CELL ALLOCATION TO THE INNER CELL MASS AND THE TROPHECTODERM IN BOVINE EMBRYOS CULTURED IN 2 DIFFERENT MEDIA

Data from other laboratories have shown that speed of bovine blastocys t development is higher when Menezo B2 is used for coculture compared to TCM199. It was our purpose to investigate whether this early blasto cyst formation was also indicative of embryo quality by studying the a llocation of inner cells in embryos generated by B2-coculture and by T CM199-coculture. For this purpose, a differential staining technique w as used. General embryo development was similar for TCM 199- and B2-em bryos expressed as rate of cleavage at day 3 and morula-blastocyst for mation at day 8 (P > 0.05), but significantly different when expressed as number of eight-cell stages at day 3 and expanded or hatched blast ocysts at day 8 (P < 0.01). B2-embryos cultured until day 5, 6, and 7 post insemination, had total cell numbers of 24, 65, and 109 respectiv ely, which was significantly higher than the cell number of TCM199-emb ryos cultured over the same time period (18, 41, and 71 respectively, P < 0.001). Morphological differentiation was significantly more advan ced for B2-embryos at day 7 and 8 (P < 0.0001 and P < 0.001, respectiv ely). First presumptive inner cells appeared in eight- to 16-cell stag es at day 3. Because the determination of inner cells by differential staining is depending upon the presence of functional tight junctions, we concluded that the establishment of the tight junction seal in B2- embryos differed from that in TCM 199-embryos: Inner cells appeared 0. 56 cell cycle later in B2-embryos (P < 0.001) and a larger variation e xisted in the number of ICM-cells in B2-blastocysts (P < 0.001). The h igher total cell number of B2-expanded blastocysts was mainly acquired by trophectoderm growth (P < 0.06). These data indicate that the appa rent better quality of B2-embryos (faster cleavage, earlier blastocyst formation) is not reflected in a reliable number of inner cells of B2 -blastocysts. (C) 1996 Wiley-Liss, Inc.