It is possible that mouse spermatozoa are exceptional, in that they do not rely on acrosomal enzymes to penetrate the zona, because sperm acrosin activity is weaker in mice compared with in other mammalian species such as rats and hamsters ( 8). However, in contrast to these findings, acrosin-deficient mouse spermatozoa could pass through the zona, although dispersion of the cumulus oophorus was delayed to some extent ( 7). Indeed, it was reported that anti-acrosin antibodies significantly decreased the incidence of in vivo fertilization in rabbits ( 5), and inhibition of acrosin by soybean trypsin inhibitor prevented human spermatozoa from penetrating the zona ( 6). Of the many acrosomal enzymes, acrosin has been thought to be a major player in this process, because of its strong hydrolyzing activity and widespread distribution in mammals ( 3, 4). A biophysical analysis suggested that the calculated force generated by the sperm alone is not sufficient to penetrate the zona mechanically ( 2). It is generally assumed that spermatozoa penetrate the zona by mechanical force, aided by the acrosomal enzymes that are bound to the inner acrosomal membrane ( 1). Mammalian spermatozoa deposited in the vagina or uterus ascend the female genital tract and penetrate the outer layers of the oocyte (cumulus oophorus and zona pellucida) before reaching its plasma membrane. This study also suggests that the KO hamster system would be a useful model for identifying new gene functions or analyzing human and animal disorders because of its technical facility and reproducibility. This indicates that in hamsters, acrosin plays an indispensable role in allowing fertilizing spermatozoa to penetrate the zona. When the zona pellucida was removed before IVF, all oocytes were fertilized. In vitro fertilization (IVF) experiments revealed that mutant spermatozoa attached to the zona, but failed to penetrate it. Acrosin-KO spermatozoa ascended the female genital tract and reached ovulated oocytes in the oviduct ampulla, but never fertilized them. Homozygous mutant males were completely sterile.
We generated acrosin-knockout (KO) hamsters, using an in vivo transfection CRISPR/Cas9 system. Here we present evidence that acrosin, an acrosomal serine protease, plays an essential role in sperm penetration of the zona. It is assumed that this step involves partial lysis of the zona by sperm acrosomal enzymes, but there has been no unequivocal evidence to support this view. During natural fertilization, mammalian spermatozoa must pass through the zona pellucida before reaching the plasma membrane of the oocyte.
0 kommentar(er)
