cDNA microarray analysis of bovine embryo gene expression profiles during the pre-implantation period

Reproductive Biology and Endocrinology - Tập 2 - Trang 1-16 - 2004
Koichi Ushizawa1, Chandana B Herath1, Kanako Kaneyama1, Satoshi Shiojima2, Akira Hirasawa2, Toru Takahashi1, Kei Imai1,3, Kazuhiko Ochiai4, Tomoyuki Tokunaga5, Yukio Tsunoda6, Gozoh Tsujimoto2, Kazuyoshi Hashizume1,4
1Reproductive Biology and Technology Laboratory,, Developmental Biology Department, National Institute of Agrobiological Sciences, Tsukuba, Japan
2Department of Genomic Drug Discovery Science, Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Japan
3Department of Technology,, National Livestock Breeding Center, Odakura, Japan
4Department of Veterinary Medicine,, Faculty of Agriculture, Iwate University, Ueda, Japan
5Development and Differentiation Laboratory,, Developmental Biology Department, National Institute of Agrobiological Sciences,, Tsukuba, Japan
6Laboratory of Animal Reproduction,, College of Agriculture, Kinki University, Nakamachi, Japan

Tóm tắt

After fertilization, embryo development involves differentiation, as well as development of the fetal body and extra-embryonic tissues until the moment of implantation. During this period various cellular and molecular changes take place with a genetic origin, e.g. the elongation of embryonic tissues, cell-cell contact between the mother and the embryo and placentation. To identify genetic profiles and search for new candidate molecules involved during this period, embryonic gene expression was analyzed with a custom designed utero-placental complementary DNA (cDNA) microarray. Bovine embryos on days 7, 14 and 21, extra-embryonic membranes on day 28 and fetuses on days 28 were collected to represent early embryo, elongating embryo, pre-implantation embryo, post-implantation extra-embryonic membrane and fetus, respectively. Gene expression at these different time points was analyzed using our cDNA microarray. Two clustering algorithms such as k-means and hierarchical clustering methods identified the expression patterns of differentially expressed genes across pre-implantation period. Novel candidate genes were confirmed by real-time RT-PCR. In total, 1,773 individual genes were analyzed by complete k-means clustering. Comparison of day 7 and day 14 revealed most genes increased during this period, and a small number of genes exhibiting altered expression decreased as gestation progressed. Clustering analysis demonstrated that trophoblast-cell-specific molecules such as placental lactogens (PLs), prolactin-related proteins (PRPs), interferon-tau, and adhesion molecules apparently all play pivotal roles in the preparation needed for implantation, since their expression was remarkably enhanced during the pre-implantation period. The hierarchical clustering analysis and RT-PCR data revealed new functional roles for certain known genes (dickkopf-1, NPM, etc) as well as novel candidate genes (AW464053, AW465434, AW462349, AW485575) related to already established trophoblast-specific genes such as PLs and PRPs. A large number of genes in extra-embryonic membrane increased up to implantation and these profiles provide information fundamental to an understanding of extra-embryonic membrane differentiation and development. Genes in significant expression suggest novel molecules in trophoblast differentiation.

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