EFFECT OF IN-VITRO INCUBATION ON SPONTANEOUS ACROSOME REACTION IN FRESH AND CRYOPRESERVED HUMAN SPERMATOZOA

Objectives-Capacitation and acrosome reaction are prerequisites for fe rtilization. However, in vitro capacitation is not necessary for an ag onist-induced acrosome reaction. We studied whether in vitro capacitat ion is important in spontaneous acrosome reaction and analyzed how cap acitation before cryopreservation influences the acrosomal status of t hawed spermatozoa. Methods-Semen specimens from normal donors (n = 15) were processed by the swim-up technique and divided into two aliquots . One aliquot was capacitated (capacitation induced) for three hours b y incubation in a modified-BWW medium with 3% HSA at 37 degrees C unde r 5% carbon dioxide. The other aliquot did not receive any treatment. Both aliquots were analyzed by CASA to assess the capacitation status of the spermatozoa and then cryopreserved. Spontaneous acrosome reacti on was assessed by FITC-PNA lectin before and after cryopreservation. Sperm viability was measured using Hoechst-33258 stain. Results-Before freezing, the frequency of spontaneous acrosome reaction was higher i n the capacitation-induced sperm preparation (median, 20.5% [interquar tile range, 17.2-37.8]) than in swim-up-induced specimens (median, 10. 6% [range, 4.8-23.2]; P < .001). The percentage of viable cells showin g acrosome reaction increased after cryopreservation in both swim-up-i nduced specimens (median, 241.4% [interquartile range, 37.1-678.6]; P < .001) and capacitation-induced specimens (median, 48.2% [range, 6.1- 63.3]; P = 0.002). Although this increase was higher in the swim-up-in duced specimens (P = 0.002), frequency of postthaw spontaneous acrosom e reaction was similar in bath groups (P = 0.18). Conclusions-We concl ude that sperm capacitation significantly optimizes the acrosome react ion. However, a small proportion of normal spermatozoa do not require capacitation to undergo spontaneous acrosome reaction in vitro. After cryopreservation, the percentage of spermatozoa that had intact acroso mes was similar in both groups, despite the fact that one aliquot unde rwent prefreeze capacitation. These findings suggest that the acrosome reaction may involve complex mechanism(s) rather than a physiological change induced by capacitation.