RT info:eu-repo/semantics/article
T1 Reactive oxygen species generators affect quality parameters and apoptosis markers differently in red deer spermatozoa
A1 Martínez Pastor, Felipe
A1 Aisen, Eduardo
A1 Fernández Santos, María Rocío
A1 Esteso, Milagros
A1 Maroto Morales, Alejandro
A1 García Álvarez, Olga
A1 Garde López-Brea, Julián
A2 Biologia Celular
K1 Veterinaria
K1 Apoptosis
K1 Esperma
K1 Ciervo rojo
AB 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+ (hydroxyl radical generator); 1 mM, 100, and 10 μM H2O2; and 100, 10, and 1 mU/ml XOD (superoxide and H2O2 generator), incubated at 37 °C for 180 min. Intracellular reactive oxygen species (ROS; H2DCFDA) increased with dose and time similarly for the three systems at each concentration level. Motility and mitochondrial membrane potential (Δψm) were considerably decreased by H2O2 (1 mM and 100 μM) and XOD (100 and 10 mU/ml). Only 1 mM H2O2 reduced viability. The antioxidant Trolox (10 μM) reduced intracellular ROS, but could not prevent the H2O2 or XOD effects. In a second experiment, YO-PRO-1 and M540 were used as apoptotic and membrane stability markers respectively. Only H2O2 increased the proportion of apoptotic and membrane-destabilized spermatozoa. Catalase added to XOD prevented Δψm loss, confirming that H2O2 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 H2O2 increased their proportion after 60 min. There were important differences between ROS generators, H2O2 being the most cytotoxic. Although H2O2 and XOD caused Δψm dissipation, this was not reflected in increasing apoptotic markers.
PB Bioscientifica
YR 2019
FD 2019-04-22
LK http://hdl.handle.net/10612/10530
UL http://hdl.handle.net/10612/10530
NO Reproduction, 2009, vol. 137, n. 2
NO P. 225–235
DS BULERIA. Repositorio Institucional de la Universidad de León
RD 26-abr-2024