Effect of Mitoquinone on sperm quality of cryopreserved stallion semen.

Overview

• This study examined the impact of mitochondria-targeted antioxidant Mitoquinone (MitoQ) on the quality of stallion semen after freezing and thawing.
• Researchers tested various concentrations of MitoQ to determine its effect on sperm motility and other quality parameters post-cryopreservation.

Study Objectives

• To assess whether MitoQ supplementation improves the quality of cryopreserved stallion semen.
• To determine the optimal concentration of MitoQ for enhancing sperm motility and viability after thawing.
• To investigate if MitoQ affects other sperm quality parameters such as membrane integrity, DNA integrity, mitochondrial potential, apoptosis, and reactive oxygen species (ROS) levels.

Methodology

• Semen samples were collected from three fertile stallions aged between 10 and 13 years.
• Samples were filtered and centrifuged using a skimmed milk-based extender and then diluted to a concentration of 50 × 10⁶ sperm/mL in a freezing medium.
• The diluted semen was divided into five groups with different MitoQ concentrations: 0 (control), 25, 50, 100, and 200 nM.
• Samples were cooled, equilibrated, frozen, and then thawed for analysis.
• Post-thaw sperm were evaluated at multiple time points for:
• Motility and kinetic parameters (e.g., amplitude of lateral head displacement)
• Sperm viability
• Integrity of plasma membrane and acrosome
• DNA integrity
• Mitochondrial membrane potential
• Apoptotic status
• Intracellular reactive oxygen species (ROS) concentrations

Key Findings

• MitoQ at concentrations of 25, 50, and 100 nM significantly improved total sperm motility 30 minutes after thawing compared to the control group.
• The 25 nM concentration also improved sperm amplitude of lateral head displacement, an indicator of sperm movement quality.
• High concentration (200 nM) of MitoQ negatively affected sperm motility, kinetics, and viability, indicating a toxic or detrimental effect at this level.
• No significant effects were observed at any tested concentrations on:
• Plasma membrane integrity
• Acrosome integrity
• DNA integrity
• Mitochondrial membrane potential
• Intracellular ROS levels

Conclusions and Implications

• MitoQ supplementation can mildly enhance sperm motility and movement characteristics in cryopreserved stallion semen, especially at the concentration of 25 nM.
• Higher concentrations (200 nM) are harmful, reducing sperm quality parameters and should be avoided.
• MitoQ does not appear to affect structural sperm integrity or oxidative stress markers post-thaw, suggesting its main benefit is on motility.
• The findings provide guidance for optimizing cryopreservation protocols with mitochondria-targeted antioxidants to improve fertility outcomes in stallions.