Studies on Cytogenetic and Morphological Assessment of Bovine Embryos Fertilized in Vitro

Cytogenetic and morphological study was carried out to assess the quality of bovine embryos fertilized in vitro. The major results were summarized as follows. 1) An optimal condition for the chromosome preparation of bovine preimplantation embryos was established using 2-cell embryos fertilized in vitro. 2) Chromosomal anomalies were observed in 7.1-36.4% of 89 two- to 32-cell embryos and these anomalies were caused by abnormal fertilization, especially by polyspermy, and abnormal cleavage. 3) At blastocyst stage, the incidence (18.6%) of chromosomal anomalies in the inner cell mass (ICM) separated from trophectoderm cells by immunosurgery was significantly lower than that (44.2%) in whole blastocyst. 4) Anti-bovine spleen cells rabbit serum was prepared and the method to count the cell numbers of ICM and trophectoderm of bovine blastocysts separately was developed by a differential fluorochrome staining technique using this antiserum. 5) Using this technique, it was found that the cell-cell contacts of ICM cells in blastocysts derived from in vitro fertilization were tighter than those from in vitro fertilization followed by culture in vitro and in a rabbit oviduct. 6) The delineation of each blastomere of ICM cells in blastocysts derived from in vitro fertilization of in vitro matured oocytes was improved by the transfer of the embryos to a rabbit oviduct from the 4-cell stage. 7) The dead ICM cells were observed in ICM cells from survived blastocysts after freezing and thawing by three-step or one-step methods. These findings show that the low developmental ability of bovine embryos fertilized in vitro is caused by an abnormal fertilization and their low pregnancy rates may be caused by the reduced cell proliferation and the loose cell-cell contact of ICM cells.