DEVELOPMENTAL POTENTIAL OF RHESUS-MONKEY EMBRYOS PRODUCED BY IN-VITROFERTILIZATION

This study was an examination of the developmental potential of in vit ro fertilization (IVF)-produced rhesus monkey embryos that were cultur ed in medium alone or cocultured with various cell types. End points w ere the quality and yield of embryos attaining the expanded or hatched blastocyst stage. A total of 96 IVF-produced embryos were cryopreserv ed and thawed, and 90 embryos were considered intact and suitable for culture. These embryos were placed into one of five treatment groups c onsisting of four different cell supports and medium alone. Two primar y cultures (bovine oviductal cells [bOVID] and bovine cumulus cells [b CUM])) and two established cell lines (Vero cells and buffalo rat live r cells [BRL]) were utilized for coculture of embryos. Embryos were cu ltured for up to 14 days, and growth curves were established for all e mbryos that expanded and/or hatched. The developmental rate for embryo s classified as viable varied substantially; in number of days to reac h a given stage, early morulae ranged from Days 3 to 9 post-inseminati on, morulae from Days 4 to 3, blastocysts from Days 6 to 11, expanded blastocysts from Days 7 to 12, and hatched blastocysts from Days 9 to 15. On the basis of developmental curves, 30% of the embryos were arre sted upon thawing or shortly after. Of the remaining embryos classifie d as viable, developmental efficiencies to the hatched blastocyst stag e for the various treatments were 1) bOVID, 33%; 2) bCUM, 15%; 3) Vero cells, 9%; 4) BRL, 45%; and 5) medium alone, 8%. The nuclei of the in ner cell mass (ICM) and trophoblastic cells of those embryos that hatc hed (14 of 90) were differentially stained and cell counts were determ ined. The total cell number per embryo ranged from 177 to 1674, with t he percentage of ICM per embryo varying from 12 to 68%. While some coc ulture treatments produced a significantly greater number of expanded and hatched blastocysts compared to medium alone, there was no correla tion between cell counts of hatched blastocysts and any treatment grou p. Therefore, while cocultures improved the developmental efficiency o f rhesus monkey embryos compared to culture in medium alone, the major ity of embryos that reached the hatched blastocyst stage in all treatm ents showed a well-developed ICM, suggesting that embryo quality was n ot improved with coculture. In conclusion, BRL were found to be benefi cial for the development of rhesus monkey hatched blastocysts of good quality while providing the convenience of a rigorously controlled est ablished cell line for coculturing.