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Öğe A comparative study of seminal plasma and blood serum macro and trace elements in the the breeding (October) and the non-breeding (April) seasons in Merino ram(Hellenic Veterinary Medical Society, 2019) Bülbül, Bülent; Akalın, Pınar Peker; Başpınar, Nuri; Bucak, Mustafa Numan; Kırbaş, Mesut; Öztürk, Caner; Güngör, Şükrü; Akbulut, KübraIn this study, it was aimed to investigate the concentrations of macro and trace elements in seminal plasma and blood serum in the breeding (October) and the non-breeding (April) seasons in Merino Ram. Nineteen Merino Rams, aged 18-24 months, were involved in the study. Blood (once) and ejaculate samples (6 replicates) were taken in the breeding (October) and the non-breeding (April) seasons. Blood serum, seminal plasma and diet Calcium, Sodium, Potassium, Magnesium, Phosphorus, Sulfur, Zinc, Selenium, Chrome, Manganese, Nickel, Molybdenum and Boron concentrations were determined by ICP-AES. In blood serum, Sodium and Selenium concentrations were higher (p<0.05 and 0.001, respectively) in the the breeding season than in non-breeding season, whereas Potassium, Chromium and Boron concentrations were lower (p<0.05, 0.001 and 0.001, respectively) in the breeding season than in the non-breeding season. In seminal plasma Calcium, Sodium, Zinc and Manganese concentrations were higher (p<0.05, 0.001, 0.01 and 0.05, respectively) in the breeding season than in the non-breeding season, whereas Phosphorus, Chrome, Molybdenum and Boron concentrations were lower (p<0.001, 0.001, 0.05 and 0.001, respectively) in the breeding season than in the non-breeding season. No difference was detected regarding the other elements. The higher levels of Cr and B in the non-breeding season compared to the breeding season both in serum and seminal plasma, regardless of diet intake, suggest that these elements may play a crutial role on male fertility in Merino Ram.Öğe Antioxidative effects of cysteamine, hyaluronan and fetuin on post-thaw semen quality, DNA integrity and oxidative stress parameters in the Brown Swiss bull(Wiley, 2015) Sarıözkan, Serpil; Tuncer, Pürhan Barbaros; Büyükleblebici, Serhat; Bucak, Mustafa Numan; Cantürk, Fazile; Eken, AyşeThe aim of this study was to compare the effectiveness of antioxidants including cysteamine (2.5, 7.5mm), hyaluronan (0.25, 1mgml(-1)) and fetuin (5, 10mgml(-1)) in the freezing of Brown Swiss bull semen. The best percentages of CASA motilities were achieved with 10mgml(-1) of fetuin and 2.5mm of cysteamine. For sperm morphology, 10mgml(-1) of fetuin and 2.5mm of cysteamine had better protective effects (P<0.001). The results of hypo-osmotic swelling test showed that the percentage values of membrane integrity in all the groups, excluding that supplemented with 5mgml(-1) of fetuin, were higher than those of the control group (P<0.001). Results obtained for the DNA damage of sperm cells demonstrated that 0.25mgml(-1) of hyaluronan, and 2.5 and 7.5mm of cysteamine led to lower rates of spermatozoa with damaged DNA, compared with the control group (P<0.001). The maintenance of superoxide dismutase and glutathione peroxidase antioxidant activities following freeze-thawing with 2.5 and 7.5mm of cysteamine and 10mgml(-1) of fetuin was demonstrated to be at a higher level in comparison with the control group (P<0.001). Malondialdehyde formation was found to be lower in the groups supplemented with 0.25mgml(-1) of hyaluronan and 7.5mm of cysteamine after the freeze-thawing process (P<0.001).Öğe Combination of cysteamine and lipoic acid ımproves the post-thawed bull sperm parameters(2016) Güngör, Şükrü; Aksoy, Adil; Yeni, Deniz; Avdatek, Fatih; Öztürk, Caner; Ataman, Mehmet Bozkurt; Coyan, Kenan; Bucak, Mustafa NumanThe present study was conducted to examine the protective roles of cysteamine, trehalose, alpha-lipoic acid and combinations of these antioxidants on post-thawed bull sperm and oxidative stress parameters. Five healthy Holstein bull (3-4 years old) were used. Eight ejaculates for each bull were collected and pooled. Pooled ejaculate, splitted into seven equal aliquots and diluted at 37 °C with base extenders containing cysteamine 2 mM, trehalose 50 mM, alpha-lipoic acid (ALA) 1 mM, cysteamine 2 mM + trehalose 50 mM, ALA 1 mM + trehalose 50 mM, cysteamine 2 mM + ALA 1 mM and no antioxidant (control), was cooled to 5 °C and then frozen. Frozen straws were thawed in a water bath for evaluation. The combination of cysteamine 2 mM and ALA 1 mM of the semen extender improved the percentages of post-thawed subjective motility (68 ± 2.7%), and progressive motility (42.9 ± 4.7%), compared with the controls (61 ± 4.2% and 37.5 ± 8%, respectively, non- significantly, P>0.05). The supplementation of the semen extender with combination of cysteamine 2 mM and ALA 1 mM produced a higher acrosome integrity and mitochondrial activity (52.02 ± 6.4% and 32 ± 4.1%, respectively), compared with the controls (30.5 ± 1.7 and 14.02 ± 3.5% respectively, P < 0.05). Combination of cysteamine and ALA antioxidants in semen extenders provided the benefit in terms of sperm motilities, acrosome integrity and mitochondrial activity on frozen-thawed bull spermÖğe Combination of fetuin and trehalose in presence of low glycerol has beneficial effects on freeze-thawed ram spermatozoa(Wiley, 2021) Bucak, Mustafa Numan; Akalın, Pınar Peker; Keskin, Nazan; Bodu, Mustafa; Öztürk, Ali Erdem; İli, Pınar; Özkan, Hüseyin; Topraggaleh, Tohid Rezaei; Arslan, Halil Özancan; Başpınar, Nuri; Dursun, ŞükrüBackground: Freeze-thawing process negatively affects ram spermatozoa in terms of sperm quality, DNA integrity and antioxidant defence system. Thus, antioxidant supplementation of spermatozoa during freeze-thawing is suggested to improve sperm parameters. Objectives: The aim of this study was to determine the effects of fetuin and trehalose added into ram semen extender on sperm parameters, antioxidant parameters, antioxidant-related gene expressions and DNA integrity during the freeze-thawing process, in low glycerol concentration. Methods: Semen samples collected from six mature rams were pooled and splitted into equal aliquots and diluted with a tris-based extender containing different concentrations of glycerol (G5; %5 and G3; %3), fetuin (F; 2.5, 5 and 15 mg/mL) and trehalose (60 mm) as eight groups (G5F0, G5F2.5, G5F5, G5F15, G3F0, G3F2.5, G3F5 and G3F15). Results: G3F5 group resulted in the highest motility, mitochondrial activity and viability and the lowest DNA fragmentation and DNA damage (p < 0.05). Also, G3F0 displayed considerably more cryoprotective effect compared with G5F0 group (p < 0.05) in terms of motility, mitochondrial activity and viability rates. Lipid peroxidation levels decreased in G5F5 group compared with G5F0 group (p < 0.05). The levels of total glutathione increased in G3F2.5 group (p < 0.05) in comparison with the G5F0 group. NQO1 gene levels were upregulated approximately twofold in G5F5, G5F15, G3F2.5, G3F5 and G3F15 groups compared with G5F0 group (p < 0.05). The levels of GCLC gene were approximately twofold higher in G3F0, G3F2.5, G3F5 and G3F15 groups compared with G5F0 group (p < 0.05). GSTP1 gene levels were significantly higher with different levels in all treatment groups except for G5F2.5 and G3F0 groups in comparison with G5F0 group (p < 0.05). Conclusions: Co-supplementation of tris-based extender having low glycerol (3%) with trehalose and fetuin to enhance the quality of ram spermatozoa after freeze-thawing process is recommended.Öğe Combination of trehalose and low boron in presence of decreased glycerol improves post-thawed ram sperm parameters: A model study in boron research(Wiley, 2022) Bucak, Mustafa Numan; Keskin, Nazan; Bodu, Mustafa; Bülbül, Bülent; Kırbaş, Mesut; Öztürk, Ali Erdem; Frootan, Fateme; İli, Pınar; Özkan, Hüseyin; Başpınar, Nuri; Dursun, ŞükrüBackground: Sperm cryopreservation has been widely used in the field of reproductive biotechnology. It applies to certain males of economic and scientific values, including livestock breeds or endangered animal species. The development of a semen extender with a low cryoprotectant concentration and an appropriate amount of trehalose and boron can prevent the deterioration of sperm parameters. Objective: The main goal of this study is to establish a suitable ram extender model, by examining different combinations of high (5%) and low (3%) glycerol concentrations (to reduce its toxic effects on sperm freezing), a fixed amount of trehalose and an increased dose of boron to prevent the deterioration of sperm parameters, and investigate the levels of gene expressions. Materials and methods: The Merino ram ejaculates were collected. The collected ejaculates providing the defined criteria were pooled. The pooled ejaculates were divided into eight aliquots and diluted with the Tris extender including different combinations of glycerol (5% and 3%) and boron (0.25, 0.5, and 1 mm) concentrations and a fixed amount of trehalose, then frozen. After freeze-thawing process, sperm motility, mitochondrial membrane activity, plasma membrane integrity, acrosomal membrane integrity, DNA damage (single cell gel electrophoresis (COMET) and TUNEL assays) as well as NAD(P)H quinone oxyreductase (NQO1), glutamate-cycteine ligase (GCLC), and glutathione S-transferase (GSTP1) for molecular mechanisms of sperm cell response to oxidative stress were assessed for different extender groups following freeze-thawing process: 5% glycerol + 0 mm boron (G5B0.00), 5% glycerol + 0.25 mm boron (G5B0.25), 5% glycerol + 0.5 mm boron (G5B0.50), 5% glycerol + 1 mm boron (G5B1.00), 3% glycerol + 0 mm boron (G3B.00), 3% glycerol + 0.25 mm boron (G3B0.25), 3% glycerol + 0.5 mm boron (G3B0.50), and 3% glycerol + 1 mm boron (G3B1.00). Results: G3B0.25 presented higher percentages of subjective motility, mitochondrial activity, and viability of spermatozoa comparing with G5B0.00 and groups with boron. Supplementation of 0.25 mm boron with and without trehalose (G3B0.25 and G5B0.25) showed higher acrosome integrity, compared with G5B0.00, G5B1.00, G3B0.50, and G3B1.00. For TUNEL analysis, G3B1.00 showed the highest DNA integrity among the experimental groups which was statistically significant only with G5B0.50 (p < 0.05). The mRNA levels of NQO1 were significantly decreased in G5B1.00, G3B0.50, and G3B1.00, when compared to G5B0.00. In comparison with G5B0.00, supplementation of 1 mm boron with and without trehalose had significantly lower expression of GCLC. The level of GSTP1 gene was significantly lower (approximately threefold) in G3B1.00, compared to G5B0.00 (p < 0.05). Discussion and conclusion: It can be assumed that the increase of the boron concentration in the extender may have important adverse effects on sperm parameters and antioxidant gene expression after thawing. The results obtained from this study will help to understand the toxicity limits of boron and eliminate the toxicity of glycerol in studies of gametes and tissue freezing. Therefore, it can be concluded that the use of sufficient boron can decrease cryodamages of cryopreservation of mammalian spermatozoa as well tissue engineering.Öğe Comparing ethylene glycol with glycerol and with or without dithiothreitol and sucrose for cryopreservation of bull semen in egg-yolk containing extenders(Academic Press, 2014) Büyükleblebici, Serhat; Tuncer, Pürhan Barbaros; Bucak, Mustafa Numan; Taşdemir, Umut; Eken, Ayşe; Büyükleblebici, Olga; Durmaz, Emre; Sarıözkan, Serpil; Endirlik, Burcu ÜnlüThere are few studies performed for investigating the roles of different ratio and cryoprotectants with dithiothreitol or sucrose on sperm motility characteristics and antioxidant capacities of post-thawed bull spermatozoa. The objectives of this study were to compare glycerol (G) and ethylene glycol (EG) at different concentrations as cryoprotectants and dithiothreitol (D) or sucrose (S) (with/without) as antioxidants in Tris extender for cryopreservation of bull semen. Twenty-four ejaculates obtained from three bulls were included in the study. Each ejaculate was split into four equal aliquots and diluted using both of the Tris extenders with glycerol (5% or 7%) or ethylene glycol (3% or 5%). After that, each extenders were split into three equal aliquots and diluted using both of the dithiothreitol 5 mM or sucrose 25 mM, and control (without additives) was cooled to 4 degrees C and frozen in 0.25-ml French straws. when compared to control, different doses cryoprotectants and antioxidants addition no significantly increased the percentages of post-thaw sperm progressive and motitilities, acrosome abnormality and plasma membrane integrity (P > 0.05). However, EG3 + S yielded the greatest percentages of the total abnormality (P < 0.05). As regard to antioxidant activities G7 and EG5 led to lowest MDA activity with or without D or S but, these results were not supported to the GPx activity (P < 0.01). The sperm motion characteristics such as VAP, VCL, ALH and BCF gave significantly different results (P < 0.05). When compared the DNA integrity, different doses cryoprotectants without antioxidants addition significantly increased the percentages the tail intensity and tail moment (P < 0.05). There were no significant differences observed in non-return rates among all treatment groups (P > 0.05). (C) 2014 Elsevier Inc. All rights reserved.Öğe Cryopreservation effects on ram sperm ultrastructure(Mary Ann Liebert Inc., 2020) Keskin, Nazan; Erdoğan, Cennet; Bucak, Mustafa Numan; Öztürk, Ali Erdem; Bodu, Mustafa; İli, Pınar; Başpınar, Nuri; Dursun, ŞükrüCryoprotectants are known to have protective effects against cryodamage to spermatozoa. In this study, the cryoprotective effects of two cryoprotectants (glycerol, ethylene glycol) and cryoprotectants/trehalose combinations on frozen-thawed ram spermatozoa were investigated at the ultrastructural level. For this purpose, ejaculates collected from Konya Merino rams were pooled and diluted with a tris-based extender containing additives, including 5% glycerol, 3% glycerol +60 mM trehalose, 1.5% glycerol +100 mM trehalose, 5% ethylene glycol, 3% ethylene glycol +60 mM trehalose, and 1.5% ethylene glycol +100 mM trehalose. They were all cooled to 5°C and then frozen in 0.25 mL French straws in liquid nitrogen. The samples were thawed at 37°C and centrifuged to remove the diluents. Then, they were processed using a scanning transmission electron microscope. In the statistical analysis, the number of ultrastructurally cryodamaged and intact spermatozoa were counted in longitudinal and transverse ultrathin sections in all groups by electron microscopic examination. The amount of intact spermatozoa in the groups containing 5% ethylene glycol and 1.5% ethylene glycol +100 mM trehalose was found to be higher than other groups (p < 0.05). As a result, it was suggested that the groups of 5% ethylene glycol and 1.5% ethylene glycol +100 mM trehalose provided the highest protection for the ultrastructural morphology of frozen-thawed Konya Merino ram spermatozoa among the groups.Öğe Cryopreservation of bull sperm: Effects of extender supplemented with different cryoprotectants and antioxidants on sperm motility, antioxidant capacity and fertility results(Elsevier Science Bv., 2014) Büyükleblebici, Serhat; Tuncer, Pürhan Barbaros; Bucak, Mustafa Numan; Eken, Ayşe; Sarıözkan, Serpil; Taşdemir, Umut; Endirlik, Burcu ÜnlüThe objectives of this study were to compare glycerol (G) and ethylene glycol (EG) at different concentrations and trehalose (T) or cysteine (C; with/without) in Tris extender for cryopreservation of bull semen. Twenty-four ejaculates obtained from three bulls were included in the study. Each ejaculate was divided into four equal aliquots and diluted using both of the Tris extenders with G (5% or 7%) or EG (3% or 5%). After that, each extenders were divided into three equal aliquots and diluted using both of the 5 mM C or 25 mM T, and control (without additives) was cooled to 4 degrees C and frozen in 0.25 ml French straws. The addition of 3% and 5% EG without antioxidants resulted in the least Computer-Assisted Sperm motility Analysis (CASA) motility as compared with the other groups. Treatment with 25 mM Tin 3% EG beneficially effected acrosome morphology as compared with the other groups. Also, treatment with 3% EG with 25 mM T and 5% EG resulted in a greater rate of total abnormalities. Treatment with 3% G yielded a slightly greater percentage of membrane integrity by Hypo-Osmotic Swelling Test (HOST) assessment than that of the other groups. Treatment with 3% EG with 5 mM C resulted in the greatest concentration of malondialdehyde (MDA). The glutathione peroxidase (GPx) antioxidant activity was increased in the C-treatment groups when compared to the other groups. Treatment with 5% EG and 5 mM C resulted in less chromatin damage and detrimental impacts on tail moment. Treatment with 5% EG led to greater non-return rates of inseminated cows. However, this result was not considered to be statistically important.Öğe Decreasing glycerol content by co-supplementation of trehalose and taxifolin hydrate in ram semen extender: Microscopic, oxidative stress, and gene expression analyses(Academic Press Inc., 2020) Bucak, Mustafa Numan; Keskin, Nazan; İli, Pınar; Bodu, Mustafa; Akalın, Pınar Peker; Öztürk, Ali Erdem; Özkan, Hüseyin; Topraggaleh, Tohid Rezaei; Sarı, Fikret; Başpınar, Nuri; Dursun, ŞükrüThis study aimed to evaluate the comparative effects of taxifolin hydrate and trehalose on the quality of frozen-thawed ram spermatozoa for the first time. Ejaculates collected from six mature rams were pooled, and divided to eight equal aliquots to extend them with different concentrations of glycerol (%5 and %3), taxifolin hydrate (10, 100, and 500 ?M), and trehalose (60 mM) as eight groups (G5T0, G5T10, G5T100, G5T500, G3T0, G3T10, G3T100, and G3T500). After freeze-thawing process of cryopreservation, microscopic and oxidative stress parameters, and gene expression levels were investigated for understanding of possible impacts of taxifolin hydrate and trehalose. The study showed that G3T10 resulted in the highest post-thawed viability and mitochondrial activity. Moreover, all extenders with taxifolin hydrate reduced DNA fragmentation in comparison to G5T0, but DNA damage was prevented at the highest rate in presence of G5T10. The level of LPO significantly decreased in the groups G5T500 and G3T100, and the expression levels of NQO1, GCLC, and GSTP1 genes significantly increased in the groups G5T100, G5T500, G3T10, and G3T100 compared to the group G5T0. Finally, co-supplementation of tris-based extender having 3% glycerol with 60 mM trehalose and 10 ?M taxifolin hydrate in cryopreservation extender may be recommended to improve the quality of post-thawed ram spermatozoa. However, further in vivo and in vitro studies are suggested to evaluate fertility rates of frozen-thawed ram spermatozoa co-supplemented with trehalose and taxifolin hydrate.Öğe DNA damaging effect of paclitaxel in the epididymal sperms as a chemotherapeutic agent and possible remedies to prevent this effect: A study on reproductive potential of male cancer patients of reproductive age(Elsevier Ltd., 2019) İli, Pınar; Sarı, Fikret; Bucak, Mustafa Numan; Öztürk, Caner; Güngör, Şükrü; Ataman, Mehmet BozkurtCancer is a major public health problem, young cancer patients therefore undergo chemotherapy, andmost of them may lose their fertility. DNA damage level provides important clues about the quality andreproductive potential of spermatozoa. In this study, we evaluated the levels of both DNA fragmentationand abnormal DNA integrity in the epididymal sperms of New Zealand rabbit (Oryctolagus cuniculus)after cryopreservation using the terminal deoxyribonucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL) assay and the toluidine blue (TB) staining methods and assessed the effects of pacli-taxel, resveratrol,L-glutamine (LG), and basal medium eagle (BME) solution on DNA damage. Paclitaxelinduced the levels of both DNA damages in the sperms, but resveratrol ameliorated this effect. LG andBME supplementation to the extender prevented the sperm samples from DNA fragmentation aftercryopreservation. Chemotherapy drugs containing paclitaxel can cause the sperm DNA to be damaged,and hence adversely affect the fertility of male cancer patients of reproductive age. The administration ofresveratrol together with paclitaxel may ameliorate the DNA damage inducing effect of paclitaxel. Spermbanking and cryopreservation with the appropriate cryoprotectants such as LG and BME prior to cancertreatment can also be suggested to all male cancer patients of reproductive age facing cancer treatmentfor fertility preservation.Öğe Effects of arginine and trehalose on post-thawed bovine sperm quality(Akademiai Kiado Rt., 2017) Öztürk, Caner; Güngör, Şükrü; Ataman, Mehmet Bozkurt; Bucak, Mustafa Numan; Başpınar, Nuri; İli, Pınar; İnanc, Muhammed EnesThe present study was conducted to examine the protective role of arginine and trehalose on post-thaw bull sperm and oxidative stress parameters. Five ejaculates for each bull were used in the study. Each ejaculate, split into three equal aliquots and diluted at 37 degrees C with base extenders containing 2 mM arginine, 25 mM trehalose and no antioxidant (control) was cooled to 5 degrees C and then frozen. Frozen straws were thawed in a water bath for evaluation. Supplementation of the semen extender with arginine decreased the percentages of post-thawed subjective motility (29 +/- 8.21%), CASA motility (12.2 +/- 5.69%) and progressive motility (3.52 +/- 2.13%), compared with the controls (43 +/- 2.73%, 55.4 +/- 6.78% and 33.48 +/- 4.14%, respectively, P < 0.05). Supplementation of the semen extender with trehalose produced a higher mitochondrial activity and sperm viability (36.3 +/- 3.99% and 44.1 +/- 2.18%) compared with the control (13 +/- 8.15 and 31.7 +/- 3.94%, respectively, P < 0.05). It was established that trehalose (95.1%) and arginine (92.8%) protect DNA integrity compared to the control (90.4%) (P < 0.05). Trehalose supplementation in semen extenders provided great benefit in terms of viability, mitochondrial activity, and intact sperm DNA on frozen-thawed bull sperm.Öğe Influence of Ellagic Acid and Ebselen on Sperm and Oxidative Stress Parameters during Liquid Preservation of Ram Semen(Royan Inst, 2019) Bucak, Mustafa Numan; Bodu, Mustafa; Başpınar, Nuri; Güngör, Şükrü; İli, Pinar; Acibaeva, Begimay; Topraggaleh, Tohid Rezaei; Dursun, ŞükrüObjective: The purpose of the present study was to assess the effects of ellagic acid and ebselen on sperm and oxidative stress parameters during liquid preservation of ram semen. Materials and Methods: In this experimental study, sixty ejaculates from six mature Merino rams were used. In experiment 1, the ejaculates were diluted in base extender contained ellagic acid at 0 (control), 0.5, 1, and 2 mM. In experiment 2, ebselen at 0 (control), 10, 20, and 40 mu M were added to the extender. Sperm motility, viability, mitochondrial membrane potential, DNA integrity, lipid peroxidation (LPO), the antioxidant potential (AOP), and total glutathione (tGSH) were evaluated at 0, 24, 48, and 72 hours of preservation. Results: Supplementation of ellagic acid at 1 and 2 mM resulted in higher sperm motility and viability at 0 hours of storage. Ellagic acid at 2 mM led to higher motility and viability compared to controls after 0, 24, and 48 hours of preservation and increased AOP after 24 and 72 hours. Higher tGSH was at 1 mM ellagic acid, compared to control after 72 hours. Addition of ebselen at a concentration of 40 mu M increased motility at 24 and 48 hours and 10 mu M produced the same effect after 48 and 72 hours of storage as well as higher viability, compared to the controls after 0 hours of storage. Sperm DNA integrity was significantly improved after 24, 48, and 72 hours with the addition of ebselen at 10 mu M, and after 72 hours at 40 mu M. Addition of 40 mM ebselen also reduced the LPO levels after 24 hours of storage compared to the controls. Conclusion: The results showed that supplementation of ellagic acid and ebselen in semen extender has a potential effect on sperm and oxidative stress parameters during liquid preservation of ram semen.Öğe Influence of fetuin and hyaluronan on the post-thaw quality and fertilizing ability of Holstein bull semen(Elsevier, 2015) Sarıözkan, Serpil; Bucak, Mustafa Numan; Tuncer, Pürhan Barbaros; Büyükleblebici, Serhat; Eken, Ayşe; Akay, CemalIt was determined that fetuin and hyaluronan supplementation did not provide any significant effect on the post-thaw subjective and CASA motility percentages and sperm motion characteristics, in comparison to the controls (P > 0.05). Sperm acrosome and total abnormalities were similar in all groups (P > 0.05). Groups M (hyaluronan + fetuin) and H (hyaluronan) displayed a higher rate of sperm membrane integrity, compared to that of Group C (control) (P < 0.01). According to the results of the comet assay, the lowest percentage of sperm with damaged DNA was achieved in Group H, when compared to all of the experimental groups (P < 0.01). Furthermore, all of the additives resulted in a lower rate of sperm with damaged DNA than that of the controls, and thus, reduced DNA damage (P < 0.01). For pregnancy rates, there were no significant differences between the extender groups (P > 0.05). MDA formation was found to be lower in Groups M and F (P < 0.01). In Group M, SOD activity was determined to have significantly increased (23.61 +/- 5.62 U/ml) compared to the other groups (P < 0.01). All experimental groups had a GSH-Px activity higher than that of the control group (P < 0.01). Crown Copyright (C) 2015 Published by Elsevier Inc. All rights reserved.Öğe Influence of various antioxidants added to TCM-199 on post-thaw bovine sperm parameters, DNA integrity and fertilizing ability(Elsevier, 2014) Sarıözkan, Serpil; Bucak, Mustafa Numan; Tuncer, Pürhan B.; Büyükleblebici, Serhat; Cantürk, FazileSupplementation of the semen extender with antioxidants did not produce any significant effect on CASA and progressive motilities and sperm motility characteristics, in comparison to the control group (P > 0.05). For sperm acrosome and total abnormalities, TCM-199 supplemented with cysteine (2.60 +/- 0.24% and 4.80 +/- 0.20%), glutamine (2.80 +/- 0.20% and 6.40 +/- 0.40%), carnitine (2.60 +/- 0.24% and 6.00 +/- 0.63%) and methionine (3.40 +/- 0.51% and 9.20 +/- 0.86%) at doses of 2 mM provided a better protective effect, compared to that of the controls (8.00 +/- 0.44 and 15.60 +/- 1.895). As regards sperm membrane integrity, supplementation with 2 mM of glutamine and methionine (56.00 +/- 1.70% and 62.40 +/- 1.78%, respectively) resulted in higher rates, when compared to the control group (41.40 +/- 4.74%). According to the results of the COMET assay, only the use of TCM-199 supplemented with 2 mM of cysteine reduced DNA damage and resulted in percentages of sperm with damaged DNA (2.17 +/- 0.18%) lower than those of the control group (3.16 +/- 0.32%) (P < 0.001). For pregnancy rates, there were no significant differences among the extender groups (P > 0.05). Crown Copyright (C) 2014 Published by Elsevier Inc. All rights reservedÖğe Investigation of conception rates achieved with the transfer of sexed and unsexed bovine embryos(TÜBİTAK, 2014) Karaşahin, Tahir; Akyol, Numan; Satılmış, Muharrem; Bucak, Mustafa Numan; Çoyan, KenanThe objective of this study was to investigate the conception rates achieved with the transfer of in-vivo–derived sexed and unsexed Holstein bovine embryos to appropriate recipients and to determine the accuracy of nonelectrophoretic PCR sexing. Sevenday-old embryos were derived in vivo by the nonsurgical flushing of the uterus. Before being vitrified and frozen some of the embryos obtained were sexed, while others were not sexed and were maintained as the control group. After thawing, the sexed and unsexed embryos were transferred to 23 and 21 bovine recipients, respectively. The conception rates achieved with the transfer of the sexed and unsexed embryos were 30.4% (7/23) and 42.9% (9/21), respectively. The difference between conception rates achieved in the 2 groups was not statistically significant (P > 0.05). For the sexed embryos the conception rates achieved with the transfer of male and female embryos were 27.2% (3/11) and 33.3% (4/12), respectively. The accuracy of embryo sexing with nonelectrophoretic PCR was 66.6% for male embryos and 100% for female embryos postdelivery. The mean rate of accuracy determined for embryo sexing at the end of the study was 83.33%.Öğe The effects of different doses of ROCK inhibitor, antifreeze protein III, and boron added to semen extender on semen freezeability of Ankara bucks(MDPI, 2022) Karaşör, Ömer Faruk; Bucak, Mustafa Numan; Cenariu, Mihai; Bodu, Mustafa; Taşpınar, Mehmet; Taşpınar, FilizIn the presented study, the effects of ROCK inhibitor Y-27632, antifreeze protein III, and boron at two different doses were investigated on the spermatological parameters of Ankara buck semen after freeze–thawing. Ejaculates were collected from bucks using an electroejaculator during the breeding season. The ejaculates that showed appropriate characteristics were pooled and used in the dilution and freezing of semen. The extender groups were formed by adding two different doses of three different additives (ROCK inhibitor Y-27632, 5 and 20 µM; antifreeze protein III, 1 and 4 µg/mL; boron, 0.25 and 1 mM) to the control extender. The semen was diluted with the different extenders at 35–37 °C and loaded into straws. Sperm samples frozen in liquid nitrogen vapors, following equilibration, were stored in liquid nitrogen. It was observed that extender supplementation improved post-thaw motility of Ankara buck semen after freeze–thawing. Differences were significant (p < 0.01) for 5 and 10 µM doses of ROCK inhibitor (71.82% and 74.04 % motility), as well as for 0.25 and 1 mM doses of boron (76.36% and 72.08% motility), compared to the control group (66.15% motility). With respect to the evaluation of acrosomal integrity and mitochondrial activity after freeze–thawing, although supplementation provided protection at all doses, the efficacy was not statistically significant (p > 0.05). It was observed that DNA damage was improved by antifreeze protein III at 1 µg/mL (1.23% ± 0.23%) and by boron at all doses (0.25 mM: 1.83% and 1 mM: 1.18%) compared to the control group (3.37%) (p < 0.01), following the thawing process. In the present study, it was determined that some additives added to the extender provided significant improvements in buck spermatozoa motility and DNA damage after thawing.Öğe The synergistic effect of trehalose and low concentrations of cryoprotectants can improve post-thaw ram sperm parameters(Academic Press Inc., 2020) Öztürk, Ali Erdem; Bodu, Mustafa; Bucak, Mustafa Numan; Ağır, Vahit; Özcan, Ayşe; Keskin, Nazan; İli, Pınar; Topraggaleh, Tohid Rezaei; Sidal, Hümeyra; Başpınar, Nuri; Dursun, ŞükrüThe objective of this study was to compare the effects of different concentrations of two different cryoprotectants (glycerol, G and ethylene glycol, EG) and trehalose (T), added to the semen extender, on post-thaw ram sperm parameters. Ejaculates, collected from 6 Merino rams, were pooled and evaluated at 37 °C. The pooled samples were divided into six equal aliquots, and diluted in Tris-based extenders containing 5% G, 3% G + 60 Mm T, 1.5% G + 100 Mm T, 5% EG, 3% EG + 60 mM T, and 1.5% EG + 100 Mm T. Subsequently, the samples were cooled to 5 °C, frozen in 0.25-ml French straws, and stored in liquid nitrogen (LN2). Frozen samples were thawed individually, at 37 °C for 25 s in a water bath, for evaluation. Sperm motility was assessed using a phase-contrast microscope with a warm stage maintained at 37 °C. Acrosome integrity (FITC/PNA-PI), sperm viability (SYBR-14/PI), mitochondrial activity (JC-1/PI), DNA damage (COMET assay) and DNA fragmentation (TUNEL test) were determined. The group of samples diluted in an extender containing 5% of glycerol (Group 5% G) displayed higher percentages of subjective motility, viability and mitochondrial activity of sperm, compared to the other groups (P < 0.05). On the other hand, Group 3% G + 60 mM T yielded the second-best results for subjective motility, viability and mitochondrial activity of sperm, when compared to the other groups. The post-thaw sperm parameters of Group 3% G + 60 Mm T did not show any statistically significant difference from those of Group 5% G. There were no statistically significant differences between the groups for acrosome integrity (P > 0.05). The results of the COMET assay showed that the use of low concentrations of cryoprotectants in combination with trehalose decreased sperm DNA damage. Accordingly, Group 1.5% G + 100 mM T and Group 3% EG + 60 mM T benefited from a significantly stronger cryoprotective effect on DNA integrity, in comparison to Group 5% G (P < 0.05). According to the results of the TUNEL test, the combined use of low concentrations of cryoprotectants with trehalose decreased sperm DNA damage, when compared to the use of 5% glycerol, but this difference was statistically insignificant (P > 0.05). In conclusion, G and EG concentrations can be reduced by adding various amounts of T (60 mM, 100 mM) to the semen extender. The addition of 5% of glycerol and 3% G + 60 mM T to the semen extender did not yield statistically different post-thaw sperm parameters, when compared for protection against cryoinjury. Post-thaw sperm parameters can be improved by the supplementation of the semen extender with 3% G + 60 mM T. Thus, we recommend the use of freezing extenders containing low cryoprotectant concentrations (3% G) combined with trehalose to avoid the high level of toxic and osmotic damage caused by 5% G.
