[en] Myeloperoxidase (MPO) is a pro-oxidant enzyme contained in, and released by, neutrophils during degranulation or after lysis. Post-thaw semen contains MPO and its concentration is associated with decreased sperm motility. Recently, MPO concentration in post-thaw semen was shown to be associated with presence of non-sperm cells. The aim of this study was to describe the effect of centrifugal fractionation of semen prior to cryopreservation on post-thaw concentrations of non-sperm cells and MPO.
The experimental design consisted in freezing semen with or without previous centrifugation through two concentrations of single layer colloid media. Non-sperm cells and MPO concentrations were assessed in pellet and upper layer at each step of the procedure and MPO was detected in cells by immunocytochemistry.
Single layer colloid centrifugation decreased non-sperm cells and MPO concentrations in post-thaw semen. The MPO concentration was correlated with concentration of non-sperm cells in the upper layer of the supernatant. In post-thaw semen, with or without previous single layer colloid centrifugation, MPO concentration was correlated with concentration of non-sperm cells. Overall, neutrophils were rarely observed and non-sperm cells were mainly epithelial cells or cellular debris, as demonstrated by MPO immunocytochemistry.
Following single layer colloid centrifugation, MPO concentration was decreased, and correlations observed in different samples of the experiments revealed an association between concentrations of MPO and non-sperm cells. At all steps of the semen processing and cryopreservation, MPO immunostaining was clearly identified only on non-sperm cells. Our results demonstrate that non-sperm cells present in fresh semen release MPO during freezing.
Research Center/Unit :
Clinique Equine, Département des sciences cliniques des animaux de compagnie et équidés, Faculté de Médecine Vétéri Equine theriogenology, Large Animal Clinical Sciences Department, Texas A&M university
Disciplines :
Animal production & animal husbandry Veterinary medicine & animal health Biotechnology
Author, co-author :
Ponthier, Jérôme ; Université de Liège - ULiège > Département clinique des animaux de compagnie et des équidés > Obstétrique et pathologie de la reproduction des équidés
Teague, Sheila; Texas A&M university > Departement of Large Animal Clinical Sciences > Equine Theriogenology
Franck, Thierry ; Université de Liège - ULiège > Département clinique des animaux de compagnie et des équidés > Anesthésiologie gén. et pathologie chirurg. des grds animaux
de la Rebière de Pouyade, Geoffroy ; Université de Liège - ULiège > Département clinique des animaux de compagnie et des équidés > Anesthésiologie gén. et pathologie chirurg. des grds animaux
Serteyn, Didier ; Université de Liège - ULiège > Département clinique des animaux de compagnie et des équidés > Anesthésiologie gén. et pathologie chirurg. des grds animaux
Brinsko, Steven; Texas A&M university > Departement of Large Animal Clinical Sciences > Equine theriogenology
Love, Charles; Texas A&M university > Departement of Large Animal Clinical Sciences > Equine theriogenology
Blanchard, Terry; Texas A&M university > Departement of Large Animal Clinical Sciences > Equine theriogenology
Varner, Dickson; Texas A&M university > Departement of Large Animal Clinical Sciences > Equine theriogenology
Deleuze, Stefan ; Université de Liège - ULiège > Département clinique des animaux de compagnie et des équidés > Obstét. et path. de la reprod. des anim. de comp. et équidés
Language :
English
Title :
Effect of non-sperm cells removal with single layer colloidal centrifugation on myeloperoxidase concentration in post-thaw equine semen
Publication date :
December 2013
Journal title :
Theriogenology
ISSN :
0093-691X
eISSN :
1879-3231
Publisher :
Elsevier, New York, United States - New York
Volume :
80
Issue :
9
Pages :
1082-1087
Peer reviewed :
Peer Reviewed verified by ORBi
Name of the research project :
SPERMPO
Funders :
ULiège - Université de Liège Texas A&M university Clinique Equine, Département des sciences cliniques des animaux de compagnie et équidés, Faculté de Médecine Vétérinaire, Ulg
Agarwal A., Ikemoto I., Loughlin K.R. Relationship of sperm parameters with levels of reactive oxygen species in semen specimens. J Urol 1994, 152:107-110.
O'Flaherty C.M., Beorlegui N.B., Beconi M.T. Reactive oxygen species requirements for bovine sperm capacitation and acrosome reaction. Theriogenology 1999, 52:289-301.
Baumber J., Ball B.A., Gravance C.G., Medina V., Davies-Morel M.C. The effect of reactive oxygen species on equine sperm motility, viability, acrosomal integrity, mitochondrial membrane potential, and membrane lipid peroxidation. J Androl 2000, 21:895-902.
Peris S.I., Bilodeau J.F., Dufour M., Bailey J.L. Impact of cryopreservation and reactive oxygen species on DNA integrity, lipid peroxidation, and functional parameters in ram sperm. Mol Reprod Dev 2007, 74:878-892.
Henkel R., Kierspel E., Stalf T., Mehnert C., Menkveld R., Tinneberg H.R., et al. Effect of reactive oxygen species produced by spermatozoa and leukocytes on sperm functions in non-leukocytospermic patients. Fertil Steril 2005, 83:635-642.
Wang A., Fanning L., Anderson D.J., Loughlin K.R. Generation of reactive oxygen species by leukocytes and sperm following exposure to urogenital tract infection. Arch Androl 1997, 39:11-17.
Wang A.W., Zhang H., Ikemoto I., Anderson D.J., Loughlin K.R. Reactive oxygen species generation by seminal cells during cryopreservation. Urology 1997, 49:921-925.
Baumber J., Vo A., Sabeur K., Ball B.A. Generation of reactive oxygen species by equine neutrophils and their effect on motility of equine spermatozoa. Theriogenology 2002, 57:1025-1033.
Franck T., Grulke S., Deby-Dupont G., Deby C., Duvivier H., Peters F., et al. Development of an enzyme-linked immunosorbent assay for specific equine neutrophil myeloperoxidase measurement in blood. J Vet Diagn Invest 2005, 17:412-419.
Serteyn D., Grulke S., Franck T., Mouithys-Mickalad A., Deby-Dupont G. La myeloperoxydase des neutrophiles, une enzyme de défense au capacité oxydantes. Annales de Médecine Vétérinaire 2003, 147:79-93.
Taurog A., Dorris M.L. Myeloperoxidase-catalyzed iodination and coupling. Arch Biochem Biophys 1992, 296:239-246.
Mathy-Hartert M., Bourgeois E., Grulke S., Deby-Dupont G., Caudron I., Deby C., et al. Purification of myeloperoxidase from equine polymorphonuclear leucocytes. Can J Vet Res 1998, 62:127-132.
de la Rebiere G., Franck T., Deby-Dupont G., Salciccia A., Grulke S., Peters F., et al. Effects of unfractionated and fractionated heparins on myeloperoxidase activity and interactions with endothelial cells: possible effects on the pathophysiology of equine laminitis. Vet J 2008, 178:62-69.
Grulke S., Franck T., Gangl M., Peters F., Salciccia A., Deby-Dupont G., et al. Myeloperoxidase assay in plasma and peritoneal fluid of horses with gastrointestinal disease. Can J Vet Res 2008, 72:37-42.
Ponthier J., Desvals M., Franck T., de la Rebiere de Pouyade G., Spalart M., Palmer E., et al. Myeloperoxidase in equine semen: concentration and localization during freezing processing. J Equine Vet Sci 2012, 32:32-37.
Ponthier J., Franck T., Detilleux J., Mottart E., Serteyn D., Deleuze S. Association between myeloperoxidase concentration in equine frozen semen and post-thawing parameters. Reprod Domest Anim 2010, 45:811-816.
Ponthier J., de la Rebiere de Pouyade G., Desvals M., Spalart M., Franck T., Serteyn D., et al. Is neutrophil elastase associated with myeloperoxidase concentration and post-thawing parameters in equine frozen semen?. Anim Reprod Sci 2010, 121S:S200-S202.
Edmond A.J., Teague S.R., Brinsko S.P., Comerford K.L., Waite J.A., Mancill S.S., et al. Effect of density-gradient centrifugation on quality and recovery rate of equine spermatozoa. Anim Reprod Sci 2008, 107. 318.
Villegas J., Schulz M., Soto L., Iglesias T., Miska W., Sanchez R. Influence of reactive oxygen species produced by activated leukocytes at the level of apoptosis in mature human spermatozoa. Fertil Steril 2005, 83:808-810.
Johansson C.S., Matsson F.C., Lehn-Jensen H., Nielsen J.M., Petersen M.M. Equine spermatozoa viability comparing the NucleoCounter SP-100 and the eosin-nigrosin stain. Anim Reprod Sci 2008, 107:325-326.
Waite J.A., Love C.C., Brinsko S.P., Teague S.R., Salazar J.L., Mancill S.S., et al. Factors impacting equine sperm recovery rate and quality following cushioned centrifugation. Theriogenology 2008, 70:704-714.
Pillet E., Duchamp G., Batellier F., Beaumal V., Anton M., Desherces S., et al. Egg yolk plasma can replace egg yolk in stallion freezing extenders. Theriogenology 2011, 75:105-114.
Art T., Franck T., Lekeux P., de Moffarts B., Couetil L., Becker M., et al. Myeloperoxidase concentration in bronchoalveolar lavage fluid from healthy horses and those with recurrent airway obstruction. Can J Vet Res 2006, 70:291-296.
Brinsko S.P. Retrograde ejaculation in a stallion. J Am Vet Med Assoc 2001, 218:551-553.
Villegas J., Schulz M., Soto L., Sanchez R. Bacteria induce expression of apoptosis in human spermatozoa. Apoptosis 2005, 10:105-110.
Johnston S.D., Kamolpatana K., Root-Kustritz M.V., Johnston G.R. Prostatic disorders in the dog. Anim Reprod Sci 2000, 60-61:405-415.
Smith J. Canine prostatic disease: a review of anatomy, pathology, diagnosis, and treatment. Theriogenology 2008, 70:375-383.
Roumeguere T., Delree P., Van Antwerpen P., Rorive S., Vanhamme L., de Ryhove L.D., et al. Intriguing location of myeloperoxidase in the prostate: a preliminary immunohistochemical study. Prostate 2012, 72:507-513.
Black S.J. Extracellular matrix, leukocyte migration and laminitis. Vet Immunol Immunopathol 2009, 129:161-163.
de la Rebiere de Pouyade G., Riggs L.M., Moore J.N., Franck T., Deby-Dupont G., Hurley D.J., et al. Equine neutrophil elastase in plasma, laminar tissue, and skin of horses administered black walnut heartwood extract. Vet Immunol Immunopathol 2010, 135:181-187.
Riggs L.M., Franck T., Moore J.N., Krunkosky T.M., Hurley D.J., Peroni J.F., et al. Neutrophil myeloperoxidase measurements in plasma, laminar tissue, and skin of horses given black walnut extract. Am J Vet Res 2007, 68:81-86.
Graham-Pole J., Davie M., Willoughby M.L. Cryopreservation of human granulocytes in liquid nitrogen. J Clin Pathol 1977, 30:758-762.
