PROMOTION OF HUMAN SPERM CAPACITATION BY SUPEROXIDE ANION

Capacitation of spermatozoa is an essential procedure for fertilizatio n. Capacitated spermatozoa have an increase in the intracellular cAMP and acrosome reaction (AR) occurs immediately. The effect of exogenous superoxide anion (O(-)2) on the level of intracellular cAMP and the p ercentages of both spontaneous AR and lysoyhosphatidylcholine-induced AR (LPC-AR) were studied using semen samples collected from 10 healthy and fertile volunteers working or studying in Lanzhou Medical College . Spermatozoa were separated by Percoll and incubated at 37 degrees C in Ham's F-10 medium with O(-)2 generation system: xanthine + xanthine oxidase + catalase + diethylenetriaminepentaacetic acid + sodium form ate. The intracellular cAMP was determined by (H-3) - cAMP radioimmuno assay at 3 h of incubation, and the percentages of AR and LPC-AR were evaluated by the triple-stain technique at 3.5 h of incubation. The ef fects of SOD with different concentration were also determined. The re sults showed: the level of intracellular cAMP (pmol/10(8) spermatozoa) of spermatozoa increased from 14.0 +/- 1.3 to 23.2 +/- 2.5 (P < 0.01) , and the percentages of AR and LPC-AR increased from 4.5 +/- 1.1% and 14 +/- 1.9% to 16 +/- 2.0% and 32.5 +/- 1.7%, respectively (P < 0.01 in both comparisons). SOD inhibited these processes concentration depe ndently. To investigate the source of O(-)2 during in vivo sperm capac itation, female genital tract fluids collected from 6 healthy nonpregn ant donors of reproductive age, and seminal plasma, capacitated and no ncapacitated spermatozoa from 10 fertile volunteers were investigated by spin trapping method. The results showed: A typical electron parama gnetic resonance spectrum for O(-)2 spin adduct was exhibited only in capacitated spermatozoa but not in vaginal or cervical secretions, ute rine and fallopian tubal fluids, nor in seminal plasma and noncapacita ted spermatozoa. These results suggested that only capacitated spermat ozoa themselves are able to generate O(-)2 which stimulated their capa citation in turn. Furthermore, on the basis of these data, we propose that it may be possible to utilize the inhibitory effect of SOD on spe rm capacitation so as to regulate fertilization.