Rooster Sperm Research Articles

Effects of Melatonin On The Rooster Sperm Quality During Cryopreservation

Effects of Melatonin On The Rooster Sperm Quality During Cryopreservation

Effects of Rutin On The Quality of Rooster Sperm During Cryopreservation

Effects of Rutin On The Quality of Rooster Sperm During Cryopreservation

Chronic Dietary Exposure of Roosters to a Glyphosate-Based Herbicide Increases Seminal Plasma Glyphosate and AMPA Concentrations, Alters Sperm Parameters, and Induces Metabolic Disorders in the Progeny.

The effects of chronic dietary Roundup (RU) exposure on rooster sperm parameters, fertility, and offspring are unknown. We investigated the effects of chronic RU dietary exposure (46.8 mg kg−1 day−1 glyphosate) for 5 weeks in 32-week-old roosters (n = 5 RU-exposed and n = 5 control (CT)). Although the concentrations of glyphosate and its main metabolite AMPA (aminomethylphosphonic acid) increased in blood plasma and seminal fluid during exposure, no significant differences in testis weight and sperm concentrations were observed between RU and CT roosters. However, sperm motility was significantly reduced, associated with decreased calcium and ATP concentrations in RU spermatozoa. Plasma testosterone and oestradiol concentrations increased in RU roosters. These negative effects ceased 14 days after RU removal from the diet. Epigenetic analysis showed a global DNA hypomethylation in RU roosters. After artificial insemination of hens (n = 40) with sperm from CT or RU roosters, eggs were collected and artificially incubated. Embryo viability did not differ, but chicks from RU roosters (n = 118) had a higher food consumption, body weight and subcutaneous adipose tissue content. Chronic dietary RU exposure in roosters reduces sperm motility and increases plasma testosterone levels, growth performance, and fattening in offspring.

Rooster frozen-thawed semen quality following sublethal xanthine oxidase treatments

Reactive oxygen species are associated with cryodamage and may be a factor causing or exacerbating cellular cryodamage during freezing and thawing processes. Induction of sublethal oxidative stress as a new approach for preconditioning of sperm improves the cryo-resistance of sperm. The aim of this study was to investigate effects of sublethal concentrations of xanthine oxidase (XO), which induces oxidative stress before cryopreservation on values for semen quality variables of rooster sperm post-thawing. Semen samples were collected from 15 roosters and treated with different concentrations of XO [XO-0, XO-0.005, XO-0.05, XO-0.5, XO-5, and XO-50 U/ml]; then, the effects of treatments with XO as sublethal stressors, were examined. Results indicated the XO-0.5 and XO-5 treatments resulted in a greater percentage of sperm total motility, progressive motility, viability, and membrane functionality compared to other groups. There was no difference after treatments with XO-0, XO-0.005, and XO-0.05 on sperm total motility, membrane functionality, apoptosis, mitochondria activity, and viability. There was a greater percentage of mitochondria activity in sperm of the XO-0.05, XO-0.5, and XO-5 groups. Furthermore, there was the greatest concentration of malondialdehyde (MDA) in samples of the XO-50 group. Values for sperm abnormal morphology, acrosome integrity, and DNA fragmentation were not different among samples post-thawing. Sperm treated with XO-0.5 and XO-5 had a greater fertilization capacity than those of the control group. In conclusion, treatment of sperm with 0.5 and 5 U/ml XO as inducers of mild oxidative stress before cryopreservation, improved several function quality indices of sperm post-thawing.

Supplementation of Beltsville extender with quercetin improves the quality of frozen-thawed rooster semen

ABSTRACT 1. Excessive reactive oxygen species (ROS) production during the sperm freeze-thawing process leads to membrane lipid peroxidation, DNA damage, and motility loss. 2. This study examined the effect of supplementation of Beltsville poultry semen extender with different concentrations of quercetin (and antioxidants) on the cryopreservation of rooster sperm. 3. Semen samples were collected from six Ross broiler breeders via abdominal massage twice a week for 4 weeks (eight replicates), and were divided into five equal aliquots to be diluted in Beltsville extenders that contained different concentrations of quercetin: 0, 5, 10, 15, and 20 mM. Motility, membrane functionality, abnormal morphology, lipid peroxidation, mitochondrial activity, viability, apoptosis status, and fertility potential were assessed post thaw. 4. The addition of 10 and 15 mM quercetin to the semen extender significantly increased the total motility, straight-line velocity (VSL), and sperm membrane functionality compared with the other groups (P ≤ 0.05). Moreover, 10 mM quercetin caused higher progressive motility (34.86 ± 3.80%), curvilinear velocity (VCL; 175.11 ± 3.20 µm/s), average path velocity (VAP; 44.35 ± 11.06 µm/s), viability (59.14 ± 1.36%), mitochondrial activity (80.14 ± 2.07%), lower abnormal morphology (19.21 ± 0.45%), and lower lipid peroxidation (2.7 ± 0.13 nmol/ml) compared with the other groups (P < 0.05). The rate of fertility and hatchability after artificial insemination was not affected by experimental groups. 5. In conclusion, supplementation of Beltsville extender with 10 mM quercetin could be a suitable method to improve post-thawed rooster sperm quality resulting in better freeze/thaw characteristics.

Role of Mono- and Disaccharide Combination in Cryoprotective Medium for Rooster Semen to Ensure Cryoresistance of Spermatozoa.

The combination of saccharides in the composition of a cryopreservation medium may represent a promising method for the preservation of the reproductive cells of male birds. In the current study, cryoprotective media with a combined composition of mono- and di-saccharides were developed. The degree of penetration of reducing saccharide molecules (maltose—Mal20 medium) and non-reducing disaccharide molecules (trehalose—Treh20 medium) from the cryoprotective medium into the cytosol of rooster spermatozoa was studied. LCM control media without disaccharides were used as the control. The number of maltose molecules penetrating from the outside into the cytosol of the spermatozoon was 1.06 × 104, and the number of trehalose molecules was 3.98 × 104. Using a combination of maltose and fructose, the progressive motility of frozen/thawed semen and the fertility rates of eggs were significantly higher ((p < 0.05) 40.2% and 68.5%, respectively) than when using a combination of trehalose and fructose in a cryoprotective diluent (33.4% and 62.4%, respectively). A higher rate of chromatin integrity at the level of 92.4% was obtained when using Treh20 versus 74.5% Mal20 (p < 0.05). Maltose positively affected the preservation of frozen/thawed sperm in the genital tract of hens. On the seventh day from the last insemination when using Mal20, the fertilization of eggs was 42.6% and only 27.3% when using Treh20. Despite the same molecular weight, maltose and trehalose have different physicochemical and biological properties that determine their function and effectiveness as components of cryoprotective media.

Functional difference of ATP-generating pathways in rooster sperm (Gallus gallus domesticus)

Adenosine triphosphate (ATP) production via glycolysis and oxidative phosphorylation is essential for the maintenance of flagellar motility in sperm; however, the primary energy production pathways supporting fertilization vary among species. Inconsistency in thought exists regarding which pathways maintain ATP production and sperm motility in poultry. Glycolysis and mitochondrial oxidation contribute to flagellar motion in chicken sperm, but the relative dependence on these pathways for motility and penetrability into the inner perivitelline layer remains unclear. In the present study, there was use of various inhibitors and energy substrates to evaluate the relative contribution of anaerobic glycolysis and mitochondrial oxidation to chicken sperm flagellar motility, ATP production, and penetrating capacity through the perivitelline layer. Although both pathways contributed to these processes to varying extent, glucose was the primary substrate for sperm penetration into the inner perivitelline layer in chickens. Furthermore, results from metabolic stress analyses indicated that there was less perivitelline penetrability in response to pyruvate that was not due to changes in reactive oxygen species or intracellular pH. Overall, results from the present study indicate glycolysis and mitochondrial oxidation pathways have distinct functions in the flagellar motility and penetrability of the perivitelline membrane by rooster sperm. There, therefore, are new insights as a result of findings in the present study into the energy production system of sperm through which there is utilization of extracellular metabolic substrates for maintaining sperm fertilization capacity.

The effect of adding pentoxifylline to the Ross rooster semen on quality parameters of spermatozoa during 6 hours of storage at cooling temperature

The most important factors involved in sperm fertility are motility, morphology and sperm membrane integrity. The studies show that, the addition of Pentoxifylline(PTX) can increase sperm motility by increasing energy production. The aim of the present study was to investigate the effect of adding Pentoxifylline to rooster semen on qualitative parameters of rooster sperm after 6 hours of storage at 4°C. Semen samples were collected from 15 Ross 308 roosters at 28 weeks of age. After initial evaluations, semen samples were mixed together to eliminate individual differences. After diluting the semen and adding different levels of Pentoxifylline (0, 0.25, 0.5, 0.75 and 1 μM), the samples were kept at 4°C for 6 hours. Then, sperm kinetic parameters were evaluated. The results showed that the addition of 0.75 μM PTX significantly improved progressive mobility, curvilinear velocity, viability and plasma membrane integrity of spermatozoa compared to the control group (P<0.05). In overall, it can be concluded that methylxanthine-derived drugs such as Pentoxifylline can have a significant effect on the motility, viability and plasma membrane integrity of rooster sperm in the short period of time.

Effect of Ginger Extract on Semen Parameters and Sperm DNA Fragmentation during the Liquid Storage at 4°C in Native Roosters (Research Note)

The goal of the current study was to determine the effect of ginger extract on some quality parameters of rooster spermatozoa during liquid storage. A total of 10 healthy adult native roosters were selected and sampled twice a week for three weeks. Treatment groups were made up of basal extender supplemented with various concentrations of ginger extract including 0, 20, 40, 60, and 80 μg/mL. Sperm plasma membrane integrity, viability, and motility were assessed at 0, 24, 48, and 72 hours after semen collection. However, the sperm DNA fragmentation test was determined only after 48 hours of storage. The sperm membrane integrity was high at 24 (81.66%) and 48 (58.66%) hours in control and 60 µg/mL of ginger extract, respectively. The highest sperm viability (70.50%) and motility (76.50%) were obtained in 20 μg/mL ginger extract at 48h. The lowest sperm DNA fragmentation (32%) was observed in the 20 μg/mL of ginger extract. According to the results, the ginger extract showed a protective effect against time-associated harmful changes in semen quality of roosters during liquid storage.

COMPARATIVE ANALYSIS OF THE EFFECTS OF CURCUMIN AND EPICATECHIN ON THE VITALITY OF ROOSTER SPERMATOZOA

The objective of our study was to assess the effects of two selected antioxidants (curcumin - CUR and epicatechin - EPI) on the vitality of rooster spermatozoa. We used semen samples from 10 sexually mature Lohmann light roosters, which were collected from a local farm, diluted in PBS and cultured with different concentrations of CUR and EPI (1, 5, 10, 25, 50 and 100 µmol/L). Motility, membrane and acrosome integrity were analysed after 0, 2 and 24 h of in vitro culture. Sperm motility was evaluated by the computer-assisted sperm analysis (CASA). The membrane integrity was determined by a combination of eosin/nigrosin dyes, and the acrosome integrity was detected by fast green/rose bengal dyes. The data were statistically evaluated using One-way ANOVA and the Dunnet’s test. The results indicate that after 0 h of cultivation all selected parameters stay unchanged. After 2 h significant differences (P&lt;0.0001) were observed in case of the motility and membrane integrity. Following 24 h a significant (P&lt;0.0001) preservation of motility was recorded. The level of membrane damage and acrosome integrity was significantly decreased (P&lt;0.01). Based on our results we may conclude that protective effects of CUR and EPI on the vitality of rooster spermatozoa were observed already after 2 h. This impact was even more dominant after 24 h. CUR showed better effects at lower concentrations (1, 5, and 10 µmol/L) while EPI at higher doses (25 and 50 µmol/L). Our findings could contribute to the potential application of these antioxidants in avian reproduction technologies.

Type III antifreeze protein (AFP) improves the post-thaw quality and in vivo fertility of rooster spermatozoa

Antifreeze proteins (AFP) have the potential for improving sperm cryopreservation. We have applied Type III antifreeze protein (AFP3) on the cryopreservation of spermatozoa from broiler breeder roosters, aiming to enhance post-thawing quality and fertility. Semen was extended at 37°C in Lake's extender containing AFP3 at 0.01, 0.1, 1, 5, and 10 µg/mL (no AFP3 as control). Post-thawing sperm assessment included sperm motility (CASA), morphology, membrane functionality by hypoosmotic swelling test (HOST), lipoperoxidation as malondialdehyde (MDA) production, and sperm viability, early apoptosis (phosphatidylserine exposure as annexin V-positive staining in viable spermatozoa), and mitochondrial activity by flow cytometry. Fertility was assessed after artificial insemination (30 hens/treatment). Total and progressive motility, membrane functionality, and mitochondrial activity increased in 0.1 and 1 µg/mL AFP, compared to control and other concentrations, whereas apoptosis was significantly lower. VAP, VSL, and viability were significantly higher for 1 µg/mL AFP3 than with the other treatments except for 0.1 µg/mL (which was not always significantly different from the control or other concentrations), and with abnormal forms being significantly lower. The proportion of fertilized and hatched eggs was also higher for 1 µg/mL AFP3, with 0.1 µg/mL also showing significantly higher results than the control, and no differences with other concentrations). In conclusion, 1 µg/mL AFP3 could improve the post-thawing results of rooster spermatozoa frozen in Lake's extender. According to our results, concentrations between 1 and 0.1 µg/mL could be similarly efficient.

Poloxamer 188 and hydroxyethyl starch have a cryoprotective synergic effect improving post-thawing quality and fertility of rooster spermatozoa

Poloxamer and hydroxyethyl starch have cytoprotective effects. In the present study, effectiveness was evaluated of these compounds as a cryoprotectant for rooster semen. In Experiment 1 (E1), poloxamer 188 (1%, P188), poloxamer 407 (1%, P407), and control groups were compared after sperm cryopreservation. Experiment 2 (E2) was conducted with 3%, 5%, and 7% of hydroxyethyl starch (H3, H5, H7), also combined with P188 (H3P188, H5P188, H7P188), based on results from E1. Sperm motility was assessed using CASA, abnormal forms and hypo-osmotic swelling (HOS) were evaluated using microscopy, and viability, apoptotic-like changes, and mitochondrial activity were determined using flow cytometry. In E2, there were assessments of fertility and hatching capacity. Results from E1 indicated total and progressive motility, velocity, membrane functionality, viability, and mitochondrial activity were greater with inclusion of P188 in semen extender, with less apoptotic-like changes (P < 0.05). In E2, HES inclusion in semen extender improved total motility, membrane functionality, and mitochondrial activity (P < 0.05), especially H5, which also markedly increased sperm viability and decreased apoptotic-like changes. The combination of P188 with HES increased sperm quality overall, with inclusion of H5P188 resulting in increases of progressive motility and VSL (P < 0.05). The H5 inclusion also increased proportion of fertilized eggs (P < 0.05). Furthermore, the combination of HES and P188 increased proportions of fertilized and hatched eggs compared with the control, with inclusion of H5P188 having the greatest effects. In conclusion, supplementation of semen extender with H5P188 increases post-thawing quality and fertility of rooster sperm, being a safe and effective method for the poultry industry.

Research Note: Evaluation of two methods for adding cryoprotectant to semen and effects of bovine serum albumin on quality characteristics of cryopreserved rooster spermatozoa

Chicken semen cryopreservation is a tool for programs of genetic diversity management and endangered breeds conservation. Due to physiological features, the fertility rates of cryopreserved poultry sperm are lower than mammal species. Thus, improvement of the semen cryopreservation methods is required. A first study was performed by a 2 × 2 factorial design consisting of 2 methods of adding the cryoprotectant [Direct or Diluted (mixed with extender medium)] and 2 cryoprotectants (glycerol and dimethylacetamide). Then sperm quality indicators were evaluated after freezing. A second study with a 2 × 2 design was conducted to evaluate the effectiveness of bovine serum albumin (BSA) on the optimization of 2 different extenders (Lake and Animal Sciences Group [ASG]). Viability and motility variables were evaluated before and after freezing. There was no significant difference in sperm viability and motility variables between Direct or Diluted methods. Supplementation of extenders with BSA improved most of the sperm motility variables in both extenders before and after freezing. Progressive sperm, non-progressive sperm before freezing, and all post-thaw sperm motility parameters, except amplitude of lateral head displacement and beat-cross frequency, were increased in BSA-supplemented extenders (P < 0.05), and BSA improved sperm viability in ASG extender after thawing (P < 0.05). After thawing, the interaction between extender and BSA (P < 0.05), eliminated the differences between the 2 BSA-supplemented media in curvilinear velocity, straight-line velocity, average path velocity, and amplitude of lateral head displacement which were higher in non-supplemented ASG extender than nonsupplemented Lake medium. In conclusion, the direct or diluted methods of adding glycerol or dimethylacetamide, did not significantly affect the post-thaw sperm characteristics. BSA positively affected most of the post-thaw sperm motility indicators regardless of the type of extender and resulted in significantly higher post-thaw sperm viability in ASG medium.

The effect of methyl-beta-cyclodextrin on DNA absorption and quality of posttransfected sperm

Sperm can be selected as a natural vector for the production of transgenic animals. Methyl-beta-cyclodextrin (MBCD) removes cholesterol from the phospholipid membrane of sperm and improves the efficiency of DNA uptake by sperm. In experiment 1, fresh sperm was treated with various concentrations of MBCD. The direct effects of MBCD on sperm parameters were monitored. In experiment 2, different concentrations of MBCD (0, 1, 2, and 4 mmol) were assessed for the transfection of genetically exogenous construction to rooster sperm. Washed semen was divided into 5 equal groups for the incubation and transfection with a pcDNA3.1+/hG-CSF vector (exogenous DNA) as follows; Treatment I—Control (washed semen without DNA); Treatment II—Control (washed semen with DNA); Treatment III—(washed semen incubated with DNA and 1 mmol MBCD); Treatment IV—(washed semen incubated with DNA and 2 mmol MBCD); and Treatment V—(washed semen incubated with DNA and 4 mmol MBCD). We demonstrated that rooster spermatozoa spontaneously can uptake exogenous DNA; this was assessed using exogenous DNA amplification (sperm genomic DNA used as a template for PCR reaction) after DNase I treatment. In addition, total motility (TM), progressive motility (PM), velocity parameters [curvilinear velocity (VCL), straight linear velocity (VSL), sperm track straightness (STR), linearity (LIN)], membrane integrity (MI), and membrane functionality were posttransfectionally evaluated. The concentrations of 1 and 2 mmol MBCD significantly (P < 0.05) improved the motion characteristics and membrane integrity of fresh sperm. The presence of hG-CSF in rooster sperm was detected by PCR and based on sperm analyses MBCD (1 mmol) improved the percentage of motility (98.9 ± 0.81), membrane functionality (64 ± 1.64), and MI (76.2 ± 1.65) after transfection when compared with the other groups (P < 0.05). For the production of transgenic chicken, hens were inseminated (AI) by transfected sperm treated with 1 and 0 mmol MBCD. A PCR analysis of the blood samples and dead embryo tissues of chicks did not reveal the transgene integration.

Royal jelly may improve sperm characteristics during preservation of rooster semen: Gene expression of antioxidant enzymes.

Sustainable production and the increasing number of embryonated hatching eggs are critical aspects of the poultry production industry. The present paper aims to appraise the effectiveness of royal jelly (RJ) on the semen characteristics of Native Mazandaran roosters in both liquid and frozen storage conditions. Semen collected from 10 sexually mature roosters and following dilution was supplemented with RJ at 0.0 (control), 5 (RJ5), 10 (RJ10), 20 (RJ20) and 40 (RJ 40) mg/ml. After cooling and freezing-thawing, the percentage of forward progressive motility, viability, abnormality, hypo-osmotic swelling test (HOST) and the mRNA abundance of antioxidant enzymes of spermatozoa were measured. Our results revealed that the addition of 5mg/ml RJ to the semen extender significantly increased (p<.05) the percentages of forward progressive motility, viability and HOST during liquid and frozen storage. The abnormality of spermatozoa in the RJ5 group was significantly lower compared to the other groups. During liquid storage, a significant decrease in forward progressive motility was found after 48hr in comparison with 24hr at 4°C. High levels of RJ (from 10 to 40mg/ml) were severely decreased the characteristics of rooster spermatozoa in comparison with RJ5 and the control group. The inclusion of RJ at 5mg/ml to the semen extender enhanced the mRNA transcript of antioxidant enzymes of spermatozoa during liquid preservation. The mRNA abundance of antioxidant enzymes did not influence by cryostorage. Overall, these data suggest that supplementation of RJ at 5mg/ml to the extender improved semen characteristics and redox status of rooster spermatozoa.

Effects of the carbonated drink as an extender on semen characteristics, fertility and hatchability in Nigerian indigenous chicken

Semen extenders are liquid diluents that buffer sperm cells and preserve their fertilizing potentials. The commercial carbonated drink (CD) as an extender was evaluated on semen characteristics, fertility and hatchability in Yoruba ecotype chickens (YECs). The fructose of the CD was 1.52?0.05 mg/ml. Under the conditions of 370Celsius, 5% and 10% of CD were added to the egg yolk citrate solution to make 100%. Semen was obtained from ten matured Yoruba ecotype chicken cocks with an average weight of 1.8?0.2 kg. The semen was pooled in a test tube and added to the extenders for preservation at 0, 30 and 60 minutes, respectively, in a factorial design layout. Percentage motility of sperm cells was significantly (p&lt;0.05) higher in 5% CD inclusion compared with 10% CD inclusion and control. Motility decreased with an increase in preservation time across the treatments. The percentage of dead sperm cells decreased (p&lt;0.05) in 5% CD inclusion when compared with 10% CD inclusion and control. The sluggish sperm percentage increased significantly (p&lt;0.05) with semen preservation time. Fertility and hatchability of eggs were significantly (p&lt;0.05) higher in 5% CD inclusion. It was concluded that carbonated drinks at 5% inclusion in an extender could preserve cock sperm cells for 60 minutes with improved fertility and hatchability of eggs.

Effects of freezing extender supplementation with mitochondria-targeted antioxidant Mito-TEMPO on frozen-thawed rooster semen quality and reproductive performance

Rooster semen cryopreservation is a useful method to utilize semen samples for artificial insemination in commercial flocks, but with use of the freezing-thawing process there is a reduction in the quality and fertilization capacity of rooster spermatozoa post-thawing. The aim of the current study was to investigate the efficacy of the mitochondria-targeted antioxidant Mito-TEMPO on rooster sperm quality and fertilization capacity after conducting the freezing-thawing processes. Semen samples were diluted and there were five equal aliquots supplemented with 0, 0.5, 5, 50 and 500 μM Mito-TEMPO. Semen samples were subsequently cryopreserved in liquid nitrogen. After thawing, sperm motility, lipid peroxidation, membrane functionality, normal morphology, mitochondria active potential, acrosome integrity, viability, apoptotic-like changes, DNA fragmentation, hydrogen peroxide concentration and fertilizing capacity were evaluated. Supplementation of Lake medium with 5 and 50 μM Mito-TEMPO resulted in greater (P ≤ 0.05) total sperm motility, progressive motility, average path velocity, membrane functionality, mitochondria active potential, acrosome integrity and viability compared with semen of the other groups. Lipid peroxidation, late apoptotic-like changes, DNA fragmentation and hydrogen peroxide content, however, were less (P ≤0.05) in semen samples supplemented with 5 and 50 μM Mito-TEMPO compared to other groups. Furthermore, fertility percentages were greater when there was supplementation with 5 and 50 μM Mito-TEMPO compared to the control group. Mitochondria-targeted antioxidant Mito-TEMPO could be included in semen extender before cryopreservation to improve quality and fertilization capacity of rooster semen after thawing of cryopreserved samples.

14 Genome-wide association study of frozen sperm quality in cocks

Abstract Preservation of the genetic material of unique breeds of hens involves the use of semen cryobanks. The aim of the study was to search for genomic associations with rooster sperm quality after freezing. We took into account the decrease in sperm motility after thawing. Genotyping of 96 individuals (roosters) using the Affymetrix Axiom®600k Array chip was performed. Genomic data were obtained using the Plink 1.9 software. DNA samples were taken with genotyping quality at SNP loci more than 95%, SNP with MAF &amp;gt; 0.01. GWAS was performed using EMMAX. SNP information in the corresponding genes was determined using the genomic browsers NCBI and Ensembl. Assessment of rooster sperm quality showed high individual variability in total motility before freezing (20–95%), and after freezing (5–70%). Significant SNPs were detected on the chromosomes GGA1 (rs315598192, rs316943858), GGA2 (rs312981435), GGA3 (rs13694743), GGA11 (rs314024471), GGA19 (rs316346648), GGA22 (rs31797024248, rga31780824824, gga22 (rs3178082424). Candidate genes ZC3HC1, CACNA2D1, BCKDHB, AMFR, and GOSR1 were identified. The annotated genes were associated with cell cycle regulation, the formation of calcium cytoskeleton, and the formation of a multienzyme complex associated with the inner membrane of mitochondria, glycosylated transmembrane receptor. The experimental data and results can be used to study the localization of regions and genes associated with phenotypic traits of frozen sperm quality. Authors acknowledge financial support from Russian Science Foundation № 18-16-00071.

Effect of crocin and naringenin supplementation in cryopreservation medium on post-thaw rooster sperm quality and expression of apoptosis associated genes.

The aim of our study was to examine the effects of crocin (0.5 (C0.5), 1 (C1) and 1.5 (C1.5) mM) and naringenin (50 (N50), 100 (N100) and 150 (N150) μM) in cryopreservation extender for freezing rooster semen. Sperm motility, viability, abnormalities, membrane functionality, active mitochondria, apoptosis status, lipid peroxidation (LP), GPX, SOD, TAC, the mRNA expression of pro-apoptotic (CASPASE 3) and anti-apoptotic (Bcl-2) genes, fertile eggs, hatched eggs and hatching rate were investigated following freeze-thawing. C1 and N100 resulted in higher (P < 0.05) total motility and progressive motility in comparison to the control group. The C1 and N100 groups improved viability, membrane functionality and reduced lipid peroxidation. We found higher values for active mitochondria with C1 and N100 compared to control group. The C1 and N100 groups showed lower percentages of early apoptosis when compared with control group. Also, C1 and N100 had higher TAC, compared to the control group. The mRNA expressions of BCL-2 in the C1 and N100 groups were significantly higher than that of other treatments. The expression of CASPASES 3 was significantly reduced in C1 and N100 group (P < 0.05) when compared to control group. Significantly higher percentages of fertile eggs, hatched eggs and hatching rate were observed in C1 and N100 compared to the control group. In conclusion, crocin at 1 mM and naringenin at 100 μM seem to improve the post-thawing rooster semen quality, fertility and could protect the sperm by reducing the pro-apoptotic (CASPASE 3) and increasing anti-apoptotic (Bcl-2) genes.

Does alpha-lipoic acid–loaded nanostructured lipid carriers improve post-thawed sperm quality and ameliorate apoptosis-related genes of rooster sperm?

Oxidative stress could be prevented by antioxidant-loaded nanoparticles. The purpose of this study was to assess the effects of 10 (A10), 20 (A20), 30 (A30), 40 (A40), and 50 (A50) μM alpha-lipoic acid and alpha-lipoic acid nanostructured lipid carriers (ALN) at 10 (ALN10), 20 (ALN20), 30 (ALN30), 40 (ALN40), and 50 (ALN50) μM on post-thawed sperm quality, fertility, and apoptosis-related genes of rooster sperm. The extender supplemented with ALN30 led to higher total and progressive motility, straight-line velocity, and linearity in comparison to the control group. The ALN30 resulted in higher percentage of mitochondria activity and glutathione peroxidase level compared with control (P < 0.05). The extender supplemented with ALN30 led to lower percentage of apoptotic sperm, when compared with the control. CASPASE 3 expression in ALN30 was lower (P < 0.05) than the other groups. The results showed that BCL-2 mRNA expression of sperm was significantly (P < 0.05) higher in ALN30 compared with the other groups (P < 0.05). Higher percentages of fertility and hatchability rates were observed in ALN30 group. The results indicate that ALN30 could be regarded as a novel potential cryoprotectant for the cryopreservation of rooster semen. Therefore, nanostructured lipid carriers improve not only the active compound (such as alpha-lipoic acid) of biomedical applicability but also the potential for industrial application in sperm cryopreservation.