ارزیابی تاثیر اضافه کردن نانوسلنیوم به رقیق کننده منی بر روی کیفیت اسپرم قوچ پس از فرآیند انجماد-ذوب کردن

زمینه مطالعاتی: آنتی‌اکسیدان‌ها جهت حفظ سلامت سلول‌ اسپرم در مقابل آسیب‌های ناشی از تنش اکسیداتیو ضروری هستند. هدف: این پژوهش به منظور افزایش خصوصیات کیفی منی قوچ توسط نانوسلنیوم طی فرآیند انجماد در نیتروژن مایع طی 30 روز نگهداری انجام پذیرفت. روش کار: جمع‌آوری منی دو بار در هفته از 4 راس قوچ نژاد قزل خالص که از هر قوچ 6 بار اسپرم‌گیری صورت گرفت. منی جمع‌آوری شده با رقیق‌کننده‌ی بدون آنتی‌اکسیدان (شاهد) و نانوسلنیوم 1 میکروگرم و نانوسلنیوم 2 میکروگرم در میلی‌لیتر رقیق شد. وپایوت‌ها پس از سردسازی به مدت 90 دقیقه در یخچال و رسیدن به دمای5 درجه‌ی سانتیگراد، به مدت 8 الی 10 دقیقه در 4 سانتی‌متری بالای سطح نیتروژن مایع قرار گرفتند و سپس در داخل نیتروژن مایع غوطه‌ور گردیدند. خصوصیات کیفی اسپرم‌ها شامل زندهمانی، درصد تحرک کل و پیش‌رونده، سلامت غشای پلاسمایی، درصد اسپرم‌های غیرطبیعی و غشای آکروزوم ناسالم در روزهای صفر و 15 و 30 فرآیند انجماد مورد بررسی قرار گرفتند. نتایج: درصد زنده‌مانی، تحرک کل و پیش‌رونده و سلامت غشای سیتوپلاسمی اسپرم‌ها در اثر گذر زمان کاهش معنی‌داری داشت (0.01>p). همچنین درصد اسپرمهای غیر‌طبیعی و اسپرم‌های با غشای آکروزوم ناسالم در طی آزمایش روند افزایشی داشتند (0.01>p). با این وجود، افزودن سطوح مختلف نانوسلنیوم به رقیق کننده سبب افزایش معنی‌دار درصد زنده‌مانی، درصد اسپرم‌های با حرکت کل و پیش‌رونده و غشای پلاسمایی سالم(هاست مثبت) در روزهای صفر و 15و 30 نسبت به تیمار شاهد شده (0.01>p) و سبب کاهش درصد اسپرم‌های با غشای پلاسمایی ناسالم و درصد اسپرم‌های غیرطبیعی نسبت به تیمار شاهد شد (0.01>p). نتیجه گیری نهایی: با توجه به نتایج حاصل از این پژوهش، افزودن مقادیر مختلف نانوسلنیوم خصوصا نانوسلنیوم 1 میکروگرم در میلی‌لیتر به رقیق‌‌کننده‌ی اسپرم قوچ سبب افزایش فراسنجه‌های کیفی اسپرم در طول مدت نگهداری می‌شود

Investigating the effect of interaction of Nano selenium as antioxidant with time on qualitative parameters of Ghezel ram sperm in different thawing days

Abstract Introduction: Freezing is one branch of cryobiology knowledge that process to protecting and long term keeping of cell in very low temperature. Sperm freezing commonly done in infertility treatment centers and keeping sperm banks and livestock industry. Sperm can survive decrease out of body and environment temperature. freezing is so important one of influential destructive factors in reducing duration of sperm storage is creation of ROS. It needs very low, but it is increasing has destructive effects on sperm (lenzi et al. 1994). Use of antioxidants in sperm extender have better results to deal with these reactive oxygen species. It is believed that ROS induces lipid peroxidation in the sperm membrane and The peroxidation of the resulting fatty acids has toxic effects on the sperm that Leading to a decrease in sperm function (Sanuka and Corps 2004). Antioxidants are compounds that control, stop and neutralize intermediates derived from the energy production process in the electron transport chain of cells (aerobic activity) known as active oxygen species, and in a group of free radicals (Zinee et al. 2000). Adding antioxidants to semen diluent during freezing of sperm improves the quality and improvement of semen parameters such as motility, viability and acrosome membrane integrity (Mironoksuk et al. 2018). Selenium is an essential micronutrient element to human health Which is toxic at high concentrations. Selenium is a constituent of selenoproteins that they have an enzymatic and structural role in biochemistry. Selenium is the main component of selenoenzymes in the center of all of these proteins, there is the amino acid of selenocysteine Which acts as an oxidationreducing agent. However, one of the limiting factors for selenium consumption is bioavailability (the rate of entry into circulation and tissues) and its toxicity (Tarz et al. 2007). Therefore, the use of other forms of selenium that is less toxic and more beneficial is more appropriate. Selenium nanoparticles are low toxicity compounds and bioavailable and effectively increase the production of selenoproteins in the body (Peng et al. 2007). Selenium is a cofactor or activator of the glutathione peroxidase enzyme Which is one of the strongest antioxidants in the body and catalyzes the decomposition of lipid peroxides and hydrogen peroxide (Hill et al. 1996). The glutathione peroxidase enzyme destroys free radicals within the cytoplasm (Ozball et al. 2008). Typically, the concentration of most of the elements in the liver is higher than the other organs, but in the case of selenium, it has the highest concentration in the testis. High concentration of selenium in testis indicates the protective role of this element and its related enzymes in spermatogenesis (Schumberger et al. 1983). Material and method: Semen was collected with an artificial vagina twice a week from 4 selected adult Ghezel rams during nonreproductive season. Ejaculates were immediately evaluated primarily for parameters including volume, concentration, motility, viability and abnormal sperm. And if they had the required standards (high concentration of 3 billion sperm per ml, motility above 70% and volume greater than 0.5 ml), they were selected as suitable samples for dilution. The semen was diluted with extender with no antioxidants (control) and containing 1 µg/ml Nano Selenium and 2 µg/ml Nano Selenium and then immediately aspirated to 0.25 ml freezing straw. After recording, treatments were packaged and refrigerated in 5°C for 1.5 hours. Then, the packaged straw containing semen were placed on 4 cm above the liquid nitrogen, then were immersed in liquid nitrogen immediately after 10 minutes. Qualitative parameters of sperm included viability, total motility (TM), progressive motility (PM), plasma membrane integrity (HOST), percentage of abnormal sperm and acrosome membrane damaged was studied in days 0, 15 and 30 of freezing process. Results and Discussion: Percentage of viability, total motility, progressive motility, plasma membrane integrity (HOST) of sperm decreased significantly (p < 0.01). Also the percentage of acrosome membrane damage and abnormal sperm increased during experiment (p < 0.01). However, the addition of different levels of Nano Selenium to semen extender significantly increased the percentage of viability, percentage of sperm with total motility, progressive motility and health plasma membrane (positive HOST) on days 0, 15 and 30 compared with control group (p < 0.01). Also, the addition of different levels of Nano Selenium to extender significantly decrease the percentage of abnormal sperm and acrosome membrane damaged (p < 0.01). Conclusion: According to the results of this study, the addition of different levels of Nano Selenium, especially Nano Selenium 1 µg/ml to ram sperm extender increases the quality of sperm during storage.