Primary differentiation in the human blastocyst: Comparative molecular portraits of inner cell mass and trophectoderm cells

Whole 8-cell morulae can be aggregated with isolated inner cell masses from blastocysts. On examining semithin light microscope sections of such aggregates we found that cells of the morula changed shape and spread over the surface of the ICM, thus translocating it to the inside of the aggregate. Using single cells from 8-cell embryos in combination with single cells from other stage embryos or isolated ICMs we show that 1/8 blastomeres spread over other cells providing a suitably adhesive surface. The incidence of spreading is high with inner cells from 16-cell embryos (56 %) and 32-cell embryos (62%) and isolated inner cell masses (64%). In contrast, the incidence of spreading of 1/8 blastomeres is low over outer cells from 16-cell embryos (26%) and 32-cell embryos (13%). Blastomeres from 8-cell embryos do not spread over unfertilized 1-cell eggs, 1/2 or 1/4 cells or trophectoderm cells contaminating isolated ICMs. When 1/8 cells are aggregated in pairs they flatten on one another (equal spreading) as occurs at compaction in whole 8-cell embryos. However, if 1/8 is allowed to divide to 2/16 in culture one of the cells engulfs the other (51-62/ pairs). Based on the ideas of Holtfreter (1943) and Steinberg (1964,1978) these results are interpreted to indicate an increase in adhesiveness at the 8-cell stage as well as cytoskeletal mobilization. Following the 8-cell stage there is an increase in adhesiveness of inside cells while the outside cells decrease in adhesiveness. The difference in adhesiveness between inside and outside cells in late morulae is probably central to the divergent differentiation of (inner) ICM and (outer) trophectoderm cell populations.

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Trophectodermal processes regulate the expression of totipotency within the inner cell mass of the mouse expanding blastocyst

Development ◽

10.1242/dev.84.1.63 ◽

1984 ◽

Vol 84 (1) ◽

pp. 63-90

Author(s):

Tom P. Fleming ◽

Paul D. Warren ◽

Julia C. Chisholm ◽

Martin H. Johnson

Keyword(s):

Inner Cell Mass ◽

Cell Mass ◽

Central Area ◽

Ultrastructural Changes ◽

Trophectoderm Cells ◽

Junctional Complexes ◽

Secondary Lysosomes ◽

Cytoplasmic Processes ◽

Cellular Cover

Mouse blastocysts, aged 0, 2, 6 and 12 h from the onset of cavitation, were examined by transmission (TEM) and scanning (SEM) electron microscopy. In TEM sections, trophectoderm cells (TE) differed morphologically from those of the inner cell mass (ICM) by their flattened shape, paler cytosol staining and polarized disposition of both junctional complexes (apicolateral) and intracellular secondary lysosomes (SL; basal). Throughout this period of development, cytoplasmic processes, characterized by abundant SLs, cover approximately 80 % of the juxtacoelic face of the ICM. These processes are shown to be derived from the basal surface of TE cells intermediately placed between polar and mural regions. In SEM preparations of the juxtacoelic ICM surface, revealed by ‘cracking open’ blastocysts, the processes appear as tongue-shaped, centripetally oriented structures which terminate collectively at a central area on the ICM surface. The potential of cultured ICMs to generate TE was demonstrated following their immunosurgical isolation from blastocysts aged up to 12 h post cavitation and by examining the sequence of ultrastructural changes associated with TE generation by ICMs from 2 h blastocysts. In contrast, the juxtacoelic cells of similarly aged ICMs observed in situ in ultrasections of intact embryos showed little or no evidence of totipotency expression as judged by the absence of TE characteristics. Since TE expression within presumptive ICM cells is thought to be generated by an asymmetry of cell contacts (Johnson & Ziomek, 1983), we propose that the juxtacoelic TE processes, by providing a cellular cover to the ICM, function in suppressing the expression in situ of ICM totipotency.

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The human blastocyst: cell number, death and allocation during late preimplantation development in vitro

Development ◽

10.1242/dev.107.3.597 ◽

1989 ◽

Vol 107 (3) ◽

pp. 597-604 ◽

Cited By ~ 13

Author(s):

K. Hardy ◽

A.H. Handyside ◽

R.M. Winston

Keyword(s):

Cell Death ◽

Inner Cell Mass ◽

Cell Mass ◽

Cell Number ◽

Blastocyst Stage ◽

Human Embryos ◽

Cell Numbers ◽

Human Blastocyst ◽

Development In Vitro

The development of 181 surplus human embryos, including both normally and abnormally fertilized, was observed from day 2 to day 5, 6 or 7 in vitro. 63/149 (42%) normally fertilized embryos reached the blastocyst stage on day 5 or 6. Total, trophectoderm (TE) and inner cell mass (ICM) cell numbers were analyzed by differential labelling of the nuclei with polynucleotide-specific fluorochromes. The TE nuclei were labelled with one fluorochrome during immunosurgical lysis, before fixing the embryo and labelling both sets of nuclei with a second fluorochrome (Handyside and Hunter, 1984, 1986). Newly expanded normally fertilized blastocysts on day 5 had a total of 58.3 +/− 8.1 cells, which increased to 84.4 +/− 5.7 and 125.5 +/− 19 on days 6 and 7, respectively. The numbers of TE cells were similar on days 5 and 6 (37.9 +/− 6.0 and 40.3 +/− 5.0, respectively) and then doubled on day 7 (80.6 +/− 15.2). In contrast, ICM cell numbers doubled between days 5 and 6 (20.4 +/− 4.0 and 41.9 +/− 5.0, respectively) and remained virtually unchanged on day 7 (45.6 +/− 10.2). There was widespread cell death in both the TE and ICM as evidenced by fragmenting nuclei, which increased substantially by day 7. These results are compared with the numbers of cells in morphologically abnormal blastocysts and blastocysts derived from abnormally fertilized embryos. The nuclei of arrested embryos were also examined. The number of TE and ICM cells allocated in normally fertilized blastocysts appears to be similar to the numbers allocated in the mouse. Unlike the mouse, however, the proportion of ICM cells remains higher, despite cell death in both lineages.

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Defects in the allocation of cells to the inner cell mass and trophectoderm of parthenogenetic mouse blastocysts

Reproduction Fertility and Development ◽

10.1071/rd9961193 ◽

1996 ◽

Vol 8 (8) ◽

pp. 1193 ◽

Cited By ~ 9

Author(s):

B Mognetti ◽

D Sakkas

Keyword(s):

Inner Cell Mass ◽

Cell Mass ◽

Substantial Reduction ◽

Cell Lineage ◽

Blastocyst Stage ◽

Trophectoderm Cells ◽

Parthenogenetic Development ◽

Preimplantation Stage ◽

Parthenogenetic Mouse Embryos ◽

Parthenogenetic Embryos

Diploid parthenogenetic mouse embryos (which possess two maternally-derived genomes) can develop only as far as the 25-somite stage when transferred in utero and exhibit a substantial reduction in trophoblast tissue. The loss of cultured parthenogenetic embryos during postimplantation indicates that a defect in cell lineage may be evident as early as the blastocyst stage. The possibility that a defect may already be reflected at the preimplantation stage was investigated by examining the allocation of cells to the trophectoderm (trophoblast progenitor cells) and the inner cell mass of haploid and diploid parthenogenetic mouse blastocysts. Utilizing a differential labelling technique for counting cells, diploid parthenogenetic blastocysts were found to have fewer inner cell mass cells and trophectoderm cells than their haploid counterparts and normal blastocysts. In addition, both haploid and diploid parthenogenetic blastocysts had a lower inner cell mass: trophectoderm ratio than normal blastocysts. Thus, the relatively poor development of the trophectoderm lineage at the postimplantation stage is not reflected by a reduction in its allotment of cells at its first appearance. Nevertheless, the findings indicate that parthenogenetic development is already compromised at the blastocyst stage, and provide evidence that the expression of imprinted genes has significance for the development of the embryo at the preimplantation stage.

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