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Reactive oxygen species generators affect quality parameters and apoptosis markers differently in red deer spermatozoa.
Reproduction 2009; 137(2):225-35R

Abstract

Fe(2)(+)/ascorbate, hydrogen peroxide (H(2)O(2)), and hypoxanthine/xanthine oxidase (XOD) are commonly used for inducing oxidative stress on spermatozoa. A comparative study of these agents was carried out on thawed spermatozoa from red deer. First, we tested a high, medium, and low concentration of each agent: 100, 10, and 1 microM Fe(2)(+) (hydroxyl radical generator); 1 mM, 100, and 10 microM H(2)O(2); and 100, 10, and 1 mU/ml XOD (superoxide and H(2)O(2) generator), incubated at 37 degrees C for 180 min. Intracellular reactive oxygen species (ROS; H(2)DCFDA) increased with dose and time similarly for the three systems at each concentration level. Motility and mitochondrial membrane potential (Deltapsi(m)) were considerably decreased by H(2)O(2) (1 mM and 100 microM) and XOD (100 and 10 mU/ml). Only 1 mM H(2)O(2) reduced viability. The antioxidant Trolox (10 microM) reduced intracellular ROS, but could not prevent the H(2)O(2) or XOD effects. In a second experiment, YO-PRO-1 and M540 were used as apoptotic and membrane stability markers respectively. Only H(2)O(2) increased the proportion of apoptotic and membrane-destabilized spermatozoa. Catalase added to XOD prevented Deltapsi(m) loss, confirming that H(2)O(2) was the causative agent, not superoxide. In a third experiment, caspase activation was tested using the (FAM-VAD-FMK) probe. Viable spermatozoa with activated caspases could be detected in untreated samples, and only H(2)O(2) increased their proportion after 60 min. There were important differences between ROS generators, H(2)O(2) being the most cytotoxic. Although H(2)O(2) and XOD caused Deltapsi(m) dissipation, this was not reflected in increasing apoptotic markers.

Authors+Show Affiliations

Biology of Reproduction Group, National Wildlife Research Institute (IREC), CSIC-UCLM-JCCM, and Institute for Regional Development (IDR), Ciencia y Tecnología Agroforestal, ETSIA, University of Castilla-La Mancha, Albacete, Spain. felipe.martinez@uclm.esNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Comparative Study
Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

19028926

Citation

Martínez-Pastor, Felipe, et al. "Reactive Oxygen Species Generators Affect Quality Parameters and Apoptosis Markers Differently in Red Deer Spermatozoa." Reproduction (Cambridge, England), vol. 137, no. 2, 2009, pp. 225-35.
Martínez-Pastor F, Aisen E, Fernández-Santos MR, et al. Reactive oxygen species generators affect quality parameters and apoptosis markers differently in red deer spermatozoa. Reproduction. 2009;137(2):225-35.
Martínez-Pastor, F., Aisen, E., Fernández-Santos, M. R., Esteso, M. C., Maroto-Morales, A., García-Alvarez, O., & Garde, J. J. (2009). Reactive oxygen species generators affect quality parameters and apoptosis markers differently in red deer spermatozoa. Reproduction (Cambridge, England), 137(2), pp. 225-35. doi:10.1530/REP-08-0357.
Martínez-Pastor F, et al. Reactive Oxygen Species Generators Affect Quality Parameters and Apoptosis Markers Differently in Red Deer Spermatozoa. Reproduction. 2009;137(2):225-35. PubMed PMID: 19028926.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Reactive oxygen species generators affect quality parameters and apoptosis markers differently in red deer spermatozoa. AU - Martínez-Pastor,Felipe, AU - Aisen,Eduardo, AU - Fernández-Santos,María Rocío, AU - Esteso,Milagros C, AU - Maroto-Morales,Alejandro, AU - García-Alvarez,Olga, AU - Garde,J Julián, Y1 - 2008/11/21/ PY - 2008/11/26/pubmed PY - 2009/3/21/medline PY - 2008/11/26/entrez SP - 225 EP - 35 JF - Reproduction (Cambridge, England) JO - Reproduction VL - 137 IS - 2 N2 - Fe(2)(+)/ascorbate, hydrogen peroxide (H(2)O(2)), and hypoxanthine/xanthine oxidase (XOD) are commonly used for inducing oxidative stress on spermatozoa. A comparative study of these agents was carried out on thawed spermatozoa from red deer. First, we tested a high, medium, and low concentration of each agent: 100, 10, and 1 microM Fe(2)(+) (hydroxyl radical generator); 1 mM, 100, and 10 microM H(2)O(2); and 100, 10, and 1 mU/ml XOD (superoxide and H(2)O(2) generator), incubated at 37 degrees C for 180 min. Intracellular reactive oxygen species (ROS; H(2)DCFDA) increased with dose and time similarly for the three systems at each concentration level. Motility and mitochondrial membrane potential (Deltapsi(m)) were considerably decreased by H(2)O(2) (1 mM and 100 microM) and XOD (100 and 10 mU/ml). Only 1 mM H(2)O(2) reduced viability. The antioxidant Trolox (10 microM) reduced intracellular ROS, but could not prevent the H(2)O(2) or XOD effects. In a second experiment, YO-PRO-1 and M540 were used as apoptotic and membrane stability markers respectively. Only H(2)O(2) increased the proportion of apoptotic and membrane-destabilized spermatozoa. Catalase added to XOD prevented Deltapsi(m) loss, confirming that H(2)O(2) was the causative agent, not superoxide. In a third experiment, caspase activation was tested using the (FAM-VAD-FMK) probe. Viable spermatozoa with activated caspases could be detected in untreated samples, and only H(2)O(2) increased their proportion after 60 min. There were important differences between ROS generators, H(2)O(2) being the most cytotoxic. Although H(2)O(2) and XOD caused Deltapsi(m) dissipation, this was not reflected in increasing apoptotic markers. SN - 1741-7899 UR - https://www.unboundmedicine.com/medline/citation/19028926/Reactive_oxygen_species_generators_affect_quality_parameters_and_apoptosis_markers_differently_in_red_deer_spermatozoa_ L2 - https://rep.bioscientifica.com/doi/10.1530/REP-08-0357 DB - PRIME DP - Unbound Medicine ER -