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Abstract
Capacitation of spermatozoa is an essential procedure for fertilization. Capacitated spermatozoa have an in crease in the intracellular cAMP and acrosome reaction (AR) occurs immediately. The effect of exogenous superoxide anion (O−2) on the level of intracellular c AMP and the percentages of both spontaneous AR and lysophosphatidylcholine-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°C in Ham's F-10 medium with O−2 generation system: xanthine + xanthine oxidase + catalase + diethylenetriaminepentaacetic acid + sodium formate. The intracellular cAMP was deter mined by (3H)— cAMP radioimmunoassay 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 effects of SOD with different concentration were also determined. The results showed: the level of intracellular CAMP (pmol/108 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 dependently. To investigate the source of O−2 during in vivo sperm capacitation, female genital tract fluids collected from 6 healthy nonpregnant donors of reproductive age, and seminal plasma, capacitated and noncapacitated spermatozoa from 10 fertile volunteers were investigated by spin trapping method. The results showed: A typical electron paramagnetic resonance spectrum for O−2 spin adduct was exhibited only in capacitated spermatozoa but not in vaginal or cervical secretions, uterine and fallopian tubalfluids, nor in seminal plasma and noncapacitated spermatozoa. These results suggested that only capacitated spermatozoa themselves are able to generate O−2 which stimulated their capacitation in turn. Furthermore, on the basis of these data, we pro pose that it may be possible to utilize the inhibitory effect of SOD on sperm capacitation so as to regulate fertilization.