Membranous and structural damage that occur during cryopreservation of human sperm may be time-related events

ObjectiveTo evaluate sperm cryopreservation-induced injuries using sperm plasma membrane protein P34H and α-tubulin as two different subcellular compartment markers.DesignProspective experimental study.SettingAcademic hospital research center and fertility clinic.Patient(s)Semen samples obtained from healthy donors attending the fertility clinic. Sperm samples were either directly processed for sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot experiments (control group) or cryopreserved in liquid nitrogen for different periods of time before being analyzed.Intervention(s)SDS-PAGE, Western blotting, and densitometric quantification of P34H and α-tubulin before and after cryopreservation.Main Outcome Measure(s)Changes in protein quantification between the different groups as a result of sperm cryopreservation.Result(s)In the 31 sperm samples processed for P34H evaluation, a 50% decrease is observed after sperm cryopreservation as compared to the control group. The α-tubulin immunoblotting of 41 sperm samples revealed a 200% increase in the protein detection in the group of cryopreserved sperm as compared to the control group. Contrary to the P34H detection, this change in α-tubulin immunodetected levels appears to be related to the cryopreservation period as it increases during storage.Conclusion(s)These findings indicate that cryopreservation of human semen induces damages at different cellular levels. Moreover, some cryoinjuries are immediate although others seem to take place over time stored in liquid nitrogen.