Effects of human sperm cryopreservation on apoptotic markers in normozoospermic and non-normozoospermic patients

Abstract

The negative effects of cryopreservation on sperm parameters are well documented but little information is known about molecular basis of the process. The aim of the present study was to investigate the possible effects of sperm cryopreservation on main apoptotic signs including DNA fragmentation and caspase-3 activation and to determine if these effects vary according to sperm parameters. Sperm samples of 72 patients were cryopreserved. The patients were sub-grouped as normozoospermic or non-normozoospermic patients according to their semen parameters. DNA fragmentation rates and caspase-3 activation levels were analyzed before and after cryopreservation in both groups. Mean DNA fragmentation rate was increased significantly from 23.98% in neat semen samples to 27.34% after cryopreservation (P = 0.03). DNA fragmentation rates were slightly higher in non-normozoospermic patients compared with the normozoospermic patients in both the neat semen and after cryopreservation (23.25 and 24.71% vs. 26.32 and 28.36%, respectively) although the difference obtained were not statistically significant. An increasing trend for caspase-3 activations (0.093 vs. 0.116) was observed after cryopreservation but the differences were not statistically significant. Caspase-3 activation was found to be slightly higher in non-normozoospermic patients both in the neat semen and after cryopreservation compared with the normozoospermic patients but the differences were not statistically significant. Caspase-3 expression was also shown using immunocytochemistry in both fresh ejaculated sperm and thawed sperm after cryopreservation but at different localizations. The cryopreservation process had detrimental effects on sperm quality but the quality of the sperm samples was not adversely effective for the apoptotic markers including DNA fragmentation and caspase-3 activation patterns. In fact, it was the cryopreservation process itself that adversely effected the above apoptotic markers and apoptosis. It was concluded therefore that sperm cell cryopreservation triggers apoptosis after thawing and this process adversely affects semen parameters.