Expression of genes regulating chromosome segregation, the cell cycle and apoptosis during human preimplantation development.

BACKGROUND: Appropriate gene expression is vital for the regulation of developmental processes. Despite this fact there is a remarkable paucity of information concerning gene activity during preimplantation development. METHODS: We employed reverse transcription and real-time fluorescent PCR to quantify the expression of nine genes (BRCA1, BRCA2, ATM, TP53, RB1, MAD2, BUB1, APC and beta-actin) in oocytes and embryos. A full characterization of all genes was achieved in 42 embryos and four oocytes. The genes analysed have a variety of important cellular functions. RESULTS: Oocytes displayed relatively high levels of mRNA transcripts, while 2-3-cell embryos were seen to contain very little mRNA from any of the genes examined. Recovery of expression levels was not seen until the 4-cell stage or later, with the presumptive activation of the embryonic genome. Some genes displayed sharp increases in expression in embryos composed of 4-8 cells, but, for most, maximum expression was not achieved until the blastocyst stage. CONCLUSIONS: Our data show that it is possible to define characteristic gene expression profiles for each stage of human preimplantation development. The identification of genes active at defined preimplantation phases may provide clues to the cellular pathways utilized at specific stages of development. Expression of genes that function in DNA repair pathways indicate that DNA damage may be common at the cleavage stage. We suggest that specific patterns of gene expression may be indicative of embryo implantation potential.

Aneuploidy study of human oocytes first polar body comparative genomic hybridization and metaphase II fluorescence in situ hybridization analysis.

BACKGROUND: The object of this study was to determine the mechanisms that produce aneuploidy in oocytes and establish which chromosomes are more prone to aneuploidy. METHODS: A total of 54 oocytes from 36 women were analysed. The whole chromosome complement of the first polar body (1PB) was analysed by comparative genomic hybridization (CGH), while the corresponding metaphase II (MII) oocyte was analysed by fluorescence in situ hybridization (FISH) to confirm the results. RESULTS: Matched CGH-FISH results were obtained in 42 1PB-MII doublets, of which 37 (88.1%) showed reciprocal results. The aneuploidy rate was 57.1%. Two-thirds of the aneuploidy events were chromatid abnormalities. Interestingly, the chromosomes more frequently involved in aneuploidy were chromosomes 1, 4 and 22 followed by chromosome 16. In general, small chromosomes (those equal to or smaller in size than chromosome 13) were more prone to aneuploidy (chi2-test, P=0.07); 25% of the aneuploid doublets would have been misdiagnosed as normal using FISH with probes for nine-chromosomes. CONCLUSIONS: The combination of two different techniques, CGH and FISH, for the study of 1PB and MII allowed the identification and confirmation of any numerical chromosome abnormality, as well as helping to determine the mechanisms involved in the genesis of maternal aneuploidy.