Faculty Bibliography

In human embryos, blastomeres differentiate into trophectoderm (TE) cells and inner cell mass (ICM) cells of blastocysts. Although morphologically indistinguishable, blastomeres at early cleavage stages are likely to undergo changes on a molecular level that make them destined to become ICM or TE cells. While the transcription factor Oct-4 might serve as a marker for totipotent ICM cells, human chorionic gonadotrophin might be used as the equivalent for TE cells. This study reports a reverse transcription-polymerase chain reaction procedure to assess human beta-HCG mRNA concentrations as well as ploidy in individual blastomeres from normally and abnormally fertilized human embryos. beta-HCG mRNA was detected in both euploid and aneuploid cells and in oocytes. Surprisingly, beta-LH mRNA was also detected in some euploid blastomeres. In regard to preimplantation genetic diagnosis, assessment of expression levels of beta-HCG and Oct-4 mRNA in individual biopsied cells might serve as a tool to identify embryogenic blastomeres in combination with testing for chromosome and single gene abnormalities

BACKGROUND: We assessed the maturational competence and the chromosomal pattern of mouse oocytes reconstructed by germinal vesicle (GV) transfer technique using nuclear and/or cytoplasmic components from cryopreserved GV stage oocytes. METHODS: From 657 GV oocytes (326 fresh and 331 frozen/thawed), four groups of reconstructed oocytes were obtained by micromanipulation and electrofusion: fresh GV-fresh cytoplast (FF), thawed GV-thawed cytoplast (TT), fresh GV-thawed cytoplast (FT), thawed GV-fresh cytoplast (TF). All reconstructed oocytes were cultured in vitro to metaphase II. RESULTS: Survival rate after manipulation and electrofusion, as well as progression to metaphase II, did not differ significantly among the four groups. Comparing reconstructed oocytes with fresh and thawed control pools, the only difference was a slightly but significantly higher maturation rate in the TT pool versus matched controls (P < 0.01). Cytogenetic analysis of 25 reconstructed oocytes showed the expected number of 20 chromosomes in 88% of them. CONCLUSIONS: We conclude that both nuclear and cytoplasmic components derived from cryopreserved immature oocytes are suitable for GV transfer procedure, and generate chromosomally normal oocytes able to progress to metaphase II in vitro. The possibility of using cryostored immature oocytes as a source of nuclei and cytoplasm could help in applying GV transfer procedure, both in research and clinical settings