking account of lacking literature and possibility of toxicity and adverse effect of ginger on vital organ, further clinical trial especially on evaluating the safety and clinical effect must be considered. Also, dose and duration of consumption by monitoring of health indicators and biochemical changes in all species such as human, animal and poultry must be applied.

Exploring the Mechanism of Zhibai Dihuang Decoction in the Treatment of Ureaplasma Urealyticum-Induced Orchitis Based on Integrated Pharmacology

Background: Ureaplasma urealyticum (UU) infection is the most common cause of male infertility. Zhibai Dihuang Decoction (ZBDHD) can improve the rate of forwarding motility sperm, sperm deformity rate, seminal plasma zinc and refined berry sugar levels. Methods: The potential targets of ZBDHD are obtained from The Encyclopedia of Traditional Chinese Medicine (ETCM). Orchitis-related targets were collected from the Genecards and OMIM databases. The Cytoscape and the Database for Annotation, Visualization and Integrated Discovery (DAVID) were utilized to construct and analyzed the networks. Finally, a rat model of orchitis caused by UU infection was used to detect related indicators of mitochondrial energy metabolism using TUNEL apoptosis detection technology, loss cytometry, Real-Time Quantitative Reverse Transcription PCR (qRT-PCR) and Western Blot. Results: A total of 795 ZBDHD targets and 242 orchitis-related targets were obtained. The “ZBDHD- orchitis PPI network” was constructed and analyzed. ZBDHD can regulate signaling pathways and biological processes related to mitochondrial energy metabolism. The results of experimental studies have shown that ZBDHD maintains the integrity of sperm mitochondrial respiratory chain function by enhancing mitochondrial Na+-K+-ATPase and Ca2+-Mg2+-ATPase activities, promotes the synthesis of mitochondrial ATP, and improves sperm energy supply, thereby improving the motility, vitality and survival rate of sperm, and effectively improving the quality of semen in UU-infected rats (p < 0.05). Conclusion:This study discovered the multi-pathway mechanism of ZBDHD intervention in UU-induced orchitis through integrated pharmacological strategies, which provides a reference for further research on the mechanism of ZBDHD intervention in orchitis in the direction of mitochondrial energy metabolism.

Survival of Cryopreserved Stallion Spermatozoa and the Level of Oxidative Modification of Sperm Plasma Proteins

Sperm freezing is one of the ways to preserve the sperm of rare, high-productive, physically strong stallions. Freezing negatively affects the reproductive characteristics of sperm, because of an increase in oxidative stress (OS) in particular. Macromolecules, especially proteins, are damaged in the result of OS. Aldehyde and ketone derivatives are formed, which are stable and available for diagnosis. The aim of the study was to determine the level of spontaneous oxidative protein modification (OPM) of sperm plasma in stallions with high and low sperm survival rates during hypothermic storage of cryopreserved sperm. Two experimental groups were formed: the 1st group consisted of 10 stallions, the survival rate of thawed after freezing spermatozoa of which ranged from 6 to 18h (average 11.0±2.8), the 2nd group included 12 stallions with sperm survival from 72 to 96h (average value - 84.1±2.4). Sperm survival time was defined as the ability of spermatozoa to maintain progressive motility above 5% during hypothermic (+4°C) storage of sperm. To assess the level of OPM in spermoplasma, sperctrophotometric analysis of 2,4-dinitrophenylhydrazones (DNPH) formed during the interaction of carbonyl derivatives of proteins with 2,4-dinitrophenylhydrazine (spectrophotometer “SF-2000”, Russia) was used. The R.L. Levine’ technique was modified by E.E. Dubinina. Measurements of DNPH derivatives were carried out at 14 wavelengths to record neutral aldehyde dinitrophenylhydrazones (ADNPHn) in the range 260-280nm, the main character (ADNPHb) - 258-264 and 428-520nm, to determine ketone-dinitrophenylhydrazones of a neutral nature (CDNPHn) 363-370nm, basic character (CDNPHb) - 430-434 and 524-535nm. The area under the absorption spectrum of DNPH derivatives of carbonyl derivatives of proteins (S) was analyzed. The total level of OPM in sperm plasma in group 1 stallions was statistically significantly higher than in group 2 stallions (677.7 Eod/g protein and 476.2 Eod/g protein, respectively, p<0.05). The result indicates a higher severity of OS in the sperm plasma of stallions with a low sperm survival rate, more significant damage to amino acid residues of proteins. OPM of sperm plasma leads to changes in their secondary, tertiary structure that negatively affects the functional state of proteins, the morphological and reproductive characteristics of sperm deteriorate. Damaged proteins can undergo aggregation and fragmentation processes. The accumulation of OPM products, protein aggregates resistant to proteolysis, disrupts cell metabolism, leading to apoptosis. The total level of OPM of sperm plasma in stallions with low survival rates of thawed after freezing spermatozoa is higher than in animals with high survival rates.

Sperm characteristics and growth of Rhamdia quelen males fed on rations formulated with vegetable food and different digestible energy levels supplied by soy bean oil / Short Communication Características espermáticas e crescimento de machos Rhamdia quelen alimentados com rações formuladas com alimentos vegetais e diferentes níveis de energia digestível fornecidos pelo óleo de soja

This research was carried out to evaluate the effects of rations formulated with vegetable foods and different levels of digestible energy supplied by soybean oil, on sperm characteristics and growth of Rhamdia quelen males. The fishes were fed on over 255 days with rations containing 350 g crude protein (CP) kg -1 and 2700, 2950, 3200, 3450 and 3700 kcal of digestible energy (DE) kg -1 . A randomized experimental design was used with five treatments and three replicates. A 8 m 2 tank containing six females and three males was an experimental unit. The fishes were weighted fortnightly to correct the feeding rate. All males receipted an intramuscular injection of 2.5 mg of crude pituitary extract (CPE) kg -1 to perform the sperm collection. The sperm characteristics were assessed on fresh sperm immediately after collection. All data were subjected to ANOVA and regression analysis. The growth, sperm volume, sperm concentration, or time of sperm motility were not affected (p &lt; 0.05) by rations. In addition, ration containing 3318.75 kcal of DE kg -1 improved the integrity of sperm membrane, suggesting a theoretical higher sperm survival rate of 98.22%. In conclusion, ration formulated with vegetable foods and different levels of soy bean oil as energy source did not interfere on growth of Rhamdia quelen males. In addition, the diet energy supplied by mainly soy bean oil offered a nutritional condition that improved the sperm quality since males fed on ration with 3318.75 kcal of digestible energy kg -1 has been increased the membrane integrity of sperm cell.

Effect of diluent variation on cryopreservation of large yellow croaker Larimichthys crocea

The aim of this research was to investigate different factors, including cryoprotective agents (CPAs), diluents, dilution ratios, equilibrium times, freezing rates, and thawing methods to optimize cryopreservation protocols for large yellow croaker (Larimichthys crocea). The parameters evaluated were sperm motility, sperm activity index (SAI), survival rate, and DNA damage. Different types of CPAs, such as dimethyl sulfoxide (DMSO), propylene glycol (PG), ethylene glycol (EG), methanol, and glycerol, were tested for sperm preservation. The highest motility, SAI, and survival rate were observed when EG was used. Different diluents such as Stein’s solution, Hank’s balanced salt solution, marine fish Ringer’s solution, artificial seminal plasma (ASP) of small yellow croaker, and Cortland solution were investigated. The highest post-thaw motility was observed upon using ASP as the diluent. Different concentrations of EG were then mixed with ASP to identify the optimal EG concentration. Experimental results showed that the motility (70.33 ± 1.20%), SAI (5), and survival rate (78.30 ± 0.42%) of post-thaw sperm were optimum when 10% EG and ASP were used as the CPA and diluent of cryopreservation, respectively. Post-thaw sperm motility was high at equilibration times below 150 s and at an optimum dilution ratio of 1:1 (sperm: CPA + diluent) and was not significantly different compared with fresh sperm motility. The freezing rate was found to be slow below −10°C/min. The thawing temperature of 45°C was identified as ideal. The percentage of tail DNA in post-thaw sperm at 10% EG and ASP was also investigated and was found to have more significant DNA damage than that in fresh sperm but significantly lower damage than that in post-thaw sperm at EG concentrations of 5%, 15%, and 20% (p &lt; 0.05). The cryopreservation protocols obtained in this study will be useful in large yellow croaker hatcheries.

PSV-39 The relationship between structural integrity and sperm survival during hypothermic storage of stallion sperm

Abstract The aim of this work was to study the relationship between ultrastructural integrity and sperm survival during hypothermic storage of chilled and cryopreserved stallion sperm. Semen from 37 stallions, were collected during the breeding season. The structural integrity of sperm was studied using a Hitachi 700 electron microscope. Progressive motility was evaluated using an Olympus BX 41 phase contrast microscope. Sperm survival was defined as the ability of spermatozoa to maintain progressive motility above 5% during hypothermic (+4C) storage of sperm. Spearman nonparametric correlation analysis was used. Sperm survival at a temperature +4°C in chilled semen averaged 115.4±11.47 hours. After freezing sperm survival decreased on average to 48.9±5.56 hours (P &amp;lt; 0.001). A reliable correlation between the number of sperm with intact heads and sperm survival at a temperature of + 4°C in diluted (r=0.60; P &amp;lt; 0.001) and thawed (r=0.63; P &amp;lt; 0.001) sperm was established. A significant negative correlation between the number of spermatozoa with degradation of the acrosome in native sperm and their survival (r=-0.48; P &amp;lt; 0.01) under the hypothermic storage was found. For cryopreserved semen, a similar dependence has not been established. A significant correlation between the number of spermatozoa with the normal position of the acrosome and the sperm survival rate in native sperm was also established (r=0.52; P &amp;lt; 0.01). A significant negative correlation between sperm with acrosome hypoplasia and sperm survival was found (r=-0.67; P &amp;lt; 0.001). We assume that the structural integrity and ability of spermatozoa to maintain progressive motility for a long time under conditions of hypothermic storage depends on the quality of spermatogenesis and the resistance of cells to low and ultra-low temperatures. Authors acknowledge financial support from Russian Science Foundation, Grant No: 20-16-00101, development program of Bioresource collections “Cryobank of genetic recourses the All-Russian Research Institute for Horse Breeding.”

Effects of Cryoprotective Medium Composition, Dilution Ratio, and Freezing Rates on Spotted Halibut (Verasper variegatus) Sperm Cryopreservation.

Simple SummaryThe spotted halibut, Verasper variegatus, is a popular fish species occurring naturally in the East China Sea and coastal areas of Korea and Japan. However, when reared in captivity, male and female spotted halibut do not usually mature synchronously. Maintaining production of this commercial fish in hatcheries through sperm cryopreservation is important. This study investigated the effect of several factors for successful cryopreservation of fish sperm including cryoprotective agents (CPAs), diluents, dilution ratios (Milt: CPA + diluents), and freezing rates. The observed factors significantly affected movable sperm ratio (MSR), sperm activity index (SAI), survival rate, and DNA damage after cryopreservation. In the present study, the mixture of 15% dimethyl sulfoxide (DMSO) + 300 mM sucrose with a dilution ratio lower than 1:2 and a freezing rate slower than −5 °C/min provided the best treatment and reduced DNA damage. These results can contribute to the development of a cryopreservation protocol for spotted halibut hatcheries.The spotted halibut is species that has a high potential market value in Korea, but the supply of seed is unstable because of the limited milt production of males. The objective of this research was to explore different aspects, such as CPAs, diluents, dilution ratio, and freezing rates, to develop an optimal sperm cryopreservation. The parameters assessed were movable sperm ratio, sperm activity index, survival rate, and DNA damage. The CPAs tested in this research were propylene glycol, dimethyl sulfoxide (DMSO), methanol, ethylene glycol, and glycerol. Different diluents, including 300 mM sucrose, 300 mM glucose, Stain’s solution, and Ringer’s solution, were investigated. The previous experiment showed that the optimal CPA for cryopreservation was DMSO with a concentration of 15% with 300 mM as diluent. To determine the effect of the dilution ratio, sperm was diluted to 1:1, 1:2, 1:10, 1:100, and 1:1000 with 300 mM sucrose containing DMSO at a final concentration of 15%. Lastly, the optimal freezing rate of the sperm was evaluated with four different freezing rates (−1, −5, −10, and −20 °C/min). Post-thaw sperm motility was higher with a dilution ratio lower than 1:2, and the freezing rate was less than −5 °C/min. In conclusion, these findings represent the development of a cryopreservation protocol for spotted halibut.

The epigenetic alterations of human sperm cells caused by heroin use disorder.

The molecular mechanisms of drug use on sexual health are largely unknown. We investigated, the relationship between heroin use disorder and epigenetic factors influencing histone acetylation in sperm cells. The volunteers included twenty-four 20- to 50-year-old men with a normal spermogram who did not consume any drugs and twenty-four age- to BMI-matched men who consume only the drug heroin for more than last four months. HDAC1 and HDAC11 mRNA expression levels in spermatozoa and miR-34c-5p and miR-125b-5p expression levels in seminal plasma were measured. The heroin-user group showed significantly increased white blood cell counts and decreased sperm motility and survival rates (8.61±1.73, 21.50±3.11, 69.90±4.69 respectively) as compared to the control group (1.49±0.32, 38.82±3.05, 87.50±0.99 respectively) (p≤.001). An increase in DNA fragmentation index (DFI) (heroin-user group: 41.93±6.59% and control group: 10.14±1.43%, p=.003), a change in frequency of HDAC1 (heroin-user group: 1.69±0.55 and control group: 0.45±0.14, p=.045) and HDAC11 (heroin-user group: 0.29±0.13 and control group: 2.36±0.76, p=.019) in spermatozoa and a significant decrease in seminal miR-125b-5p abundance (heroin-user group: 0.37±0.11 and control group: 1.59±0.47, p=.028) were reported in heroin consumers. Heroin use can lead to male infertility by causing leukocytospermia, asthenozoospermia, DFI elevation in sperm cells and alterations in seminal RNA profile.