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    IMPACTS OF SPECIFIC CRYOPROTECTANTS ON SPERM FREEZING AND RELATIONSHIPS BETWEEN CRYODAMAGE AND OXIDATION STRESS PARAMETERS IN AWASSI RAM SPERM
    (Cryo Letters, 2021) Varisli, Omer; Erat, Serkan; Bozkaya, Faruk; Aydilek, Nurettin; Taskin, Abdullah
    BACKGROUND: The role of oxidative stress during cryoprotectant treatment has received little attention. OBJECTIVE: To assess the effects of different cryoprotectants and discover relationships between cryodamage and oxidative stress parameters on Awassi ram sperm. MATERIALS AND METHODS: The sperm samples diluted with Salamon's tris-citrate (TRIS) containing 20% centrifuged egg yolk and 0.5, 1.0 or 1.5 M Glycerol (Gly), methanol (M), 2-methoxyethanol (2-ME), dimethylacetamide (DMA) and 1.2 propanediol (PR). After 2 h of equilibration at +4 degrees C, the sperm samples were frozen in liquid nitrogen vapour and stored. RESULTS: The best post-thaw motility (43.3%, 41.7%) of sperm was achieved when protected with 0.5 and 1.0 M glycerol. Arylesterase and ceruloplasmin parameters were significantly different after equilibration, whereas sulfhydryl groups were significantly different after freezing in their respective groups (P< 0.05). CONCLUSION: The increased use of glycerol caused greater loss of motility. The role of oxidative stress in freezing was also found to be limited.
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    Relationship Between Toxicity of Cryoprotectants, Osmotic and Oxidative Stresses In Awassi Ram Sperm
    (Cryo Letters, 2022) Varışlı, Ömer; Bozkaya, Faruk; Aydilek, Nurettin; Taşkın, Abdullah
    BACKGROUND: The relationship between the toxicity of cryoprotectants and their osmotic and/or oxidative stresses remains to be further investigated. OBJECTIVE: To investigate the toxic effects of different cryoprotectants and osmotic stress on Awassi ram sperm and to determine the relationship between oxidative and antioxidative status of the sperm. MATERIALS AND METHODS: Pooled sperm samples were exposed to sucrose solutions of different concentrations (75 to 900 mOsm) and isosmotic condition (290-325 mOsm) was re-established by adding HEPES buffered Tyrode's lactate. Sperm samples were mixed with 0.5, 1.0 and 1.5 M of glycerol, methanol, 2-methoxyethanol, dimethylacetamide or 1,2-propanediol for 5 min and returned to isosmotic condition. Sperm samples were exposed to cryoprotectants at 4 degrees C for 2 hours and isosmotic conditions were re-established. Motility, viability, acrosome integrity and oxidative or antioxidative parameters were determined. RESULTS: Treatment with hypo- or hyperosmotic sucrose solution reduced motility and viability without affecting acrosome integrity. The addition and removal of glycerol and dimethylacetamide (1.0 or 1.5 M) decreased sperm motility, while cryoprotectants had no effect on viability except for 1.5 M glycerol. Chilling significantly reduced the motility and viability of the sperm, but not the acrosome integrity. Rapid addition or removal of cryoprotectants also did not affect the acrosome integrity. Cryoprotectants changed only the ceruloplasmin level, while there were significant post-chilling differences in lipid hydroperoxide, paraoxonase and ceruloplasmin levels. CONCLUSION: Cryoprotectants without other additives have limited protection and glycerol can be toxic to spermatozoa. The oxidative stress plays a role in cryoprotectant toxicity and chilling stress.