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Home / Equine Research Bank / Theriogenology / Generation of reactive oxygen species by equine neutrophils ...
Theriogenology2002; 57(3); 1025-1033; doi: 10.1016/s0093-691x(01)00710-5

Generation of reactive oxygen species by equine neutrophils and their effect on motility of equine spermatozoa.

Abstract: Contaminating leukocytes in the ejaculate are an important source of reactive oxygen species (ROS) in human semen. When present in sufficient numbers, they can have a detrimental influence on sperm function in humans. Unfortunately, there is little published information regarding the importance of leukocytes in stallion semen. The objectives of this study were to determine the production of hydrogen peroxide (H2O2) by activated equine neutrophils and to examine the effect of this ROS production on equine sperm motility in vitro. Motile equine spermatozoa (two ejaculates each from four stallions) and peripheral blood neutrophils were isolated on discontinuous Percoll gradients, washed and resuspended in a modified Tyrode's medium. Spermatozoa (25 x 10(6)/ml) were incubated for 30 min at 38 C with neutrophils (0,0.5 x 10(6),1 x 10(6), 5 x 10(6) and 10 x 10(6)/ml) activated by either the protein kinase C agonist, 12-myristate, 13-acetate phorbol ester (PMA; 100 nM) or the leukocyte chemotactic peptide, formyl-methionyl-leucyl-phenylalanine (FMLP; 0.1 mM). Sperm motility was determined by computer-assisted semen analysis (CASA) at time 0 min (T0) and time 30 min (T30), and H2O2 was measured at T30 with the Amplex Red assay kit. At T30, there was a significant (P < 0.01) increase in H2O2 with the addition of 5 x 10 and 10 x 10(6) neutrophils/ml activated by FMLP (0.76 +/- 0.3 and 0.99 +/- 0.4 microM, respectively, versus 0.0024 +/- 0.002 microM in sperm alone), and this increase was associated with a significant (P < 0.001) decrease in total motility (52 +/- 5.1 and 48 +/- 6.0%, respectively, versus 80 +/- 4.7% in sperm alone). At T30, there was also a significant (P < 0.001) increase in H2O2 with the addition of 5 x 10(6) and 10 x 10(6) neutrophils/ml activated by PMA (1.88 +/- 0.2 and 2.07 +/- 0.3 microM, respectively, versus 0.0009 +/- 0.0006 microM in sperm alone). The results of this study demonstrate that 5 x 10(6) activated neutrophils/ml are sufficient to impair equine sperm motility in vitro.
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Publication Date: 2002-06-04 PubMed ID: 12041897DOI: 10.1016/s0093-691x(01)00710-5Google Scholar: Lookup
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  • Journal Article
  • Research Support
  • Non-U.S. Gov't

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This study explores the impact of reactive oxygen species (ROS) produced by leukocytes (white blood cells) present in stallion semen on the motility of sperm. It demonstrates that equine neutrophils, a type of white blood cell, produce hydrogen peroxide (H2O2), a ROS, which significantly decreases the motility of sperm in horse semen when present in significant numbers.

Objectives and Methodology

  • The research aimed to investigate the production of hydrogen peroxide (H2O2), a ROS, by activated equine neutrophils, and to understand its effect on the motility (movement) of equine spermatozoa in vitro.
  • The researchers used semen from four stallions and isolated motile spermatozoa and peripheral blood neutrophils (a type of leukocyte) for this study.
  • The isolated spermatozoa and neutrophils were resuspended in a medium and incubated with varying quantities of neutrophils, which were activated using PMA (a protein kinase C agonist) or FMLP (a leukocyte chemotactic peptide).
  • Sperm motility was measured using computer assistance at two times (T0 and T30) and hydrogen peroxide production was measured at T30.

Findings

  • The study found that an increase in the concentration of H2O2 occurred with the presence of 5 x 10^6 and 10 x 10^6 neutrophils/mL activated by FMLP and PMA. This increase was much higher than what was found in sperm samples measured alone.
  • This increase in H2O2 was significantly (P < 0.001) linked to a reduction in sperm motility, illustrating that the ROS negatively influences sperm functionality.
  • These results were further confirmed by similar decreases in motility when neutrophils were activated by PMA.

Conclusion

  • The results indicate that reactive oxygen species, specifically hydrogen peroxide produced by activated equine neutrophils, are sufficient to impair equine sperm motility in vitro. This finding may have broader implications for the study of fertility in horses and potentially other species.

Cite This Article

APA
Baumber J, Vo A, Sabeur K, Ball BA. (2002). Generation of reactive oxygen species by equine neutrophils and their effect on motility of equine spermatozoa. Theriogenology, 57(3), 1025-1033. https://doi.org/10.1016/s0093-691x(01)00710-5

Publication

Theriogenology
ISSN: 0093-691X
NlmUniqueID: 0421510
Country: United States
Language: English
Volume: 57
Issue: 3
Pages: 1025-1033

Researcher Affiliations

Baumber, J
  • Department of Population Health and Reproduction, University of California, Davis 95616, USA.
Vo, A
    Sabeur, K
      Ball, B A

        MeSH Terms

        • Animals
        • Horses / physiology
        • Hydrogen Peroxide / metabolism
        • Male
        • N-Formylmethionine Leucyl-Phenylalanine / pharmacology
        • Neutrophil Activation / drug effects
        • Neutrophils / drug effects
        • Neutrophils / metabolism
        • Reactive Oxygen Species / metabolism
        • Sperm Motility / drug effects
        • Tetradecanoylphorbol Acetate / pharmacology

        Citations

        This article has been cited 10 times.
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        5. Pintus E, Ros-Santaella JL. Impact of Oxidative Stress on Male Reproduction in Domestic and Wild Animals. Antioxidants (Basel) 2021 Jul 20;10(7).
          doi: 10.3390/antiox10071154pubmed: 34356386google scholar: lookup
        6. Millán de la Blanca MG, Martínez-Nevado E, Castaño C, García J, Bernal B, Toledano-Díaz A, Esteso MC, Bóveda P, Martínez-Fresneda L, López-Sebastián A, Santiago-Moreno J. Sperm Cryopreservation in American Flamingo (Phoenicopterus Ruber): Influence of Cryoprotectants and Seminal Plasma Removal. Animals (Basel) 2021 Jan 15;11(1).
          doi: 10.3390/ani11010203pubmed: 33467758google scholar: lookup
        7. Miró J, Marín H, Catalán J, Papas M, Gacem S, Yeste M. Seminal Plasma, Sperm Concentration, and Sperm-PMN Interaction in the Donkey: An In Vitro Model to Study Endometrial Inflammation at Post-Insemination. Int J Mol Sci 2020 May 14;21(10).
          doi: 10.3390/ijms21103478pubmed: 32423134google scholar: lookup
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        9. McCarthy MJ, Baumber J, Kass PH, Meyers SA. Osmotic stress induces oxidative cell damage to rhesus macaque spermatozoa. Biol Reprod 2010 Mar;82(3):644-51.
          doi: 10.1095/biolreprod.109.080507pubmed: 19846599google scholar: lookup
        10. Egyptien S, Deleuze S, Ledeck J, Ponthier J. Sperm Quality Assessment in Stallions: How to Choose Relevant Assays to Answer Clinical Questions. Animals (Basel) 2023 Oct 6;13(19).
          doi: 10.3390/ani13193123pubmed: 37835729google scholar: lookup
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