Related Experiment Video
Updated: Jun 24, 2026
The Examination of Peroxidase-Positive Leukocytes in Semen
Published on: January 19, 2024
Association between myeloperoxidase concentration in equine frozen semen and post-thawing parameters
J Ponthier1, T Franck, J Detilleux
1Equine Clinic, Veterinary Medicine Faculty, ULg University of Liége, Liége, Belgium.
Abstract:
Despite improvement of techniques, semen of 20% of stallions remains unfreezable. Recent studies focused on the impact of reactive oxygen species and oxidant enzymes on semen characteristics. Myeloperoxidase (MPO) is a pro-oxidant enzyme contained in and released by neutrophils during degranulation or after cell lysis. It is responsible for the formation of hypochlorous acid, a strong oxidant agent, which could damage spermatozoa. The aim of this study was to determine the relation between MPO concentration and characteristics of frozen semen from stallions. Thirty-five straws from different stallions were analysed. Post-thawing spermatozoal concentration, and progressive and total motility were determined by Computer-Assisted Semen Analysis. Freezability was determined according to post-thawing progressive motility (above or below 15%). Percentage of alive spermatozoa and abnormal forms was determined after Eosin-Nigrosin and Diff-Quick(®) staining, respectively. Post-thawing MPO concentration was measured by enzyme-linked immunosorbent assay (ELISA). Our study shows that frozen thawed semen contains large amounts of free MPO. We also observed that post-thawing MPO ELISA assay can be used as an indicator of equine semen freezability. High MPO concentration samples showed lower total and progressive motility. A higher proportion of abnormal head shape associated with acrosome reaction was observed in our late examinations of the high concentration MPO group. Our results show that MPO adversely affects total and progressive motility of equine semen. A negative correlation between normal motile forms and MPO concentration was also observed. The effect of MPO on dead or abnormal forms remains to be precised.
Insights
High concentrations of myeloperoxidase (MPO) in stallion semen negatively impact sperm motility and freezability. Measuring MPO levels can indicate semen quality and predict successful cryopreservation outcomes.
Area of Science:
- Reproductive Biology
- Animal Science
- Biochemistry
Background:
- Semen cryopreservation in stallions faces challenges, with 20% remaining unfreezable.
- Reactive oxygen species and oxidant enzymes are implicated in reduced semen quality.
- Myeloperoxidase (MPO), released by neutrophils, produces hypochlorous acid, a potent oxidant damaging spermatozoa.
Purpose of the Study:
- To investigate the relationship between myeloperoxidase (MPO) concentration and the characteristics of frozen-thawed stallion semen.
- To assess MPO as a potential biomarker for equine semen freezability.
Main Methods:
- Analysis of 35 frozen-thawed stallion semen samples.
- Evaluation of post-thawing sperm concentration, progressive motility, and total motility using Computer-Assisted Semen Analysis.
- Assessment of sperm viability and abnormality using staining techniques (Eosin-Nigrosin, Diff-Quick®).
- Quantification of post-thawing MPO concentration via enzyme-linked immunosorbent assay (ELISA).
Main Results:
- Frozen-thawed stallion semen contains significant levels of free MPO.
- Post-thawing MPO concentration is a reliable indicator of semen freezability.
- Higher MPO concentrations correlated with reduced total and progressive sperm motility.
- Elevated MPO levels were associated with increased abnormal head shape and acrosome reactions.
Conclusions:
- Myeloperoxidase (MPO) negatively affects the motility of equine spermatozoa.
- MPO concentration serves as a predictive marker for stallion semen cryopreservation success.
- Further research is needed to elucidate MPO's precise impact on dead or morphologically abnormal sperm forms.
