Evaluation of apoptosis after sperm cryopreservatıon

For long years, different studies were made and different methods were tried to store human sperm specimens that?s to say to form a sperm bank. The aim was to ensure sperm?s continuousness of fertilization capacity with few morphological and physiological damages and this was achieved by cryopreservation partially. However, there is no any standard method for cryopreservation yet and there are lots of questions exist. In our study, we investigated the effect of different cryoprotectants on DNA fragmentation of sperm after freezing and thawing.Totally fifty cases investigated in our study: forty samples of asthenozoospermia with sperm motility defects and ten samples of normozoospermia with no sperm motility defects. Semen samples were macroscopically evaluated for liquefaction, volume, viscosity, appearance and pH values and coagulation. Samples also were evaluated microscopically for motility, sperm concentration and morphology according to the World Health Organization (WHO) criterias. After this stage, the semen samples were separated into two groups. The first group consisted of pre-washed samples. Second group consisted of post-washed with the followed freezing and thawing with two different cryoprotectants (MEDICULT and TEST yolk buffer) samples. In both groups in order to show DNA fragmentation TUNEL assay was performed. Apoptosis was evaluated ultrastructurally by TEM.Results: the total motility of the sperm number was increased in the second group and meaningful decrease seen in the first group. There was no statistical difference between the two cryoprotectan solutions. There was a meaningful statistical difference in the average of DNA fragmentation between the two cryoprotectans group.Conclusions: sperm should be washed before freezing. The evaluation of DNA fragmentation in TUNEL assay which was also supported by ultrastructural findings revealed that there is a difference between the pre- washed and post-washed freezed and thawed samples.