1- Azarbaijan Shahid Madani University
2- university of kurdistan
3- university of tehran
2- university of kurdistan
3- university of tehran
Abstract: (8 Views)
Extended abstract
Introduction and purpose: The progress of breeding programs and improvement of the reproductive status of livestock in the last few decades is the result of the use of artificial insemination technique. In heavy livestock, this technique is widely used, but in light livestock, limitations such as the complex anatomical system of the female animal and the very high sensitivity of ram sperm to cold shock and oxidative stress have often made the use of artificial insemination difficult. Given the importance of the quality of the produced sperm in sheep artificial insemination, the sperm used is mostly fresh and chilled semen. However, it is not always possible to use fresh semen and sperm preservation is inevitably necessary. Therefore, sperm cooling can be used in sheep artificial insemination in the short term with minimal damage to sperm and also have the necessary fertility. However, during the cooling process, due to the decrease in temperature and the occurrence of cold shock, the production of free radicals increases and causes numerous structural and biochemical damages to sperm and reduces its fertility chances. To combat this problem, it is crucial to use the right antioxidant compound in the diluent. Gamma oryzanol is a natural antioxidant compound derived from barley, whose antioxidant properties have been proven in many studies. The antioxidant properties of gamma oryzanol are primarily related to the scavenging of free radicals, especially superoxide, hydroxyl, and hydrogen peroxide radicals, with the hydroxyl group on the phenolic ring of the ferulic acid moiety being the main inhibitor of free radicals. Gamma oryzanol acts as an activator of intracellular antioxidant enzymes such as superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase, increasing their activity. It has also been shown that gamma oryzanol can interfere with the Nrf-2 biochemical pathway and increase the expression of proteins related to some antioxidant enzymes such as superoxide dismutase, glutathione peroxidase, and catalase, and increase their production. Therefore, the aim of this study was to investigate the effects of adding different concentrations of gamma oryzanol to ram epididymal sperm diluent at different incubation times at 4°C.
Materials and methods: In this experiment, testicular tissue was transported to the laboratory within one hour after collection from the slaughterhouse. Sperm preparation was performed by cutting the epididymal tail. After initial evaluation, sperm samples with viability above 85%, progressive motility above 75%, and abnormalities below 10% were selected and diluted in Tris-egg yolk-based diluent. At 37°C, different concentrations of gamma oryzanol including 0, 50, 100, and 200 μM were added to the diluent containing sperm. Subsequently, the samples were transferred to 4°C in isothermal water for cooling. After about two hours, the samples were gradually cooled to 4°C and equilibrated. At 6, 12, 24, and 48 hours after fixation, the samples were evaluated at this temperature for the following parameters: total motility, progressive motility, viability, plasma membrane integrity, percentage of abnormalities, malondialdehyde (MDA) concentration, and antioxidant enzyme activities of superoxide dismutase (SOD), glutathione peroxidase (GPX), and catalase (CAT). The data obtained from the evaluations were analyzed by SAS software and the GLM procedure at a significance level of 0.05.
Results: The results of this experiment showed that the diluent containing 100μM gamma oryzanol significantly improved the overall motility characteristic compared to the control group at 12, 24 and 48 hours (P<0.05), but no significant difference was observed between the diluents in the progressive motility characteristic at all evaluated times (P>0.05). In examining the percentage of sperm viability, only at 12 hours the diluent containing 100μM gamma oryzanol significantly increased sperm viability, but at other times there was no significant difference between the groups. Regarding the sperm plasma membrane integrity parameter, the addition of 100μM gamma oryzanol had a significant effect compared to the control group, and regarding sperm abnormalities, no significant difference was found between the groups at any time. The results of the present experiment showed that the addition of 50 and 100 μM concentrations of gamma oryzanol significantly reduced MDA concentration at 12 and 24 hours. Also, the 100μM concentration of this compound significantly increased SOD enzyme activity at 12 and 24 hours, and the 100 and 200μM concentrations significantly increased GPX enzyme activity only at 24 hours (P<0.05), but the addition of gamma oryzanol concentrations did not significantly increase CAT enzyme activity compared to the control group at any of the evaluated times (P>0.05).
Conclusion: The results of this experiment showed that the use of the antioxidant compound gamma oryzanol in the diluent medium can improve the quality of ram epididymal sperm to some extent in short periods after refrigeration. Therefore, the use of a concentration of 100 μM gamma oryzanol in ram sperm diluent is recommended for short-term refrigeration.
Introduction and purpose: The progress of breeding programs and improvement of the reproductive status of livestock in the last few decades is the result of the use of artificial insemination technique. In heavy livestock, this technique is widely used, but in light livestock, limitations such as the complex anatomical system of the female animal and the very high sensitivity of ram sperm to cold shock and oxidative stress have often made the use of artificial insemination difficult. Given the importance of the quality of the produced sperm in sheep artificial insemination, the sperm used is mostly fresh and chilled semen. However, it is not always possible to use fresh semen and sperm preservation is inevitably necessary. Therefore, sperm cooling can be used in sheep artificial insemination in the short term with minimal damage to sperm and also have the necessary fertility. However, during the cooling process, due to the decrease in temperature and the occurrence of cold shock, the production of free radicals increases and causes numerous structural and biochemical damages to sperm and reduces its fertility chances. To combat this problem, it is crucial to use the right antioxidant compound in the diluent. Gamma oryzanol is a natural antioxidant compound derived from barley, whose antioxidant properties have been proven in many studies. The antioxidant properties of gamma oryzanol are primarily related to the scavenging of free radicals, especially superoxide, hydroxyl, and hydrogen peroxide radicals, with the hydroxyl group on the phenolic ring of the ferulic acid moiety being the main inhibitor of free radicals. Gamma oryzanol acts as an activator of intracellular antioxidant enzymes such as superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase, increasing their activity. It has also been shown that gamma oryzanol can interfere with the Nrf-2 biochemical pathway and increase the expression of proteins related to some antioxidant enzymes such as superoxide dismutase, glutathione peroxidase, and catalase, and increase their production. Therefore, the aim of this study was to investigate the effects of adding different concentrations of gamma oryzanol to ram epididymal sperm diluent at different incubation times at 4°C.
Materials and methods: In this experiment, testicular tissue was transported to the laboratory within one hour after collection from the slaughterhouse. Sperm preparation was performed by cutting the epididymal tail. After initial evaluation, sperm samples with viability above 85%, progressive motility above 75%, and abnormalities below 10% were selected and diluted in Tris-egg yolk-based diluent. At 37°C, different concentrations of gamma oryzanol including 0, 50, 100, and 200 μM were added to the diluent containing sperm. Subsequently, the samples were transferred to 4°C in isothermal water for cooling. After about two hours, the samples were gradually cooled to 4°C and equilibrated. At 6, 12, 24, and 48 hours after fixation, the samples were evaluated at this temperature for the following parameters: total motility, progressive motility, viability, plasma membrane integrity, percentage of abnormalities, malondialdehyde (MDA) concentration, and antioxidant enzyme activities of superoxide dismutase (SOD), glutathione peroxidase (GPX), and catalase (CAT). The data obtained from the evaluations were analyzed by SAS software and the GLM procedure at a significance level of 0.05.
Results: The results of this experiment showed that the diluent containing 100μM gamma oryzanol significantly improved the overall motility characteristic compared to the control group at 12, 24 and 48 hours (P<0.05), but no significant difference was observed between the diluents in the progressive motility characteristic at all evaluated times (P>0.05). In examining the percentage of sperm viability, only at 12 hours the diluent containing 100μM gamma oryzanol significantly increased sperm viability, but at other times there was no significant difference between the groups. Regarding the sperm plasma membrane integrity parameter, the addition of 100μM gamma oryzanol had a significant effect compared to the control group, and regarding sperm abnormalities, no significant difference was found between the groups at any time. The results of the present experiment showed that the addition of 50 and 100 μM concentrations of gamma oryzanol significantly reduced MDA concentration at 12 and 24 hours. Also, the 100μM concentration of this compound significantly increased SOD enzyme activity at 12 and 24 hours, and the 100 and 200μM concentrations significantly increased GPX enzyme activity only at 24 hours (P<0.05), but the addition of gamma oryzanol concentrations did not significantly increase CAT enzyme activity compared to the control group at any of the evaluated times (P>0.05).
Conclusion: The results of this experiment showed that the use of the antioxidant compound gamma oryzanol in the diluent medium can improve the quality of ram epididymal sperm to some extent in short periods after refrigeration. Therefore, the use of a concentration of 100 μM gamma oryzanol in ram sperm diluent is recommended for short-term refrigeration.
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