Redox STAte INDUCeD by hyaluronic Acid in bovine cryopreserved spermatozoa

FILOSA A.; PÉREZ MARCHETTA G. ,; FERNÁNDEZ S; CÓRDOBA M

Jornada; IV Jornadas de INITRA; 2014

Ins. de Investigacion en Reproduccion animal INITRA

Redox STAte INDUCeD by hyaluronic Acid in bovine cryopreserved spermatozoa   FILOSA A, Pérez Marchetta G, Fernandez S, CÓRDOBA, m. Cátedra de Química Biológica, Facultad Ciencias Veterinarias-UBA. Instituto de Investigación y Tecnología en Reproducción Animal Unidad ejecutora UBA-Conicet de Investigaciones en Producción Animal   Oxidative metabolism and variations in cellular redox state are important parameters to study the induction of spermatic processes. Heparin (HP) and hyaluronic acid (HA) are glycosaminoglycans found in the bovine female genital tract. The aim of this work was to study the cellular redox state variations, oxygen consumption and capacitation induction by HP and HA. Motility and vigor were observed by optical microscopy at 37°C. Oxygen consumption was evaluated by polarography. Results were analyzed by ANOVA and Tukey`s test (p<0.05). Cellular respiration of the samples incubated for 60 minutes with HA (7,29 ± 1,86 mL O2/h/108 esp) was significantly lower than results obtained by HP treatment (14,76 ± 2,89 mL O2/h/108 esp). Spectrofluorometric evaluation with 2´7´ dichlorofluorescein diacetate (DCFH-DA) after excitation at 488 nm and emission of green fluorescence at 525 nm was used for the measurement of reactive oxygen species (ROS). Furthermore, DCFH-DA fluorescence, attributed to H2O2, was determined by flow citometry (BD FacsCanto II flow cytometer). Sperm viability was analyzed by Propidium Iodide (PI) treatment, a fluorescent stain which only penetrates dead cell membranes and binds to DNA. A ROS production decrease (10%), analyzed by spectrofluorometrically, was observed in samples treated with HA respect to controls and (p<0.05). Samples treated with HA and evaluated by flow citometry produced less ROS (459,88± 122,65) than the controls (909,77± 150,21) (p<0.05). No significant differences in viability were found among treatments with HA, HP and the controls (p>0.05). HA causes a reduction in mitochondrial respiration and ROS production, which can lead to in vitro capacitation of cryopreserved spermatozoa.