Human Reproduction, Vol. 16, No. 10, 2148-2153,
October 2001
© 2001 European Society of Human Reproduction and Embryology
Cryopreservation of fractionated, highly motile human spermatozoa: effect on membrane phosphatidylserine externalization and lipid peroxidation
The Jones Institute for Reproductive Medicine, Department of Obstetrics and Gynecology, Eastern Virginia Medical School, Norfolk, Virginia, USA
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.
Key words: cryopreservation/human spermatozoa/lipid peroxidation/phosphatidylserine externalization
1 To whom correspondence should be addressed at: The Jones Institute for Reproductive Medicine, 601 Colley Avenue, Department of Obstetrics and Gynecology, Norfolk, VA, 23507, USA. E-mail: OehninSC{at}evms.edu
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