A Schuffner1, M Morshedi, S Oehninger. 1. The Jones Institute for Reproductive Medicine, Department of Obstetrics and Gynecology, Eastern Virginia Medical School, Norfolk, VA 23507, USA.
Abstract
INTRODUCTION: This study investigated lipid peroxidation (LPO) and membrane integrity following cryopreservation-thawing. METHODS: Infertile men (study group) and donors (control group) were examined. Purified populations of highly motile spermatozoa were cryopreserved using TEST-yolk buffer and glycerol (TYB-G) followed by quick thaw. LPO was measured by a spectrophotometric assay, with and without a ferrous ion promoter. Annexin V binding was used to assess membrane translocation of phosphatidylserine (PS). RESULTS: Pre-freeze LPO was significantly higher in the study than in the control group (P = 0.03). In both groups, LPO measurements after thawing were significantly higher than the pre-freeze samples not exposed to TYB-G (P = 0.002 and P = 0.001 respectively). However, when the pre-freeze samples with TYB-G were compared with the post-thaw samples (all exposed to TYB-G), these differences were not significant. There was a significant increase in PS externalization following cryopreservation in both groups (P = 0.02 and P = 0.003 respectively). In donors, pre-freeze LPO concentrations had a significant positive correlation with thawed spermatozoa depicting PS externalization (r = 0.77, P = 0.04). CONCLUSION: Although patients had higher basal LPO than donors, LPO did not differ between fresh and cryopreserved-thawed fractionated motile spermatozoa. Freezing-thawing was associated with translocation of PS to the external membrane leaflet.
INTRODUCTION: This study investigated lipid peroxidation (LPO) and membrane integrity following cryopreservation-thawing. METHODS: Infertile men (study group) and donors (control group) were examined. Purified populations of highly motile spermatozoa were cryopreserved using TEST-yolk buffer and glycerol (TYB-G) followed by quick thaw. LPO was measured by a spectrophotometric assay, with and without a ferrous ion promoter. Annexin V binding was used to assess membrane translocation of phosphatidylserine (PS). RESULTS: Pre-freeze LPO was significantly higher in the study than in the control group (P = 0.03). In both groups, LPO measurements after thawing were significantly higher than the pre-freeze samples not exposed to TYB-G (P = 0.002 and P = 0.001 respectively). However, when the pre-freeze samples with TYB-G were compared with the post-thaw samples (all exposed to TYB-G), these differences were not significant. There was a significant increase in PS externalization following cryopreservation in both groups (P = 0.02 and P = 0.003 respectively). In donors, pre-freeze LPO concentrations had a significant positive correlation with thawed spermatozoa depicting PS externalization (r = 0.77, P = 0.04). CONCLUSION: Although patients had higher basal LPO than donors, LPO did not differ between fresh and cryopreserved-thawed fractionated motile spermatozoa. Freezing-thawing was associated with translocation of PS to the external membrane leaflet.