Factors affecting the in-vitro development to blastocysts of bovine oocytes matured and fertilized in vitro

Abstract

The effects of media (TCM199 vs. synthetic oviduct fluid, SOF), sera (foetal calf serum, FCS vs. human serum, HS), gas atmosphere (5% CO2 in air vs. 5% CO2, 5% O2 and 90% N2) and coculture with bovine oviduct epithelial cells (cells vs. no cells) on the in-vitro development of in-vitro matured and fertilized bovine oocytes were examined. Immature oocytes surrounded with compacted cumulus cells were cultured for 24 h in TCM199 supplemented with 10% FCS, 10 micrograms follicle-stimulating hormone (FSH)/ml and 10 micrograms luteinizing hormone (LH)/ml, 1 microgram oestradiol/ml, and 1 x 10(6) granulosa cells at 39 degrees C under 5% CO2 in air. In-vitro fertilization was performed with frozen-thawed, heparin-treated (100 micrograms/ml, 15 min) spermatozoa from 2 bulls. Oocytes were incubated with 2.5 x 10(6) spermatozoa/ml for 24 h and then cultured in one of 16 treatments for 7 days. Cleavage (2-8-cell) and development to blastocysts were recorded on Days 2 and 7, respectively, after the start of culture. SOF was superior to TCM199 for cleavage (P less than 0.01), development to blastocysts (P less than 0.001) and for proportion of cultured ova resulting in blastocysts with at least 60 or at least 100 nuclei (P less than 0.001). FCS was superior to HS for development to blastocysts (P less than 0.001) and 5% oxygen was superior to air for the proportion of ova reaching at least 60 cells (P less than 0.01). For cleavage and development to blastocysts, there was an interaction between serum and cells (P less than 0.01). In the presence of cells, ova preferred FCS, in their absence, serum had little effect.(ABSTRACT TRUNCATED AT 250 WORDS)

The Use of Embryonic Stem Cell Derived Bioactive Material as a New Protein Supplement for the In Vitro Culture of Bovine Embryos

“…In this study, we used hES-BM recovered from a permanent hES cell line has prolonged growing potential, and so, we mentioned that our source is very stable. On the other hand, other teams have reported that a chemically defined medium (such as SOF) is very attractive for culture of bovine preimplantation embryos, although the percentages of developing embryos are lower than our results [20,21].…”

Sterile filtered paraffin oil supports in vitro developmental competence in bovine embryos comparable to co-culture

“…Researchers have optimized culture systems for various embryo stages and species (4)(5)(6), thus advancing in vitro culture technology for blastocyst development. The most commonly used culture environment has been the micro drop culture method using oil rather than large scale medium, with the exception of certain specific culture methods.…”

Low O 2 tensions and the prevention of differentiation of hES cells

“…Moreover, anaerobic bacteria are able to colonize the uterine lumen (15,16). Embryos are exceedingly sensitive to oxidative damage (17)(18)(19)(20)(21)(22)(23)(24), and blastocysts produced under low O 2 have significantly more inner cell mass cells than those produced under higher O 2 tensions (25). Accordingly, embryologists generally culture embryos under a low rather than high oxygen atmosphere (17)(18)(19)(20)(21)(22).…”