Susceptibility of Stallion Spermatozoa to Different Oxidative Challenges: Role of Seminal Plasma

Abstract

Seminal cryopreservation provides several advantages in horse breeding. However, improvement on post-thaw sperm survival is still necessary. One of the main factors known to impair post-thaw quality of stallion's sperm is the oxidative damage caused by reactive oxygen species (ROS). In this context, the aim of this study was evaluated the effects of ROS on stallion sperm and the possible influence of seminal plasma (SP). Toward this aim, 13 ejaculates from adult stallions (n = 13) were divided into two aliquots which were centrifuged (600g/10 minutes), and the SP was removed and reserved. Pellets were suspended in physiological saline solution (A) or SP (B). Each of these solutions was divided into four aliquots and subjected to challenge with different ROS (superoxide anion, hydrogen peroxide, and hydroxyl radical [OH−]) and the toxic by-product of lipid peroxidation (malondialdehyde [MDA]). Samples were then evaluated for the susceptibility to lipid peroxidation (thiobarbituric acid reactive substances) plasma membrane integrity (eosin–nigrosin), acrosome integrity (Fast Green/rose bengal), mitochondrial activity (3′3 diaminobenzidine), and DNA integrity. Samples incubated in the presence of SP were highly impaired by OH− with regard to motility, plasma membrane integrity, mitochondrial activity, and DNA integrity. On the other hand, in the absence of SP, MDA was highly deleterious, especially with regard to motility, plasma membrane integrity, and mitochondrial activity. Thus, these results indicate that SP may have an important role on the protection of stallion sperm against the damages caused by MDA, an important product of lipid peroxidation.