Koç Spermasının Dondurulmasında, İnkubasyon Direncinin Çeşitli Antioksidanlar ile Artırılması

Year 2017, Volume: 14 Issue: 3, 183 - 190, 15.12.2017

Selim Alçay Mehmed Berk Toker Elif Gökçe Zülfiye Gül Nail Tekin Önder Burcu Üstüner Zekariya Nur Hakan Sağırkaya Mustafa Kemal Soylu

Abstract

Çalışmamızda farklı antioksidanların dondurma çözdürme sonrası sperma parametreleri ve spermanın inkubasy-on direncinin artırılması üzerine etkilerinin belirlenmesi amaçlanmıştır. Birleştirilien sperma örnekleri dört eşit hacme bölünerek, antioksidan ilave edilmiş (5mM methiyonin, 5mM sisteamin, 1mM sistein) ve edilmemiş (kontrol) sulandırıcılarla iki aşamalı sulandırma yöntemi ile sulandırılmıştır. Sperma örneklerinin değerlendirilmesi amacıyla mo-tilite, plazma membran bütünlüğünün değerlendirilmesi amacıyla hipoozmotik şişme testi (HOST) ve Hoechst 33258 testi, akrozom ve DNA bozukluklarının belirlenmesi amacıyla ise sırasıyla FITC-Pisum sativum agglutinin (PSA-FITC) ve terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) boyama değerlerine bakılmıştır. Ayrıca membran lipid peroksidasyonun değerlendirilmesi için malondialdehyde (MDA) konsantrasyon ölçümü yapılmıştır. Değerlendirmeler doğrultusunda koç spermasının dondrulmasında antioksidan ilave edilen grupların mo-tilite, plazma mebran bütünlüğü ve akrozom bütünlü değerlerinin kontrol grubundan daha yüksek olduğu sonucuna varılmıştır (P<0.05). Sistein ilave edilen grubun motilite (%60.40) ve akrozomal bütünlük (%70) oranları diğer gruplar-dan daha yüksek buunmuştur.Ancak inkubasyonun 0. Saatinde grupların DNA bütünlük oranları arasında bir fark bulunmamaktadır (P>0.05). Dondurma çözdürme işlemleri ve 6 saat inkubasyon sonrası en yüksek motilite (%35), HOST (%50), Hoechst (%56.50) ve en düşük akrozomal bozukluk (%35) ve DNA bozukluk (%8.80) oranları 1mM sistein içeren sulandırıcı ile dondurulan grupta elde edilmiştir.

Keywords

Antioksidan, inkubasyon direnci, koç sperması, sperma dondurma

Improvement of Incubation Resilience with Various Antioxidants in Cryopreserved Ram Semen*

Abstract

The aim of the current study was to evaluate different antioxidant-supplemented extenders for post-thaw semen quality and incubation resilience of ram spermatozoa. Pooled semen samples were divided into four equal vol-umes and each volume were diluted with two-step dilution method in control and antoxidant supplemented groups (5mM methionine, 5mM cysteamine and 1mM cysteine). Semen samples were assessed for the sperm motility, plasma membrane integrity using hypoosmotic swelling test (HOST) and Hoechst 33258 test, damaged acrosome using FITC-Pisum sativum agglutinin (PSA-FITC) and DNA integrity using terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL). Membrane lipid peroxidation was also analyzed using the malondialdehyde (MDA) concen-tration assay. This study showed that antioxidant supplemented extenders had beneficial effect on motility, plasma membrane integrity and acrosome integrity of ram semen compared to control group (P<0.05). The motility (60.40%) and acrosome integrity (30%) values of cysteine group was significantly higher than those of the groups (P<0.05). However, there were no differences about DNA fragmentation rates at 0 h of incubation. In addition, we achieved a higher motility (35%), HOST (50%), Hoechst (56.50%) and lower defected acrosome (35%) and DNA fragmentation (8.80%) rates in post thawed ram semen even after 6 h incubation when the extender was supplemented with 1 mM cysteine.

Keywords

Antioxidants, cryopreservation, incubation resilience, ram semen

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