title: Effect of elevated Ca(2+) concentration in fusion/activation medium on the fusion and development of porcine fetal fibroblast nuclear transfer embryos.
abstract: The present study examined the effect of elevated Ca(2+) concentration in fusion/activation medium on the fusion and development of fetal fibroblast nuclear transfer (NT) porcine embryos. Frozen-thawed and serum starved fetal fibroblasts were transferred into the perivitelline space of enucleated oocytes. Cell fusion and activation were induced simultaneously with electric pulses in 0.3 M mannitol-based medium containing 0.1 or 1.0 mM CaCl(2). Some fused embryos were further activated 1 hr after the fusion treatment by exposure to an electric pulse. The NT embryos were cultured in vitro for 6 days. Fusion and blastocyst formation rates were significantly (P<0.05) increased by increasing the Ca(2+) concentration from 0.1 mM (67.1 and 6.3%) to 1.0 mM (84.7 and 15.8%). However, no difference in the number of cells in blastocysts was observed between the two groups. A higher percentage of blastocyst was also observed when control oocytes were parthenogenetically activated in the presence of elevated Ca(2+) (19.3% vs. 32.4%, P<0.05). When the reconstituted oocytes were fused in the medium containing 1.0 mM CaCl(2), increasing the number of pulses from 2 to 3 or an additional activation treatment did not enhance the blastocyst formation rate or cell number in blastocysts. These results demonstrate that increasing the Ca(2+) concentration in the fusion/activation medium can enhance the fusion and blastocyst formation rates of fetal fibroblast NT porcine embryos without an additional activation treatment.
authors.affiliation: Department of Animal Sciences, University of Missouri-Columbia, Columbia, Missouri 65211, USA.
authors.forename: Hee-Tae
authors.initials: HT
authors.lastname: Cheong
authors.forename: Kwang-Wook
authors.initials: KW
authors.lastname: Park
authors.forename: Gi-Sun
authors.initials: GS
authors.lastname: Im
authors.forename: Liangxue
authors.initials: L
authors.lastname: Lai
authors.forename: Qing-Yuan
authors.initials: QY
authors.lastname: Sun
authors.forename: Billy N
authors.initials: BN
authors.lastname: Day
authors.forename: Randall S
authors.initials: RS
authors.lastname: Prather
mesh_terms: D000818:Animals; D002118:Calcium; D002459:Cell Fusion; D019976:Cloning, Organism; D003470:Culture Media / Q000737:chemistry; D004624:Embryo Transfer; D005260:Female; D005347:Fibroblasts / Q000502:physiology* / Q000648:ultrastructure; D051379:Mice; D009865:Oocytes / Q000737:chemistry / Q000502:physiology; D013552:Swine / Q000196:embryology
chemical_list: D003470:Culture Media; D002118:Calcium