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Optimising Stallion Semen Cryopreservation: Preliminary Insights Into Pre-Centrifugation Extender Effects
Bruna Merci de Zutter1, Camila de Paula Freitas-Dell'Aqua1, José Antônio Dell'Aqua-Junior1
1Department of Veterinary Surgery and Animal Reproduction, School of Veterinary Medicine and Animal Science, São Paulo State University (UNESP), Botucatu, São Paulo, Brazil.
Reproduction in Domestic Animals = Zuchthygiene
|October 22, 2025
Summary
Adding cholesterol to equine sperm extenders before cryopreservation significantly improves post-thaw sperm quality. Cholesterol-based extenders enhance sperm kinematics and plasma membrane integrity in stallions.
Area of Science:
- Veterinary Reproduction
- Sperm Cryopreservation Biology
- Equine Science
Background:
- Cryopreservation of equine spermatozoa is crucial for genetic preservation and artificial insemination.
- Optimizing extender composition is essential to maintain sperm viability and function post-thaw.
- Previous studies have explored various additives, but the role of cholesterol in pre-centrifugation extenders requires further investigation.
Purpose of the Study:
- To evaluate the impact of cholesterol, pentoxifylline, and casein, with or without skim milk, on equine sperm cryosurvival.
- To determine the effectiveness of these additives when incorporated into extenders before centrifugation and cryopreservation.
- To assess the influence of cholesterol-based extenders on post-thaw sperm kinematics and plasma membrane integrity.
Main Methods:
- Seven ejaculates from four stallions were used (n=28).
- Sperm samples were divided into four extender groups: skim milk (SM), SM + pentoxifylline (SMP), SM + cholesterol (SMCho), and cholesterol + casein (ChoCa).
- Sperm kinematics and plasma membrane integrity were assessed immediately and 30 minutes post-thaw after centrifugation and freezing.
Main Results:
- Equine sperm extenders supplemented with cholesterol (SMCho and ChoCa) demonstrated superior cryosurvival compared to skim milk (SM) and skim milk + pentoxifylline (SMP) groups.
- Significant improvements in post-thaw sperm kinematics and plasma membrane integrity were observed in cholesterol-supplemented groups.
- These findings highlight the protective effects of cholesterol against cryodamage.
Conclusions:
- Cholesterol-based extenders significantly enhance post-thaw equine sperm quality.
- Incorporating cholesterol into extenders prior to cryopreservation is a promising strategy for improving stallion sperm viability.
- These results suggest cholesterol plays a vital role in protecting sperm membranes during the cryopreservation process.
