VIABILITY OF ONE-CELL BOVINE EMBRYOS CULTURED IN-VITRO - COMPARISON OF CELL-FREE CULTURE WITH COCULTURE

In this study, the quality (number of cells) and pregnancy rates of bo vine blastocysts produced by in vitro maturation/in vitro fertilizatio n (IVM/IVF) following cultivation in either cell-free culture or co-cu lture were compared. Bovine one-cell IVM/IVF embryos obtained 6h after insemination were stripped of cumulus cells and assigned to either ce ll-free culture or co-culture with granulosa cell monolayers for 9 day s (Expt 1) or 10 days (Expts 2 and 3). In Expt 3, day-7 (day 0=day of insemination) blastocysts, day-8 expanded blastocysts and day-9 hatche d blastocysts were air-dried, fixed and stained to determine the numbe r of cells. Expanded blastocysts obtained in Expt 1 were cryopreserved using propylene glycol as a cryoprotectant and were used later for em bryo transfer. There were no significant differences between cell-free culture and co-culture in the percentage of one-cell embryos that dev eloped to 2- to 16-cells (66.7% versus 72.4% for Expt 1, 71.0% versus 78.2% for Expt 2). However, significantly more (P < 0.05) of the one-c ell embryos co-cultured with granulosa cell monolayers developed to mo rula, blastocyst and expanded blastocyst stages compared with those in cell-free culture (35.0 versus 27.1%, 25.1 versus 14.2%, 15.6 versus 5.4% for Expt 1; 37.6 versus 24.0%, 25.7 of versus 11.0%, 16.8 versus 3.0% for Expt 2). Only embryos co-cultured with granulosa cell monolay ers hatched (Expt 2). Embryos co-cultured with granulosa cell monolaye rs also had higher (P < 0.01) numbers of cells (92 +/- 42 versus 56 +/ - 21 for blastocysts, 149 +/- 53 versus 81 +/- 29 for expanded blastoc ysts). Pregnancy rates tended to be higher for embryos produced by co- culture compared with those from cell-free culture (40.0 versus 27.3%, respectively). The results suggest that embryos produced in cell-free culture were of poorer quality than those produced in co-culture; how ever, some of them were developmentally competent as confirmed by the births of three calves after transfer to eleven recipient cows.