title: Improved survival of vitrified porcine embryos after partial delipation through chemically stimulated lipolysis and inhibition of apoptosis.
abstract: Mechanical removal of intracellular lipids has been the most effective approach to increase the cryosurvival of porcine embryos. In this experiment, we tested the hypotheses that the cryosurvival of porcine embryos can be improved after partial delipation through chemically stimulated lipolysis and that the survival can be further improved by inhibition of apoptosis. Porcine embryos were produced in vitro using sow oocytes. On Day 5 of embryonic development, embryos were cultured in the presence of 10 microM forskolin for 24h. On Day 6 blastocysts were vitrified using an open pulled straw (OPS) method and warmed blastocysts were cultured 18 h for them to recover. A caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (Z-VAD-FMK) was used at 20 microM during vitrification and subsequent culture to inhibit apoptosis. A 2 x 2 x 2 factorial design experiment was conducted to examine the effect of chemical delipation, vitrification and apoptosis inhibition. We also measured the lipolytic activity of porcine embryos cultured with or without forskolin. Chemical delipation increased the cryosurvival of porcine embryos compared to the controls (71.2+/-2.8% versus 37.1+/-5.1%). Apoptosis inhibition increased the ability of blastocysts to fully recover (23.8+/-3.1% versus 14.6+/-4.3%). However, there was no interaction between chemical delipation and apoptosis inhibition. Lipolytic agent treatment increased the lipolytic activity of porcine blastocysts. In conclusion, cryosurvival of porcine embryos was improved by partial delipation through chemical stimulation of lipolysis or apoptosis inhibition.
authors.affiliation: Comparative Medicine Center, University of Missouri-Columbia, 1600 E. Rollins Street, Columbia, MO 65211, USA.
authors.forename: Hongsheng
authors.initials: H
authors.lastname: Men
authors.forename: Yuksel
authors.initials: Y
authors.lastname: Agca
authors.forename: Lela K
authors.initials: LK
authors.lastname: Riley
authors.forename: John K
authors.initials: JK
authors.lastname: Critser
mesh_terms: D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D001755:Blastocyst / Q000187:drug effects / Q000502:physiology; D002470:Cell Survival; D005576:Colforsin / Q000494:pharmacology*; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D046149:Embryo Culture Techniques / Q000379:methods / Q000662:veterinary*; D008066:Lipolysis / Q000187:drug effects* / Q000502:physiology; D013552:Swine / Q000196:embryology*
chemical_list: D000590:Amino Acid Chloromethyl Ketones; D015853:Cysteine Proteinase Inhibitors; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D005576:Colforsin