Comparison of low and high quality embryos in human embryonic stem cell lines establishment

Abstract

Human embryonic stem cells (hESC) are pluripotent cells derived from inner cell mass (ICM) of human blastocysts. Preimplantation embryos have provided the main source of blastocysts for hESC derivation. General experience suggests that higher quality embryos result in a greater efficiency in hESC production, however few reports provide information on the quality of the embryos used in the derivation process. The present study compared the use of donated frozen/thawed low and high quality embryos for hECS establishment. A prospective study. Fifty three embryos from 11 patients, submitted to controlled ovarian stimulation (COS) for intra cytoplasmic sperm injection (ICSI) were donated for hECS research. COS was conducted according to a standard long GnRH agonist protocol and recombinant-FSH. Embryo transfer was performed on day three and surplus embryos were cryopreserved by slow-freezing method. Embryos with less than 10% fragmentation, from 6 to 10 cells and symmetric blastomeres were considered high quality embryos. Twenty three high and 30 low quality embryos were thawed and incubated until blastocyst stage when the hECS was isolated through mechanical removal and culture of the ICM. Eleven thawed embryos (20.7%) reached the blastocyst stage in vitro, nine out of 23 high quality and two out of 30 low quality embryos (39.1 vs. 6.6%, P=0,002). ICM(s) were removed from all blastocysts and nine cell cultures were established in which three were frozen in early passages for future use. Currently 6 hESC lines are in culture, one derived from two low quality embryos and five derived from nine high quality embryos (50.0% vs. 55.5%, P>0.05). Over the last decade there has been great interest in derivation of hESCs, and besides ethical limitations, donated embryos are likely the major source of hESCs. However, even though there are many cryopreserved embryos in storage, few are available for research. In the present study low and high quality embryos were compared regarding the efficiency in generate hESC lines. It has been discussed whether the long term genetic stability of lines obtained from poorer quality embryos differs from those derived from high quality embryos. We demonstrated that thawed low quality embryos have a decreased development competence until blastocyst stage when compared to high quality embryos. Nevertheless, once the blastocyst stage has been reached, both low and high quality embryos are equally potential sources of hESC lines.