CELLULAR EVALUATION OF BOVINE NUCLEAR TRANSFER EMBRYOS DEVELOPED IN-VITRO

Cloned blastocysts developed in vitro for 7 d had a mean number of cel ls (82.86 +/- 5.35) as evaluted by nuclei counting in serial optical s ections using confocal microscopy, after staining with propidium iodid e. This number was not significantly different from that of control IV F embryos cultured under the same conditions during the same period (( X) over bar = 88.89 +/- 7.53). Semi-thin sections revealed that most o f the blastocysts had an inner cell mass (10/12) and a blastocoele. Un der transmission electron microscopy, the trophectoderm appeared well differentiated as a polarized epithelium with apical microvilli and la teral junctions including desmosomes with bound intermediate filaments . The cytoplasm sometimes contained immature mitochondria or a large n umber of residual bodies. About half of the blastocysts examined had a large amount of cellular debris in the perivitelline space or inside the blastocoele cavity. The cloned blastocysts were also able to hatch in vitro by day 8 and SEM indicated a normal morphology of the trophe ctoderm cells with numerous apical microvilli. The high number of excl uded or degenerating cells found in some embryos may partially explain early embryonic mortality that follows transfer. However, these obser vations do not give a clear explanation for the high incidence of feta l losses.