title: Effects of cryopreservation on sperm viability, synthesis of reactive oxygen species, and DNA damage of bovine sperm.
abstract: The objective was to examine if there are relationships between alterations in sperm viability, reactive oxygen species (ROS) synthesis, and DNA integrity induced by cryopreservation of bovine sperm. Four ejaculates were collected from each of six bulls. Each ejaculate was diluted and divided into two aliquots; one was incubated for 24 hours at 37 °C, and the other frozen, thawed, and incubated for 24 hours at 37 °C. Analyses of quality of sperm were performed after 0, 3, 6, 12, and 24 hours of incubation. Progressive motile sperm was determined with computer assisted sperm analysis. Percentages of plasma membrane- and acrosome-intact sperm, sperm with a high mitochondrial membrane potential, sperm showing a high degree of DNA fragmentation (%DFI), and their reactive oxygen species content were assessed with dichlorofluorescein-diacetate, dihydrorhodamine, diaminofluorescein diacetate, and mitochondrial superoxide indicator using flow cytometry. Although all other sperm parameters showed alterations (P < 0.05) during the 24-hour incubation time, %DFI stayed constant (P > 0.05, 0.91 ± 0.23) in nonfrozen sperm. Cryopreservation induced changes of all sperm parameters (P < 0.05). In contrast to all other sperm parameters, dichlorofluorescein-diacetate-fluoroescence indicating the synthesis of H2O2 showed a similar exponential rise (P < 0.05) like the %DFI values in frozen sperm. In conclusion, changes of DNA integrity in frozen sperm seem to be related to synthesis of H2O2 but not to sperm viability and synthesis of other reactive oxygen species.
authors.affiliation: Clinic for Cattle, University of Veterinary Medicine Hannover Foundation, Hannover, Germany. Electronic address: gurlerhakan@hotmail.com.
authors.forename: H
authors.initials: H
authors.lastname: Gürler
authors.affiliation: Vetsuisse Faculty, Clinic of Reproductive Medicine, Zürich, Switzerland.
authors.forename: E
authors.initials: E
authors.lastname: Malama
authors.affiliation: Clinic for Cattle, University of Veterinary Medicine Hannover Foundation, Hannover, Germany.
authors.forename: M
authors.initials: M
authors.lastname: Heppelmann
authors.affiliation: Süt Kardesler A.S. Animal Breeding Center, Izmir, Turkey.
authors.forename: O
authors.initials: O
authors.lastname: Calisici
authors.affiliation: Besamungsvereins Neustadt Aisch, Neustadt an der Aisch, Germany.
authors.forename: C
authors.initials: C
authors.lastname: Leiding
authors.affiliation: Department of Production Animal Health, Faculty of Veterinary Medicine, University of Calgary, Calgary, Alberta, Canada.
authors.forename: J P
authors.initials: JP
authors.lastname: Kastelic
authors.affiliation: Vetsuisse Faculty, Clinic of Reproductive Medicine, Zürich, Switzerland.
authors.forename: H
authors.initials: H
authors.lastname: Bollwein
mesh_terms: D000818:Animals; D002417:Cattle / Q000502:physiology*; D002470:Cell Survival; D015925:Cryopreservation / Q000662:veterinary*; D004249:DNA Damage / Q000502:physiology*; D008297:Male; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013094:Spermatozoa / Q000502:physiology*; D013997:Time Factors
chemical_list: D017382:Reactive Oxygen Species
keywords: Bull spermatozoa; Reactive oxygen species; Sperm chromatin structure assay (SCSA); Sperm freezing; Sperm mitochondria