Effect of sperm survival and CTC staining pattern on in vitro fertilization of porcine oocytes.
Polyspermy in pig oocytes fertilized in vitro remains unacceptably high. In this study, we evaluated the effects of gamete coincubation time, and determined if the proportion of capacitated spermatozoa would be predictive of the fertilizing ability of frozen-thawed semen in vitro. Cumulus-oocyte complexes were collected from slaughterhouse prepubertal gilt ovaries and matured in vitro for 44 h in TCM199, with EGF, FSH, cysteamine and follicular fluid. Fertilization was induced with 2 x 10(5) frozen-thawed spermatozoa/ml in TBM. Penetration of oocytes as well as polyspermic fertilization occurred 2 h after insemination. A strong correlation between penetration and polyspermic fertilization rates has been demonstrated, but there was no correlation between the proportion of capacitated spermatozoa, as assessed by chlortetracycline staining, at the time of insemination and fertilization rates. We also compared the results of IVF in three IVF media: TBM, m199 and TALP. Penetration and polyspermy were very different in these three media: 71 +/- 19% and 25 +/- 13% in TBM, 37 +/- 11% and 6 +/- 2% in m199, 10 +/- 2% and 0% in TALP, respectively. Nevertheless, survival of spermatozoa or modifications of the capacitation status were not different in these media after 6 h incubation. We concluded that survival and capacitation characteristics of the semen used for IVF could not be predictive of the IVF results. It seems necessary to act at the oocyte level to control both variability between replicates and the incidence of polyspermy. Improving the spermatozoa penetration blocking system of the oocytes and reducing the number of sperm-binding sites on the zona pellucida (ZP) are our further objectives.
Marchal
R
R
Physiologie de la Reproduction et des Comportements, Nouzilly, France.
Peláez
J
J
Terqui
M
M
Mermillod
P
P
D000818:Animals; D002470:Cell Survival; D002751:Chlortetracycline; D003543:Cysteamine / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005260:Female; D005307:Fertilization in Vitro; D005640:Follicle Stimulating Hormone / Q000494:pharmacology; D015571:Follicular Fluid; D008297:Male; D009865:Oocytes / Q000502:physiology*; D013075:Sperm Capacitation; D013084:Sperm-Ovum Interactions; D013094:Spermatozoa / Q000502:physiology*; D013194:Staining and Labeling; D013552:Swine
D003543:Cysteamine; D004815:Epidermal Growth Factor; D005640:Follicle Stimulating Hormone; D002751:Chlortetracycline