Sustancias Oxígeno Reactivas (ROS) en Semen Congelado-Descongelado de Porcino

Abstract

The ROS generation was measured by flow citometry in thawed sperm samples incubated without (basal levels) or with (induced levels) a ROS inductor (1 mM tert-butyl hydroperoxide) for 30 min at 39 ºC and 5% CO2. Sperm cells were simultaneously stained with 2�, 7�-dichlorodihydrofluorescein diacetate, acetil ester (1 mM, CMH 2DCFDA), to estimate the production of ROS, and propidium iodide (1.5 mM) to exclude dead sperm from the analysis. The ejaculates from nine boars were frozen with 3% of glycerol and warmed at ~1200 or ~1800ºC min-1. The ROS production was measured at 0, 60, 120, 240 y 360 min in sperm samples hold at ~21-23 ºC (not incubated) or at 39 ºC and 5% CO2 (incubated) over time. Warming rate had not influence (P>0.05) on ROS production. ROS generation was constant (P>0.05) over time in not incubated samples, but it showed a progressive increase in incubated samples, being it significant (P<0.05) from the 120 min in basal levels or 60 min of incubation in induced levels. Significant (P>0.01) ejaculate/boar variability was evident in both basal and induced ROS production in the incubated sperm samples. Both basal and induced ROS production were significantly (P<0.01) correlated with the percentages of total and rapid progressive, motile and viable spermatozoa. The technique is of great utility to evaluate functional capacity in frozen-thawed sperms; however, additional studies are required to standardize the same one and to establish indicative thresholds of sperm quality loss.