Effect of different incubation conditions on phosphatidylserine externalization and motion parameters of purified fractions of highly motile human spermatozoa.

Abstract

The objective of this study was to assess the temporal effects of sperm incubation at body temperature with various amounts of human serum albumin (HSA) on motion parameters and phosphatidylserine externalization, an expression of membrane integrity. Purified sperm populations were prepared by discontinuous gradient separation, incubated at 37 degrees C in 3 different culture conditions (human tubal fluid [HTF] alone, HTF plus 0.3% HSA, and HTF plus 3% HSA) and evaluated at 0, 1, 3, 6, and 24 hours. Annexin V binding was used to monitor membrane translocation of phosphatidylserine and a computer-assisted semen analyzer was used to evaluate motion parameters. All incubation conditions led to a time-dependent, significant decline in sperm motion parameters and an increase in exposure of phosphatidylserine (annexin V+, live cells) to the outer leaflet of the plasma membrane in both patients and donors. Patients had a higher degree of motility loss and externalization of phosphatidylserine than donors. The decline in the percentage of normal cells (annexin V-, live) was greater in HTF alone up to 6 hours, and the decline in the percentages of motile and rapid sperm were greater in HTF alone throughout 24 hours when compared with HSA supplementation. We conclude that prolonged incubation of purified populations of highly motile human spermatozoa at body temperature was associated with significant motility loss and membrane changes as revealed by phosphatidylserine translocation. A higher concentration of HSA resulted in a relative protective effect against such impairments, particularly during the first 6 hours of incubation. Under the experimental conditions tested, significant differences were observed between infertile men and fertile controls.