title: Effect of cryopreservation and in vitro culture of bovine fibroblasts on histone acetylation levels and in vitro development of hand-made cloned embryos.
abstract: In this study, the relative acetylation levels of histone 3 in lysine 9 (H3K9ac) in cultured and cryopreserved bovine fibroblasts was measured and we determined the influence of the epigenetic status of three cultured (C1, C2 and C3) donor cell lines on the in vitro development of reconstructed bovine embryos. Results showed that cryopreservation did not alter the overall acetylation levels of H3K9 in bovine fibroblasts analysed immediately after thawing (frozen/thawed) compared with fibroblasts cultured for a period of time after thawing. However, reduced cleavage rates were noted in embryos reconstructed with fibroblasts used immediately after thawing. Cell passage affects the levels of H3K9ac in bovine fibroblasts, decreasing after P1 and donor cells with lower H3K9ac produced a greater frequency of embryo development to the blastocyst stage. Cryopreservation did not influence the total cell and ICM numbers, or the ICM/TPD ratios of reconstructed embryos. However, the genetic source of donor cells did influence the total number of cells and the trophectoderm cell numbers, and the cell passage influenced the total ICM cell numbers.
authors.affiliation: Audubon Center for Research of Endangered Species, New Orleans, Louisiana, USA.
authors.forename: Liliana
authors.initials: L
authors.lastname: Chacón
authors.forename: Martha C
authors.initials: MC
authors.lastname: Gómez
authors.forename: Jill A
authors.initials: JA
authors.lastname: Jenkins
authors.forename: Staley P
authors.initials: SP
authors.lastname: Leibo
authors.forename: Gemechu
authors.initials: G
authors.lastname: Wirtu
authors.forename: Betsy L
authors.initials: BL
authors.lastname: Dresser
authors.forename: C Earle
authors.initials: CE
authors.lastname: Pope
mesh_terms: D000107:Acetylation; D000818:Animals; D001755:Blastocyst / Q000166:cytology*; D002417:Cattle; D019976:Cloning, Organism; D015925:Cryopreservation; D004622:Embryo, Mammalian / Q000166:cytology; D047108:Embryonic Development; D057890:Epigenomics; D005347:Fibroblasts / Q000166:cytology*; D006657:Histones / Q000378:metabolism*; D053652:Nuclear Transfer Techniques
chemical_list: D006657:Histones