title: Effect of droplet vitrification on development competence, actin cytoskeletal integrity and gene expression in in vitro cultured mouse embryos.
abstract: The effect of modified droplet vitrification was assessed on cellular actin filament organization, apoptosis related gene expression and development competence in mouse embryos cultured in vitro. Mouse zygotes, 2-cell embryos and morulae were vitrified in ethylene glycol (VS-1) and ethylene glycol plus DMSO (VS-2) and thawed by directly placing the vitrified drop into 0.3M sucrose solution at 37 degrees C. High recovery (93-99%) of morphologically normal embryos was evident following vitrification and thawing. No detectable actin filament disruption was observed in the embryos at any development stage following vitrification and thawing and/or in vitro culture. The expression pattern of Bax, Bcl2 and p53 genes was altered (P<0.05) in vitrified zygotes and 2-cell embryos, but not in morulae. Although a large proportion of the vitrified zygotes (59.5+/-4.4% in VS-1 and 57.9+/-4.5% in VS-2; mean+/-S.E.M.) and 2-cell embryos (63.1+/-4.4% in VS-1 and 59.2+/-4.3% in VS-2) developed into blastocysts, development of control embryos (70.2+/-5.0% of zygotes and 75.5+/-4.4% of 2-cell embryos) into blastocysts was higher (P<0.05). In contrast, development of the control and vitrified morulae into blastocysts (more than 85%) was similar. We concluded that the modified droplet vitrification procedure supported better survival of morula stage compared to zygotes and 2-cell mouse embryos.
authors.affiliation: Department of Dairy Science, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061-0315, USA.
authors.forename: A
authors.initials: A
authors.lastname: Dhali
authors.forename: V M
authors.initials: VM
authors.lastname: Anchamparuthy
authors.forename: S P
authors.initials: SP
authors.lastname: Butler
authors.forename: R E
authors.initials: RE
authors.lastname: Pearson
authors.forename: I K
authors.initials: IK
authors.lastname: Mullarky
authors.forename: F C
authors.initials: FC
authors.lastname: Gwazdauskas
mesh_terms: D000199:Actins / Q000032:analysis; D000818:Animals; D001755:Blastocyst / Q000378:metabolism / Q000502:physiology / Q000648:ultrastructure; D015925:Cryopreservation / Q000379:methods*; D003599:Cytoskeleton / Q000648:ultrastructure; D046149:Embryo Culture Techniques; D047108:Embryonic Development; D005260:Female; D015870:Gene Expression; D016158:Genes, p53 / Q000235:genetics; D051379:Mice; D008813:Mice, Inbred ICR; D018613:Microscopy, Confocal; D009028:Morula / Q000378:metabolism / Q000502:physiology / Q000648:ultrastructure; D011247:Pregnancy; D011518:Proto-Oncogene Proteins / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2; D014021:Tissue Preservation / Q000379:methods*; D015053:Zygote / Q000254:growth & development / Q000378:metabolism / Q000648:ultrastructure; D051028:bcl-2-Associated X Protein / Q000235:genetics
chemical_list: D000199:Actins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; C052617:Bcl2 protein, mouse