Improvement of transgenic cloning efficiencies by culturing recipient oocytes and donor cells with antioxidant vitamins in cattle.
The present study was conducted to investigate effects of antioxidants during maturation culture of recipient oocytes and/or culture of gene-transfected donor cells on the meiotic competence of recipient oocytes, and the developmental competence and quality of the reconstructed embryos after nuclear transfer (NT) in cattle. Gene-transfected donor cells had negative effects on the proportions of blastocyst formation, total cell numbers, and DNA fragmentation indices of reconstructed embryos. Supplementation of either vitamin E (alpha-tocopherol: 100 microM) or vitamin C (ascorbic acid: 100 microM) during maturation culture significantly enhanced the cytoplasmic maturation of oocytes and subsequent development of embryos reconstructed with the oocytes and gene-transfected donor cells, but did not have synergistic effects. The supplementation of vitamin E during maturation culture of recipient oocytes increased the proportions of fusion and blastocyst formation of gene-transfected NT embryos, in which the proportions were similar to those of nontransfected NT embryos. When the gene-transfected donor cells that had been cultured with 0, 50, or 100 microM of vitamin E were transferred into recipient oocytes matured with vitamin E (100 microM), 50 microM of vitamin E increased the proportion of blastocyst formation and reduced the index of DNA fragmentation of blastocysts. In conclusion, gene-transfected donor cells have negatively influenced the NT outcome. Supplementation of vitamin E during both recipient oocyte maturation and donor cell culture enhanced the blastocyst formation and efficiently blocked DNA damage in transgenic NT embryos.
Wongsrikeao
Pimprapar
P
Laboratory of Animal Reproduction, The United Graduate School of Veterinary Science, Yamaguchi University, Yoshida, Yamaguchi, Japan.
Nagai
Takashi
T
Agung
Budiyanto
B
Taniguchi
Masayasu
M
Kunishi
Miho
M
Suto
Shizuyo
S
Otoi
Takeshige
T
D000818:Animals; D030801:Animals, Genetically Modified; D000975:Antioxidants / Q000494:pharmacology; D001205:Ascorbic Acid / Q000494:pharmacology*; D002417:Cattle / Q000235:genetics*; D002478:Cells, Cultured; D019976:Cloning, Organism / Q000379:methods*; D004338:Drug Combinations; D046149:Embryo Culture Techniques / Q000379:methods; D005260:Female; D005978:Glutathione / Q000032:analysis; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000737:chemistry / Q000187:drug effects*; D014801:Vitamin A / Q000494:pharmacology*
D000975:Antioxidants; D004338:Drug Combinations; D014801:Vitamin A; D005978:Glutathione; D001205:Ascorbic Acid