The Effects on Post-Thaw Sperm Quality and Nuclear DNA Integrity of Supplementation of Low-Density Lipoprotein to Freezing Extender in the Mouse

Year 2024, Volume: 35 Issue: 1, 94 - 100, 29.03.2024

Cengiz Yıldız , İbrahim Erol

https://doi.org/10.36483/vanvetj.1417880

Abstract

Mice are an important research tool for genetic and molecular biology, allowing researchers to explore a variety of human illness models. Egg yolk is a common component of semen extenders for domestic animals and low-density lipoproteins (LDL) from egg yolk have some cryoprotective properties. This study aimed to investigate sperm quality characteristics and nuclear DNA integrity after post-thawing in an extender (18% raffinose + 3% skim milk) supplemented with different concentrations of LDL (2.5%, 5.0%, 7.5%, or 10%) in mice. 18% Raffinose+3% skim milk extender was used as a control group without LDL. CD-1 mice were used in the study, and semen was collected from the cauda epididymis and diluted with the extender. The straws were then frozen and thawed to evaluate progressive motility, viability, plasma membrane (HOST), acrosome, and nuclear DNA integrity parameters. Fresh sperm had the highest progressive motility, viability, plasma membrane integrity, and longevity (endurance) of progressive motility for 4 h in HTF solution. The greatest spermatologic results, including nuclear DNA integrity, were determined in fresh sperm (p<0.05). The greatest post-thaw progressive motility (56.7±1.3%), viability (11.6±2.9%), membrane integrity (63.0±2.7%), and longevity at the end of the 4 h (31.3±2.1%) were found in the group with supplementation of 2.5% LDL compared with the control group. The lowest progressive motility, viability, and plasma membrane integrity were determined in the 10% LDL group (p<0.05). The cryopreservation process increased the rates of fragmented DNA in all tested LDL and control groups compared with fresh sperm (p<0.05). There was no statistical difference between the control and experimental groups in terms of nuclear DNA damage after freezing and thawing (p<0.05). It was concluded that the addition of 2.5% LDL to the extender improved the spermatological quality parameters after freezing and could be used in the freezing and preservation of mouse sperm as it showed a higher protective effect against cold shock.

Keywords

Mouse sperm, Low Density Lypoprotein, Sperm analysis

Project Number

Hatay Mustafa Kemal Üniversitesi BAP birimi (22.GAP.017) nolu proje

Farelerde Sperma Sulandırıcısına Düşük Yoğunluklu Lipoprotein İlavesinin Dondurma Çözdürme Sonrası Sperm Kalitesi ve Nükleer DNA Bütünlüğü Üzerine Etkileri

Abstract

Fareler genetik ve moleküler biyoloji için önemli bir araştırma aracıdır ve araştırmacıların çeşitli insan hastalığı modellerini keşfetmesine olanak tanır. Yumurta sarısı, evcil hayvanlara yönelik sperma sulandırıcılarının yaygın bir bileşenidir ve yumurta sarısından elde edilen düşük yoğunluklu lipoproteinlerin (LDL) bazı kriyoprotektif özellikleri vardır. Bu çalışma, farede farklı konsantrasyonlarda LDL (%2.5, %5.0, %7.5, %10) ilaveli sperma sulandırıcısında (%18 rafinoz + %3 yağsız süt) dondurma sonrası sperm kalite özelliklerini ve nükleer DNA bütünlüğünü araştırmayı amaçladı. LDL içermeyen kontrol grubu olarak %18 Raffinoz+%3 yağsız süt sulandırıcısı kullanıldı. Çalışmada CD-1 fareler kullanıldı ve kauda epididimisten sperma toplandı ve sperma sulandırıcısı ile sulandırıldı, payetlere çekilerek motilite, canlılık, plazma zarı (HOST), akrozom ve nükleer DNA bütünlüğü parametreleri dondurma sonrası değerlendirildi. Nativ sperma, HTF çözeltisinde 4 saat boyunca en yüksek motilite, canlılık, plazma zarı bütünlüğüne ve dayanıklılığa sahip bulundu. Nükleer DNA bütünlüğü taze spermada en yüksek belirlendi (p<0.05). Çözdürme sonrası en yüksek motilite (%56.7±1.3), canlılık (%11.6±2.9), membran bütünlüğü (%63.0±2.7) ve yaşam süresi (%31.3±2.1) %2.5 LDL takviyeli grupta, en düşük progresif motilite, canlılık ve plazma membran bütünlüğü ise %10 LDL grubunda belirlendi (p<0.05). Kriyoprezervasyon işlemi, taze sperm ile karşılaştırıldığında tüm LDL ve kontrol gruplarında parçalanmış DNA oranlarını arttırdı (p<0.05). Kontrol ve deney grupları arasında donma ve çözdürme sonrası nükleer DNA hasarı açısından istatistiksel fark yoktu (p<0.05). Sulandırıcıya %2.5 LDL ilavesinin dondurma sonrası spermatolojik kalite parametrelerini iyileştirdiği ve fare sperminin dondurulması ve korunmasında daha yüksek oranda soğuk şokuna karşı koruyucu etki gösterdiği için kullanılabileceği sonucuna varıldı.

Keywords

Fare spermi, Düşük Yoğunluklu Lipoprotein, Sperm analizi

Ethical Statement

Hatay Mustafa Kemal Üniversitesi Hayvan Deneyleri Yerel Etik Kurulu’ndan 21/05/2019 tarihli 20879420--825 sayılı izin alınarak yapılmıştır.

Supporting Institution

Hatay Mustafa Kemal Üniversitesi BAP birimi

Project Number

Hatay Mustafa Kemal Üniversitesi BAP birimi (22.GAP.017) nolu proje

Thanks

Bu çalışma Hatay Mustafa Kemal Üniversitesi BAP birimi tarafından (22.GAP.017) desteklenmiştir.

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