Optimal sperm fertility relies on a delicate balance between ROS and antioxidant activity. The most pernicious ROS is H₂O₂, and species differences in susceptibility to H₂O₂-induced damage has been poorly characterised. As such, the aim of this study was to compare the susceptibility of human and horse spermatozoa to H₂O₂-mediated damage.
Spermatozoa from each species were exposed to H₂O₂(0,0.25,0.5,1.0,2.0mM) for up to 2h at 37ºC, after which analyses of motility (CASA), oxidative stress and viability (MitoSox Red, DCF-DA, LIVE/DEAD violet), and DNA damage (SCSA) were performed. Horse spermatozoa were more resilient to H₂O₂ insult compared to human spermatozoa, with the total motility of human spermatozoa decreasing within 1h exposure to 0.25mM H2O2(0h;76.6±2.6%,1h;51.2±9.3%,P≤0.01), while horse spermatozoa did not suffer any decrease in total motility until 2h exposure at 2mM H2O2 (0h;82.5±3.6%, 2h;34.7±14.5%, P≤0.001). At 2mM H₂O₂, oxidative stress (viable, MitoSox Red and DCF-DA positive cells) increased after 2h of exposure in human spermatozoa (0h;7.852±5.9%,2h;33.3±7%, P≤0.001), though no such increase was observed in the horse. Similarly, DNA damage significantly increased after 2mM H₂O₂ insult in humans (0h;24.5±10.5%,2h;87.2±4.6%, P≤0.001) whilst there was no detectable increase in DNA damage in the horse.Despite the deleterious effects on oxidative stress and DNA damage in the human, H₂O₂ exposure had no effect on sperm viability.
This study revealed that horse spermatozoa are more resilient to H₂O₂-mediated oxidative stress compared to human spermatozoa. This may be due to evolutionary adaptations associated with their use of oxidative phosphorylation for ATP production, a process which generates large amounts of ROS, compared to humans, who primarily rely on glycolysis. Further research in this field will inform on species-specific antioxidant requirements both in vivo (dietary) and in vitro (sperm storage and IVF).