Effect of Embryo Density on In Vitro Development and Gene Expression in Bovine In Vitro-fertilized Embryos Cultured in a Microwell System

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  • SUGIMURA Satoshi
    National Livestock Breeding Center, Fukushima 961-8511, Japan The Robinson Institute, Research Centre for Reproductive Health, School of Paediatrics and Reproductive Health, The University of Adelaide, Adelaide SA 5005, Australia
  • AKAI Tomonori
    Dai Nippon Printing Co., Ltd., Chiba 277-0871, Japan
  • HASHIYADA Yutaka
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • AIKAWA Yoshio
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • OHTAKE Masaki
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • MATSUDA Hideo
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • KOBAYASHI Shuji
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • KOBAYASHI Eiji
    National Livestock Breeding Center, Fukushima 961-8511, Japan NARO Institute of Livestock and Grassland Science, Ibaraki 305-0901, Japan
  • KONISHI Kazuyuki
    National Livestock Breeding Center, Fukushima 961-8511, Japan
  • IMAI Kei
    National Livestock Breeding Center, Fukushima 961-8511, Japan

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タイトル別名
  • Effect of Embryo Density on <i>In Vitro</i> Development and Gene Expression in Bovine <i>In Vitro</i>-fertilized Embryos Cultured in a Microwell System

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To identify embryos individually during in vitro development, we previously developed the well-of-the-well (WOW) dish, which contains 25 microwells. Here we investigated the effect of embryo density (the number of embryos per volume of medium) on in vitro development and gene expression of bovine in vitro-fertilized embryos cultured in WOW dishes. Using both conventional droplet and WOW culture formats, 5, 15, and 25 bovine embryos were cultured in 125 μl medium for 168 h. The blastocysts at Day 7 were analyzed for number of cells and expression of ten genes (CDX2, IFN-tau, PLAC8, NANOG, OCT4, SOX2, AKR1B1, ATP5A1, GLUT1 and IGF2R). In droplet culture, the rates of formation of >4-cell cleavage embryos and blastocysts were significantly lower in embryos cultured at 5 embryos per droplet than in those cultured at 15 or 25 embryos per droplet, but not in WOW culture. In both droplet and WOW culture, developmental kinetics and blastocyst cell numbers did not differ among any groups. IFN-tau expression in embryos cultured at 25 embryos per droplet was significantly higher than in those cultured at 15 embryos per droplet and in artificial insemination (AI)-derived blastocysts. Moreover, IGF2R expression was significantly lower in the 25-embryo group than in the 5-embryo group and in AI-derived blastocysts. In WOW culture, these expressions were not affected by embryo density and were similar to those in AI-derived blastocysts. These results suggest that, as compared with conventional droplet culture, in vitro development and expression of IFN-tau and IGF2R in the microwell system may be insensitive to embryo density.

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