Cryopreservation-thawing of fractionated human spermatozoa and plasma membrane translocation of phosphatidylserine

Abstract

Objective: [1] To evaluate sperm membrane damage during cryopreservation-thawing by the assessment of phosphatidylserine (PS) translocation and [2] to examine the relationship between reactive oxygen species (ROS) and cryopreservation-related alterations. Design: Prospective cohort study. Setting: University-based center. Patient(s): Men consulting for infertility and fertile donors (controls). Intervention(s): Semen processing was performed by density gradient separation followed by cryopreservation and thawing. Main Outcome Measure(s): Membrane PS translocation was evaluated with annexin V binding, generation of ROS was detected by chemiluminescence, and motion parameters were assessed by computer analysis. Result(s): Annexin V binding was detected in the prefreeze fractions with high and low sperm motility. In the patient group, there were significantly higher postthaw levels of annexin V binding in both fractions when compared with prefreezing values. However, such induction of PS translocation was significantly higher in the fractions with high sperm motility. Significantly higher ROS levels were detected in prefreeze samples of the fractions with low sperm motility. Conclusion(s): In the population of men studied, [1] cryopreservation-thawing was associated with induction of membrane PS translocation; [2] postthaw ROS levels were lower than before freezing; and [3] neither annexin V binding results nor the generation of ROS were able to accurately predict sperm cryosurvival rates.