Reactive oxygen species generators affect quality parameters and apoptosis markers differently in red deer spermatozoa.
Fe2+/ascorbate, hydrogen peroxide (H2O2) 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 agen...
| Autores: | , , , , , , |
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| Tipo de recurso: | artículo |
| Fecha de publicación: | 2009 |
| País: | España |
| Institución: | Universidad de Castilla-La Mancha |
| Repositorio: | RUIdeRA. Repositorio Institucional de la UCLM |
| OAI Identifier: | oai:ruidera.uclm.es:10578/7462 |
| Acceso en línea: | http://hdl.handle.net/10578/7462 |
| Access Level: | acceso abierto |
| Palabra clave: | Apoptosis Espermatozoides Mitocondrias Toxicología Ciervos |
| Sumario: | Fe2+/ascorbate, hydrogen peroxide (H2O2) 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 μM Fe2+ (hydroxil radical generator); 1 mM, 100 and 10 μM H2O2; and 100, 10 and 1 mU/mL XOD (superoxide and H2O2 generator), incubating at 37 C for 180 min. Intracellular ROS (H2DCFDA) increased with dose and time, and similarly for the three systems at each concentration level. Motility and mitocondrial membrane potential ( m) were considerably decreased by H2O2 (1 mM and 100 μM) and XOD (100 and 10 mU/mL). Only H2O2 1 mM reduced viability. The antioxidant Trolox (10 μM) reduced intracellular ROS, but could not prevent H2O2 or XOD effects. In a second experiment, we used YO-PRO-1 and M540 as apoptotic and membrane-stability markers, respectively. Only H2O2 increased the proportion of apoptotic and membrane-destabilized spermatozoa. Catalase added to XOD prevented m loss, confirming that H2O2 was the causative agent, not superoxide. In a third experiment, caspase activation was tested using the FLICA probe. Only H2O2 increased the proportion of viable spermatozoa with activated caspases. There were important differences between ROS generators, being H2O2 the most cytotoxic. Although H2O2 and XOD caused m dissipation, it did not reflect on increasing apoptotic markers. We hypothesize that a subpopulation of spermatozoa may lack apoptotic markers. |
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