The effects of antioxidant supplementation during Percoll preparation on human sperm DNA integrity
ABSTRACT The integrity of sperm DNA is crucial for the maintenance of genetic health. A major source of damage is reactive oxygen species (ROS) generation; therefore, antioxidants may afford protection to sperm DNA. The objectives of the study were, first, to measure the effects of antioxidant supplementation in vitro on endogenous DNA damage in spermatozoa using the single cell gel electrophoresis (comet) assay and, second, to assess the effect of antioxidant supplementation given prior to X-ray irradiation on induced DNA damage. Spermatozoa from 150 patients were prepared by Percoll centrifugation in the presence of ascorbic acid (300, 600 microM), alpha tocopherol (30, 60 microM), urate (200, 400 microM), or acetyl cysteine (5, 10 microM). DNA damage was induced by 30 Gy X-irradiation. DNA strand breakage was measured using the comet assay. Sperm DNA was protected from DNA damage by ascorbic acid (600 microM), alpha tocopherol (30 and 60 microM) and urate (400 microM). These antioxidants provided protection from subsequent DNA damage by X-ray irradiation. In contrast, acetyl cysteine or ascorbate and alpha tocopherol together induced further DNA damage. Supplementation in vitro with the antioxidants ascorbate, urate and alpha tocopherol separately has beneficial effects for sperm DNA integrity.
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ABSTRACT: The present study was aimed to test the efficacy of adding vitamins C or E to Tris-fructose-egg yolk diluent to increase Awassi ram sperm storage period at 5 ˚C. Semen samples from six mature Awassi rams were used in this study. The semen samples were diluted by Tris-glucose-egg yolk. Diluted semen sample was divided into three parts. The first part was added with 0.9 mg mL(-1) vitamin C, the second part was added with 1 mg mL(-1) vitamin E and the third part was considered as a control without any addition. The diluted semen samples were cooled gradually and preserved at 5 ˚C for five days. Sperms in cooled diluted semen samples were examined for motility, vitality, abnormalities and acrosomal defects every 24 hr for five days. Results of the present study showed an increase in the viability of spermatozoa diluted in the Tris diluent containing vitamins C or E stored at 5 ˚C for 120 hr compared with the control group. There were significant (p < 0.05) effects of vitamins C and E addition to semen diluents on sperm motility as well as the sperm viability in different times of preservation at 5 ˚C. Significant (p < 0.05) higher sperm abnormalities and acrosomal defects values (37.6 ± 1.3% and 71.5 ± 1.1%, respectively) were found after 120 hr incubation in Tris free vitamin C (Control) at 5 ˚C compared with those of containing vitamin C (18.8 ± 1.8% and 52.8 ± 4.3%, respectively). From the results of the present study, it could be concluded, that the addition of antioxidants such as vitamins C and vitamin E to semen preservation media could improve longevity and quality of cooled sperm in Awassi ram semen.
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ABSTRACT: Excess of reactive oxygen species (ROS) on in vitro embryo production systems negatively affects the quality and developmental potential of embryos, as result of a decreased sperm quality and increased DNA fragmentation. This issue is of major importance in assisted fertilisation procedures such as intracytoplasmic sperm injection (ICSI), because this technique does not allow the natural selection of competent spermatozoa, and therefore, DNA-damaged spermatozoa might be used to fertilise an egg. The aim of this study was to investigate a new strategy to prevent the potential deleterious effect of ROS on cryopreserved bovine spermatozoa. We evaluated the effect of a sperm pre-treatment with different concentrations of N-acetyl-L-cysteine (NAC) on ROS production, viability and DNA fragmentation and assessed the effect of this treatment on the in vitro developmental potential and quality of embryos generated by ICSI. The results show a strong scavenging effect of 1 and 10 mm NAC after exposure of spermatozoa to a ROS inducer, without compromising the viability and DNA integrity. Importantly, in vitro developmental potential and quality of embryos generated by ICSI with spermatozoa treated with NAC were not affected, confirming the feasibility of using this treatment before an ICSI cycle. © 2015 Blackwell Verlag GmbH.Andrologia 03/2015; DOI:10.1111/and.12412 · 1.55 Impact Factor
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ABSTRACT: This study was designed to investigate enzymatic antioxidants' activity and nonenzymatic antioxidants' levels in seminal plasma of stallions and to relate them with season, age, and fertility of stallions. Fifty ejaculates were collected from six healthy Arabian stallions, 4-22 years old. Ejaculates were evaluated by conventional methods. Five milliliters of each semen sample was centrifuged, and the supernatant seminal plasma was stored at -20 degrees C. Five antioxidants, in addition to osteopontin (OPN) and testosterone, were determined in stallion seminal plasma by using commercial enzyme-linked immunosorbent assay kits. Results revealed that uric acid, ascorbic acid, OPN, and testosterone concentrations and glutathione peroxidase (GPx) activity in stallions' seminal plasma were high (P<.05) during spring. GPx activity was higher (P<.05) in age group B (11-18 years old) than in age group A(4-10 years old). The effect of stallions' age on GPx activity in the fertility groups was highly significant (P<.01). OPN concentration was highest (P<.05) in age group A. Uric acid and OPN concentrations and GPx activity in stallions' seminal plasma and percent of sperm motility were higher (P<.05) in fertility group III (>70%) than in fertility group I (<50%). However, ascorbic acid concentration, catalase activity and percentage of sperm abnormalities were lower (P<.05) in fertility group III than in fertility group I. It was concluded that season and stallion age may affect antioxidant defense systems in stallions' seminal plasma. The impairment of seminal antioxidants and OPN could lead to low fertility.Journal of Equine Veterinary Science 09/2013; 33(9-9):705-709. DOI:10.1016/j.jevs.2012.11.006 · 0.89 Impact Factor