During natural fertilization, mammalian spermatozoa must pass through the zona pellucida before reaching the plasma membrane of the oocyte. 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. Here we present evidence that acrosin, an acrosomal serine protease, plays an essential role in sperm penetration of the zona. We generated acrosin-knockout (KO) hamsters, using an in vivo transfection CRISPR/Cas9 system. Homozygous mutant males were completely sterile. Acrosin-KO spermatozoa ascended the female genital tract and reached ovulated oocytes in the oviduct ampulla, but never fertilized them. In vitro fertilization (IVF) experiments revealed that mutant spermatozoa attached to the zona, but failed to penetrate it. When the zona pellucida was removed before IVF, all oocytes were fertilized. This indicates that in hamsters, acrosin plays an indispensable role in allowing fertilizing spermatozoa to penetrate the zona. 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.
- MeSH
- akrosin genetika metabolismus MeSH
- akrozom metabolismus MeSH
- fertilizace in vitro MeSH
- genový knockout MeSH
- interakce spermie a vajíčka * MeSH
- křečci praví genetika metabolismus MeSH
- spermie enzymologie fyziologie MeSH
- zona pellucida metabolismus MeSH
- zvířata MeSH
- Check Tag
- křečci praví genetika metabolismus MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
BACKGROUND: High-throughput studies provide a wide spectrum of genes for use as predictive markers during testicular sperm extraction (TESE) in combination with ICSI. In this work, we used the specimens from testicular biopsies of men with non-obstructive azoospermia who underwent TESE to investigate the expression of spermatogenesis-related genes MND1, SPATA22, GAPDHS and ACR. METHODS: Testicular biopsy specimens were subdivided into three groups: hypospermatogenesis (HS); maturation arrest (MA); and Sertoli cell-only syndrome (SCO). The levels of expression of the spermatogenesis-related genes MND1, SPATA22, GAPDHS and ACR in the testes were compared among these three groups using the reverse transcription polymerase chain reaction (RT-PCR) technique. RESULTS: Analysis of the expression of spermatogenic genes in human testes with abnormal spermatogenesis showed different expression patterns in patients from different groups. Fertilization rate for studied set of patients was 66% and pregnancy rate 29%. For HS group fertilization rate was 72% and pregnancy rate 32%, while for MA group fertilization and pregnancy rates were 54% and 26%, respectively. Fertilization rates in relation to the studied genes were uniformly around 70%, pregnancy rates for ACR and GAPDHS genes were surprisingly low at 6% and 8% correspondingly. CONCLUSIONS: Analysis of the expression of genes involved in spermatogenesis can be a fast additional test for the level of spermatogenesis in testicular samples.
- MeSH
- akrosin genetika MeSH
- azoospermie genetika patologie MeSH
- biopsie MeSH
- dospělí MeSH
- fertilizace MeSH
- glyceraldehyd-3-fosfátdehydrogenasy genetika MeSH
- intracytoplazmatické injekce spermie MeSH
- lidé středního věku MeSH
- lidé MeSH
- nemoci varlat genetika patologie MeSH
- odběr spermií MeSH
- oligospermie genetika patologie MeSH
- polymerázová řetězová reakce s reverzní transkripcí MeSH
- proteiny buněčného cyklu genetika MeSH
- Sertoli cell only syndrom genetika patologie MeSH
- spermatogeneze genetika MeSH
- stanovení celkové genové exprese MeSH
- těhotenství MeSH
- testis metabolismus patologie MeSH
- úhrn těhotenství na počet žen v reprodukčním věku MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- těhotenství MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
