Cryopreservation of stallion semen: Effect of adding antioxidants to the freezing medium on sperm physiology

Abstract

Cryopreservation of stallion semen has not reached the level of efficiency and positive results described in other species. This is mainly due to the greater sensitivity of stallion sperm to the freezing process, showing higher rates of oxidative stress and plasma membrane damage, which trigger the activation of several cell damage pathways that ultimately culminate in DNA fragmentation and cell death. Therefore, finding molecules that improve the efficiency of this technique in stallion by preventing oxidative stress and cell damage is required. Thus, the aim of the present study was to evaluate the effect of adding three antioxidants (MnTBAP, NAC and FeTPPS) to the freezing medium on the quality and functional parameters of stallion sperm. Semen samples from three stallions frozen with the antioxidants were evaluated in two conditions: (a) adding the antioxidants before freezing, and (b) before and after freezing. Plasma membrane integrity, mitochondrial membrane potential, lipid peroxidation, intracellular ROS levels, membrane lipid disorder, DNA damage, sperm motility and binding to the zona pellucida were assessed. The results showed that MnTBAP was the antioxidant treatment that best controlled the oxidative stress process and post‐thaw cell damage, showing higher plasma membrane integrity, mitochondrial membrane potential, sperm motility, number of spermatozoa bound to the zona pellucida of bovine oocytes and lower lipid disorder. Additionally, it was determined that a second post‐thaw application of antioxidants is detrimental since induced higher cell damage and lower sperm motility, without showing any beneficial effect on the spermatozoa.

Efficiency of Tris-Based Extender Steridyl for Semen Cryopreservation in Stallions

“…The composition of extender is one of the main factors in determining the viability of frozen semen [ 18 ]. Although most extenders for freezing equine semen provide sperm viability and pregnancy rates, new freezing extenders or modifications of freezing protocols are still required [ 4 , 18 , 19 , 20 ]. Stallion semen freezing protocols vary from laboratory to laboratory, using different extenders, both commercial and homemade, and different cryoprotectants.…”

“…Fertility is the main indicator of sperm quality. Fertility is influenced by many factors, one of which is the extender [ 4 , 6 , 18 , 26 ]. In our study, semen frozen in both diluents had good fertility.…”

“…However, breeders prefer to use fresh or chilled semen [ 1 , 2 ]. The fertilizing ability of stallion sperm after freezing is lower than in other species [ 3 , 4 ]. One of the reasons is the selection of stallions based on the exterior sports quality, and productivity, and not focused on reproductive qualities [ 4 , 5 ].…”

“…The fertilizing ability of stallion sperm after freezing is lower than in other species [ 3 , 4 ]. One of the reasons is the selection of stallions based on the exterior sports quality, and productivity, and not focused on reproductive qualities [ 4 , 5 ]. Sperm freezing in horses is very important for the preservation of valuable, rare, and endangered genotypes [ 6 , 7 , 8 ].…”

In Vitro Effect of Resorcinol on Bovine Spermatozoa in Process of Cryopreservation

“…Some individuals can have problem with cryotolerance and it only depends on a suitable freezing buffer (Šichtař et al, 2017). Several substances were exanimated, which can be a potential part of freezing extender (Akhter et al, 2012;Mousavi et al, 2019;Contreras et al, 2019). 1,mdihydroxyphenol) is widely used in the manufacture of tires and rubber goods.…”