Koç spermasının dondurulmasına alfa lipoik asit ve trehalozun etkisi

Öz

Bu çalışmada, koç sperma sulandırıcısına farklı dozlardaki Alfa Lipoik asit (ALA) ve Trehalozun eklenerek dondurulup çözdürülmesinden sonra spermatozoon motilitesi, plazma membran ve akrozom bütünlüğü (PMAI), mitokondrial membran potansiyeli ve mitokondrial reaktif oksijen türleri üzerine etkilerinin araştırılması amaçlanmıştır. 10 adet Pırlak koçtan suni vajen kullanılarak alınan spermalar. üç eşit parçaya bölünerek Alfa lipoik asit (1 mM), Trehaloz (100 mM) içeren ve hiçbir katkı maddesi içermeyen (kontrol) sulandırıcılar ile 5x108/ml spermatozoa olacak şekilde sulandırıldı. Spermalar + 4 ° C'de yaklaşık 2 saat ekibrasyona bırakıldı, sıvı azot buharında (-120 C⁰) donduruldu, 37 C⁰’de 30 saniyede çözdürüldü. Çözdürülen sperma örneklerinde motilite, 37 C⁰'de ısıtma tablalı faz kontrast mikroskop ile subjektif olarak, PMAI, mitokondrial membran potansiyeli ve mitokondrial reaktif oksijen türleri (MitoSOX+) flow sitometre ile değerlendirildi. En düşük motilite kontrol grubunda tespit edilirken en yüksek plazma membran akrozom bütünlüğü Trehaloz ve ALA gruplarında belirlendi (P<0,05). En düşük Hmmp ve en yüksek Lmmp değeri kontrol grubunda tespit edilirken en düşük MitoSOX+ Trehaloz grubunda belirlendi (P<0,05). Sonuç olarak; Pırlak koçlarda sezon dışı permanın dondurulmasında ALA ve Tehalozun spermatolojik parametreleri iyileştirdiği belirlenmiştir.

Anahtar Kelimeler

• Alfa Lipoik Asit
• trehaloz
• dondurma
• çözdürme
• sperma
• flow cytometry

Effect of alpha lipoic acid and trehalose on ram semen cryopreservation

Abstract

In this study, it was aimed to investigate the effects on spermatozoon motility, plasma membrane and acrosome integrity (PMAI), mitochondrial membrane potential and mitochondrial reactive oxygen species after freezing and thawing by adding different doses of A Lipoic acid (ALA) and Trehalose to ram semen extender. Semen was collected from 10 Pirlak rams using artificial vagina, divided into three equal parts, extended with extender containing A lipoic acid (1 mM), Trehalose (100 mM) and no additives (control) to contain 5x108/ ml spermatozoa. Sperm was equilibrated about 2 hours at + 4 ° C, frozen in liquid nitrogen vapor (-120 ° C), thawed in 30 seconds at 37 ° C. Thawed sperm sample were evaluated. Subjective motility evaluated by phase contrast microscope with heating table at 37 °C, PMAI, mitochondrial membrane potential and mitochondrial reactive oxygen species were evaluated by flow cytometer. While the lowest motility was detected in the control group, the highest plasma membrane integrity was found in the Trehalose group (P <0.05). While the lowest Hmmp and the highest Lmmp value were detected in the control group, the lowest MitoSOX+ was detected in Trehalose group. It has been determined that ALA and Trehalose improve spermatological parameters in Pirlak ram semen after freezing and thawing procedure on non-breeding season.

Keywords

• Alpha Lipoic Acid
• trehalose
• freezing
• thawing
• sperm
• flow cytometry

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