Differential preimplantation development of rhesus monkey embryos in serum-supplemented media.

Several media, some augmented with amino acids, have been formulated recently, based on simplex optimization, to support the preimplantation development of mouse embryos. For the highly limited studies on preimplantation development of nonhuman primate embryos, a complex medium (CMRL-1066) has been employed. Our objective was to compare the developmental ability of rhesus monkey embryos in a simple medium containing amino acids, KSOM/AA, with the complex media used previously. Zygotes (99) were recovered following in vitro fertilization (IVF) from six monkeys, allocated to either CMRL or KSOM/AA both containing 10% fetal calf serum (FCS), and monitored daily until reaching the expanded or hatched blastocyst stage. The distribution of cells between the inner cell mass (ICM) and trophectoderm was determined at the end of culture by differential nuclear staining. Although a greater number of embryos cultured in KSOM/AA vs. CMRL developed to the morula stage (80%) and beyond (66% to expanded blastocyst), the differences were not significant. Such embryos in KSOM/AA did, however, develop at a significantly faster rate, on average, reaching the expanded blastocyst stage 26 hr earlier than did embryos cultured in CMRL. KSOM/AA embryos hatched in less time and had a higher percentage (43 vs. 34) of cells allocated to the ICM. These results indicate that a simple medium, KSOM/AA, in the presence of serum, supports the development of rhesus monkey embryos at high efficiency and at a faster rate than that observed for embryos cultured in the complex medium, CMRL-1066.

Developmental potential of embryos produced by in-vitro fertilization from gonadotrophin-releasing hormone antagonist-treated macaques stimulated with recombinant human follicle stimulating hormone alone or in combination with luteinizing hormone.

We previously demonstrated, in luteinizing hormone (LH)-deficient macaques, that follicular growth and maturation occurred with administration of exogenous (recombinant human) follicle stimulating hormone (r-hFSH) alone, and that the oocytes recovered fertilized at a notably higher rate than their counterparts from animals receiving both r-hFSH and r-hLH (Zelinski-Wooten et al., 1995). Here, the developmental potential of embryos produced from animals treated with r-hFSH alone or in combination with r-hLH was evaluated. Embryos (n = 127) were cryopreserved, thawed and either co-cultured on buffalo rat liver cells until the hatched blastocyst stage or transferred to synchronized recipients. Although embryos from each treatment group demonstrated a similar ability to develop to hatched blastocysts with a definitive inner cell mass, a significant difference was seen in cryosurvival (56 versus 78%) and in developmental rate to the hatched blastocyst (12 versus 10 days) between embryos from the r-hFSH alone and the combination group respectively. Pregnancies resulted following oviductal embryo transfers in both groups, with corpus luteum rescue occurring on days 12-16 of the luteal phase. In summary, r-hFSH alone during the pre-ovulatory interval is adequate for the gametogenic events required to produce embryos that develop either in vitro or in vivo; however, exposure to r-hLH may improve embryo viability and the rate of development.

Developmental potential of rhesus monkey embryos produced by in vitro fertilization.

This study was an examination of the developmental potential of in vitro fertilization (IVF)-produced rhesus monkey embryos that were cultured in medium alone or cocultured with various cell types. End points were the quality and yield of embryos attaining the expanded or hatched blastocyst stage. A total of 96 IVF-produced embryos were cryopreserved and thawed, and 90 embryos were considered intact and suitable for culture. These embryos were placed into one of five treatment groups consisting of four different cell supports and medium alone. Two primary cultures (bovine oviductal cells [bOVID] and bovine cumulus cells [bCUM]) and two established cell lines (Vero cells and buffalo rat liver cells [BRL]) were utilized for coculture of embryos. Embryos were cultured for up to 14 days, and growth curves were established for all embryos that expanded and/or hatched. The developmental rate for embryos classified as viable varied substantially; in number of days to reach a given stage, early morulae ranged from Days 3 to 9 post-insemination, morulae from Days 4 to 9, 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 arrested upon thawing or shortly after. Of the remaining embryos classified as viable, developmental efficiencies to the hatched blastocyst stage for the various treatments were 1) bOVID, 33%; 2) bCUM, 15%; 3) Vero cells, 9%; 4) BRL, 45%; and 5) medium alone, 8%.(ABSTRACT TRUNCATED AT 250 WORDS)