CENTRIFUGATION OF HUMAN SPERMATOZOA INDUCES SUBLETHAL DAMAGE - SEPARATION OF HUMAN SPERMATOZOA FROM SEMINAL PLASMA BY A DEXTRAN SWIM-UP PROCEDURE WITHOUT CENTRIFUGATION EXTENDS THEIR MOTILE LIFETIME

While washing of human sperm cells by centrifugation and resuspension is a procedure in widespread use, there have been indications that thi s procedure per se may be harmful to the cells. The objective of this study was to investigate this question. To this end, a method for the clean separation of motile human spermatozoa from seminal plasma in th e absence of centrifugation was developed, using a modified swim-up pr ocedure, in which liquefied semen was mixed with an equal volume of 30 mg/ml dextran in medium, and the mixture overlaid with medium contain ing 5 mg/ml bovine serum albumin, forming two discreet layers with sta ble interface. The percentage of motile cells in a given sample was co nsistently >80% immediately after recovery. Damage to the cells was as sessed by loss of motile cells during incubation up to 96 h post-recov ery. Comparison of aliquots of spermatozoa obtained by the dextran swi m-up procedure showed that the aliquot subjected to centrifugation had 4 +/- 3% motile cells after 48 h, while the untreated aliquot had 52 +/- 12%. The aliquots showed no difference 1 h post-recovery. Similar results were obtained with spermatozoa that had been centrifuged in se minal plasma and resuspended in fresh plasma, then recovered by dextra n swim-up. The delayed onset of motility loss in the centrifuged sampl es implies that this treatment induces sublethal damage in the cells. Comparison of the standard swim-up and Percoll gradient methods for sp erm recovery, both of which involve centrifugation steps, showed decli ne in motility of the samples similar to that seen with dextran swim-u p of centrifuged cells. We conclude that centrifugation per se induces sublethal damage in human spermatozoa, independently of treatment met hod, and suggest that recovery methods for human spermatozoa which avo id centrifugation might partially alleviate the damage incurred by the se cells during cryopreservation.