title: Pre-fertilization zona pellucida hardening by different cross-linkers affects IVF in pigs and cattle and improves embryo production in pigs.
abstract: Zona pellucida (ZP) hardening (resistance to proteolysis) has been classically identified as a post-fertilization event that contributes to the block to polyspermy. Di-(N-succinimidyl)-3,3'-dithiodipropionate (DSP), a permeable amine-reactive cross-linker, was recently shown to induce pre-fertilization ZP hardening and to improve porcine IVF productivity. The objectives of this study were to investigate i) how DSP affects pre-fertilization ZP hardening and IVF in cattle, ii) if a non-permeable amine-reactive cross-linker such as bis(sulfosuccinimidyl) suberate (BS3) affects ZP hardening and IVF in cattle and pigs, and iii) whether DSP or BS3, if improvement in IVF productivity was demonstrated in either species, affects in vitro embryo development. Bovine and porcine in vitro matured oocytes were incubated with the cross-linkers (0.06, 0.3, and 0.6 mg/ml) for 30 min. Then they were subjected to ZP digestion or IVF. In cattle, both DSP and BS3 induced ZP hardening and decreased the penetration rate, although monospermy, penetration, or male pronuclear formation was not affected. In pigs, BS3 treatment induced ZP hardening, decreased penetration and male pronuclear formation, and increased monospermy. IVF productivity only improved when porcine oocytes were exposed to DSP. When porcine zygotes derived from this treatment were further cultured in vitro, the cleavage and blastocyst formation rates increased. These results support the idea that mechanisms involved in the prevention of polyspermic fertilization in cattle and pigs have different efficiencies, and ZP hardening induced by DSP cross-linker may be useful for improving porcine embryo production.
authors.affiliation: Department of Physiology, School of Medicine, University of Murcia, Murcia 30071, Spain.
authors.forename: Sebastian
authors.initials: S
authors.lastname: Canovas
authors.forename: Raquel
authors.initials: R
authors.lastname: Romar
authors.forename: Luis Alberto
authors.initials: LA
authors.lastname: Grullon
authors.forename: Manuel
authors.initials: M
authors.lastname: Aviles
authors.forename: Pilar
authors.initials: P
authors.lastname: Coy
mesh_terms: D000818:Animals; D002417:Cattle / Q000502:physiology*; D003432:Cross-Linking Reagents / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D046149:Embryo Culture Techniques; D005260:Female; D005300:Fertility Agents, Female / Q000494:pharmacology*; D005307:Fertilization in Vitro / Q000662:veterinary*; D008297:Male; D009865:Oocytes / Q000187:drug effects* / Q000502:physiology; D011247:Pregnancy; D011402:Pronase / Q000494:pharmacology; D013084:Sperm-Ovum Interactions / Q000187:drug effects; D013388:Succinimides / Q000494:pharmacology; D013552:Swine / Q000502:physiology*; D015044:Zona Pellucida / Q000187:drug effects* / Q000502:physiology
chemical_list: D003432:Cross-Linking Reagents; D005300:Fertility Agents, Female; D013388:Succinimides; C035760:bis(sulfosuccinimidyl)suberate; D011402:Pronase; C011240:dithiobis(succinimidylpropionate)