Mori T, Hirayama M, Suzuki K, Shimizu H, Nagai T
Department of Animal Science, Faculty of Agriculture, Hokkaido University, Sapporo, Japan.
Biol Reprod. 1996 Aug;55(2):364-9. doi: 10.1095/biolreprod55.2.364.
To investigate the promoting effect of casein phospho peptides (CPP) on the penetration of ejaculated boar spermatozoa into in vitro-matured oocytes, preincubated spermatozoa were coincubated with in vitro-matured oocytes in fertilization medium with or without CPP for 60-240 min. Sperm penetration into zona-intact oocytes was observed within 90 min of coincubation. The penetration rate reached a maximum earlier in the presence of CPP than in the absence of CPP. Furthermore, spermatozoa preincubated in fertilization medium with CPP before coincubation with matured oocytes retained the ability to penetrate oocytes for a longer time than spermatozoa preincubated in fertilization medium without CPP. In fertilization medium containing CPP, Ca2+ uptake by spermatozoa increased within 90 min of incubation, with a peak of 113 nm/1 x 10(7) cells. On the other hand, in fertilization medium with no CPP, the intracellular Ca2+ concentration remained constant or gradually decreased until 180 min of incubation. When preincubated spermatozoa were coincubated with oocytes for 5 h in a fertilization medium containing CPP and/or EGTA, or with no addition, EGTA decreased the rate of oocytes penetrated by spermatozoa and the number of spermatozoa penetrating each oocyte. On the other hand, when CPP was added to the medium containing EGTA, the inhibitory effect of EGTA was neutralized. These results indicate that CPP promotes Ca2+ uptake of boar spermatozoa, resulting in a more rapid and longer-lasting shift of a subpopulation of spermatozoa that can penetrate oocytes; the findings also suggest that CPP neutralizes the inhibitory effect of EGTA on sperm penetration.
为研究酪蛋白磷酸肽（CPP）对体外成熟卵母细胞中射精公猪精子穿透的促进作用，将预孵育的精子与体外成熟的卵母细胞在含有或不含有CPP的受精培养基中共同孵育60 - 240分钟。在共同孵育90分钟内观察精子对完整透明带卵母细胞的穿透情况。与不添加CPP相比，添加CPP时精子穿透率更早达到最大值。此外，在与成熟卵母细胞共同孵育前在含有CPP的受精培养基中预孵育的精子，比在不含CPP的受精培养基中预孵育的精子保留穿透卵母细胞能力的时间更长。在含有CPP的受精培养基中，精子在孵育90分钟内Ca²⁺摄取增加，峰值为113 nm/1×10⁷个细胞。另一方面，在不含CPP的受精培养基中，细胞内Ca²⁺浓度在孵育180分钟前保持恒定或逐渐降低。当预孵育的精子在含有CPP和/或乙二醇双乙醚二胺四乙酸（EGTA）或不添加任何物质的受精培养基中与卵母细胞共同孵育5小时时，EGTA降低了精子穿透卵母细胞的比率以及每个卵母细胞被穿透的精子数量。另一方面，当将CPP添加到含有EGTA的培养基中时，EGTA的抑制作用被中和。这些结果表明，CPP促进公猪精子摄取Ca²⁺，导致能够穿透卵母细胞的精子亚群更快且更持久地发生变化；研究结果还表明，CPP中和了EGTA对精子穿透的抑制作用。
