Cosegregation of monoclonal antibody reactivity and cell behaviour in the mouse preimplantation embryo

Development ◽  
1982 ◽  
Vol 70 (1) ◽  
pp. 261-278
Author(s):  
Beverley J. Randle
Keyword(s):  
Monoclonal Antibody ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Antigen Expression ◽  
Cell Stage ◽  
Cell Behaviour ◽  
Minor Population ◽  
Cell Embryo ◽  
Surface Labelling

Expression of an antigen, recognized by a monoclonal antibody raised against PCI 3 embryonal carcinoma, is described in mouse preimplantation embryogenesis. The antigen is found in the cytoplasm of ovulated ova and is first noted on the cell surface of the 1-cell embryo 20 h post-ovulation. Surface labelling of blastomeres is uniform until the 8-cell stage when antigen expression becomes polarized along the radial axis of the embryo. Two major populations of blastomeres are distinguishable on division to the 16-cell morula. Dissociation of morulae in calcium-free medium yields large, polar, antigen-positive cells and small apolar cells with reduced levels of detectable antigen. A third, minor population of small, antigen-negative cells is also found in vivo. Large and small blastomeres differ in their ability to relocate within the embryo when aggregated with intact 16-cell-stage embryos. The small blastomeres of the 16-cell morula contribute significantly to the inner cell mass while the large antigen-positive cells are found only in the trophectoderm.

When and how does cell division order influence cell allocation to the inner cell mass of the mouse blastocyst?

Development ◽  
1987 ◽  
Vol 100 (2) ◽  
pp. 325-332
Author(s):  
C.L. Garbutt ◽  
M.H. Johnson ◽  
M.A. George
Keyword(s):  
Cell Division ◽  
Cell Population ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Lineage ◽  
The Other ◽  
Mouse Blastocyst ◽  
Short Term ◽  
Cell Stage ◽  
Cell Embryo

Aggregate 8-cell embryos were constructed from four 2/8 pairs of blastomeres, one of which was marked with a short-term cell lineage marker and was also either 4 h older (derived from an early-dividing 4-cell) or 4 h younger (derived from a late-dividing 4-cell) than the other three pairs. The aggregate embryos were cultured to the 16-cell stage, at which time a second marker was used to label the outside cell population. The embryos were then disaggregated and each cell was examined to determine its labelling pattern. From this analysis, we calculated the relative contributions to the inside cell population of the 16-cell embryo of older and younger cells. Older cells were found to contribute preferentially. However, if the construction of the aggregate 8-cell embryo was delayed until each of the contributing 2/8 cell pairs had undergone intercellular flattening and then had been exposed to medium low in calcium to reverse this flattening immediately prior to aggregation, the advantage possessed by the older cells was lost. These results support the suggestion that older cells derived from early-dividing 4-cell blastomeres contribute preferentially to the inner cell mass as a result of being early-flattening cells.

scholarly journals Embryo cell allocation patterns are not altered by biopsy but can be linked with further development

Reproduction ◽  
2017 ◽  
Vol 154 (6) ◽  
pp. 807-814
Author(s):  
L P Sepulveda-Rincon ◽  
N Islam ◽  
P Marsters ◽  
B K Campbell ◽  
N Beaujean ◽  
...  
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Blastocyst Stage ◽  
Mouse Strains ◽  
Random Pattern ◽  
Cell Stage ◽  
Cell Counts ◽  
Cell Embryo ◽  
Allocation Patterns ◽  
Further Development

It has been suggested that first embryo cleavage can be related with the embryonic–abembryonic axis at blastocyst stage in mice. Thus, cells of the 2-cell embryo might be already biased to form the inner cell mass or trophectoderm. This study was conducted to observe the possible effects of embryo biopsy on cell allocation patterns during embryo preimplantation in two different mouse strains and the effects of these patterns on further development. First, one blastomere of the 2-cell embryo was injected with a lipophilic tracer and cell allocation patterns were observed at blastocyst stage. Blastocysts were classified into orthogonal, deviant or random pattern. For the first experiment, embryos were biopsied at 8-cell stage and total cell counts (TCC) were annotated. Furthermore, non-biopsied blastocysts were transferred into foster mothers. Then, pups and their organs were weighed two weeks after birth. Random pattern was significantly recurrent (≈60%), against orthogonal (<22%) and deviant (<22%) patterns among groups. These patterns were not affected by biopsy procedure. However, TCC on deviant embryos were reduced after biopsy. Moreover, no differences were found between patterns for implantation rates, litter size, live offspring and organ weights (lungs, liver, pancreas and spleen). However, deviant pups presented heavier hearts and orthogonal pups presented lighter kidneys among the group. In conclusion, these results suggest that single blastomere removal does not disturb cell allocation patterns during pre-implantation. Nonetheless, the results suggest that embryos following different cell allocation patterns present different coping mechanisms against in vitro manipulations and further development might be altered.

The effect of prolonged decompaction on the development of the preimplantation mouse embryo

Development ◽  
1979 ◽  
Vol 54 (1) ◽  
pp. 241-261
Author(s):  
M. H. Johnson ◽  
J. Chakraborty ◽  
A. H. Handyside ◽  
K. Willison ◽  
P. Stern
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Rabbit Antiserum ◽  
Embryonal Carcinoma Cell ◽  
Cell Stage ◽  
Embryonal Carcinoma Cell Line ◽  
Preimplantation Mouse Embryo ◽  
Preimplantation Mouse ◽  
Cell Embryo ◽  
Relationship Of

A rabbit antiserum to a mouse embryonal carcinoma cell line blocks compaction of cleaving mouseembryos. Cell division is not affected up to the 32-cell stage but intracellular junctions fail to develop. Removal of the antibody at this stage permits compaction to occur and a normal blastocyst develops. Prolonged decompaction beyond the 32-cell embryo results in an increasing proportion of malformed blastocysts in which trophectodermal cells predominate and functional inner cell mass (ICM) cells are reduced or absent. The relationship of compaction to the generation of ICM and trophectoderm lineages in the intact embryo is discussed.

The effects of embryo stage and cell number on the composition of mouse aggregation chimaeras

Zygote ◽  
2000 ◽  
Vol 8 (3) ◽  
pp. 235-243 ◽  
Author(s):  
Pin-chi Tang ◽  
John D. West
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Number ◽  
Mouse Embryos ◽  
Cell Stage ◽  
Advanced Embryo ◽  
Preimplantation Mouse Embryos ◽  
Preimplantation Mouse ◽  
Cell Embryo ◽  
Embryo Stage

Studies with intact preimplantation mouse embryos and some types of chimaeric aggregates have shown that the most advanced cells are preferentially allocated to the inner cell mass (ICM) rather than the trophectoderm. Thus, differences between 4-cell and 8-cell stage embryos could contribute to the tendency for tetraploid cells to colonise the trophectoderm more readily than the ICM in 4-cell tetraploid[harr ]8 cell diploid chimaeras. The aim of the present study was to test whether 4-cell stage embryos in 4-cell diploid[harr ]8-cell diploid aggregates contributed equally to all lineages present in the E12.5 conceptus. These chimaeras were compared with those produced from standard aggregates of two whole 8-cell embryos and aggregates of half an 8-cell embryo with a whole 8-cell embryo. As expected, the overall contribution of 4-cell embryos was lower than that of 8-cell embryos and similar to that of half 8-cell stage embryos. In the 4-cell[harr ]8-cell chimaeras the 4-cell stage embryos did not contribute more to the trophectoderm than the ICM derivatives. Thus, differences between 4-cell and 8-cell embryos cannot explain the restricted tissue distribution of tetraploid cells previously reported for 4-cell tetraploid[harr ]8-cell diploid chimaeras. It is suggested that cells from the more advanced embryo are more likely to contribute to the ICM but, for technical reasons, are prevented from doing so in simple aggregates of equal numbers of whole 4-cell and whole 8-cell stage embryos.

scholarly journals Development of Preimplantation Mouse Embryos in vivo and in vitro

1982 ◽  
Vol 35 (2) ◽  
pp. 187 ◽  
Author(s):  
GM Harlow ◽  
P Quinn
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Number ◽  
Blastocyst Stage ◽  
Culture Conditions ◽  
Cell Stage ◽  
Preimplantation Mouse ◽  
Development In Vitro

The culture conditions for the development in vitro of (C57BL/6 X CBA) F2 hybrid two-cell embryos to the blastocyst stage have been optimized. Commercially available pre-sterile disposable plastic culture dishes supported more reliable development than re-usable washed glass tubes. The presence of an oil layer reduced the variability in development. An average of 85 % of blastocysts developed from hybrid two-cell embryos cultured in drops of Whitten's medium under oil in plastic culture dishes in an atmosphere of 5% O2 : 5% CO2 : 90% N2 ? The time taken for the total cell number to double in embryos developing in vivo was 10 h, and in cultured embryos 17 h. Embryos cultured in vitro from the two-cell stage to blastocyst stage were retarded by 18-24 h in comparison with those remaining in vivo. Day-4 blastocysts in vivo contained 25-70 cells (mean 50) with 7-28 (mean 16) of these in the inner cell mass. Cultured blastocysts contained 19-73 cells (mean 44) with 8-34 (mean 19) of these in the inner cell mass. In the uterine environment, inner-cell-mass blastomeres divided at a faster rate than trophectoderm blastomeres and it is suggested that a long cell cycle is associated with terminal differentiation. Although cultured blastocysts and inner cell masses contained the same number of cells as blastocysts and inner cell masses in vivo, the rate of cell division in cultured inner cell masses was markedly reduced.

Muscle cell differentiation in ascidian embryos analysed with a tissue-specific monoclonal antibody

Development ◽  
1987 ◽  
Vol 99 (2) ◽  
pp. 163-171
Author(s):  
T. Nishikata ◽  
I. Mita-Miyazawa ◽  
T. Deno ◽  
N. Satoh
Keyword(s):  
Monoclonal Antibody ◽  
Cell Differentiation ◽  
Muscle Cell ◽  
Antigen Expression ◽  
Relative Molecular Mass ◽  
Specific Monoclonal Antibody ◽  
Cell Stage ◽  
Muscle Cell Differentiation ◽  
Cell Embryo ◽  
Ascidian Embryos

Utilizing a muscle-specific monoclonal antibody (Mu-2) as a probe, we analysed developmental mechanisms involved in muscle cell differentiation in ascidian embryos. The antigen recognized by Mu-2 was a single polypeptide with a relative molecular mass of about 220 X 10(3). It first appeared at the early tailbud stage and continued to be expressed until the swimming larva stage. There were distinct and separate puromycin and actinomycin D sensitivity periods during the occurrence of the antigen, suggesting the new synthesis of the polypeptide by developing muscle cells. Embryos that had been permanently arrested with aphidicolin in the early cleavage stages up to the 32-cell stage did not express the antigen. DNA replications may be required for the antigen expression. Embryos that had been arrested with cytochalasin B in the 8-cell and later stages developed the antigen, and the number and position of the arrested blastomeres exhibiting the differentiation marker almost corresponded to those of the B4.1-line muscle lineage. Furthermore, in quarter embryos developed from each blastomere pair isolated from the 8-cell embryo, all the B4.1 as well as a part of b4.2 partial embryos expressed the antigen, while the a4.2 and A4.1 partial embryos did not show the antigen expression. These results may provide further support for the existence of cytoplasmic determinants for muscle cell differentiation in this mosaic egg.

Distribution of antibody- and lectin-binding sites on dissociated blastomeres from mouse morulae: evidence for polarization at compaction

Development ◽  
1980 ◽  
Vol 60 (1) ◽  
pp. 99-116
Author(s):  
Alan H. Handyside
Keyword(s):  
Binding Sites ◽  
Inner Cell Mass ◽  
Lectin Binding ◽  
Cell Mass ◽  
Spatial Differentiation ◽  
Antibody Fragments ◽  
Cell Stage ◽  
Surface Labelling ◽  
Increase Surface Area ◽  
Lectin Binding Sites

The distribution of binding sites for rabbit anti-species antiserum, Concanavalin A (Con A) and peanut agglutinin (PNA) on dissociated blastomeres from 2- to 16-cell mouse embryos has been investigated using direct and indirect fluorescence techniques. With each ligand, paraformaldehyde-fixed blastomeres from 2- to 8-cell precompact embryos were uniformly surface labelled; the majority (77%) of late compact 8-cell blastomeres showed quantitative polarization of surface labelling; and 16-cell blastomeres were either polarized (53·3%) or uniformly surface labelled. Binding of fluorescein-conjugated PNA increased at the 16-cell stage. Labelling patterns on unfixed blastomeres were similar to those on fixed blastomeres except that surface label was patched and became internalized, most rapidly from the less heavily labelled areas of 8- and 16-cell blastomeres. Quantitative polarization of binding sites at postcompaction stages was detected after (i) fixation, (ii) pretreatment and labelling in the presence of azide, cytochalasin D and/or colcemid, or (iii) labelling with monovalent Fab1 antibody fragments. It is probably due, therefore, to the presence of microvilli at the heavily labelled pole, which increase surface area and are known to become localized to the outer surface of the compact morula (Ducibella, Ukena, Karnovsky & Anderson, 1977). The possibility that the cleavage of polarized blastomeres into dissimilar daughter blastomeres could provide a mechanism for the spatial differentiation of the inner cell mass and trophectoderm of the blastocyst is briefly discussed.

scholarly journals The role of Trp53 in the mouse embryonic response to DNA damage

2019 ◽  
Vol 25 (7) ◽  
pp. 397-407
Author(s):  
Yvonne Wilson ◽  
Ian D Morris ◽  
Susan J Kimber ◽  
Daniel R Brison
Keyword(s):  
Dna Damage ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Blastocyst Stage ◽  
Human Embryos ◽  
Cell Stage ◽  
X Irradiation

Abstract Apoptosis occurs primarily in the blastocyst inner cell mass, cells of which go on to form the foetus. Apoptosis is likely to play a role in ensuring the genetic integrity of the foetus, yet little is known about its regulation. In this study, the role of the mouse gene, transformation-related protein 53 (Trp53) in the response of embryos to in vitro culture and environmentally induced DNA damage was investigated using embryos from a Trp53 knockout mouse model. In vivo-derived blastocysts were compared to control embryos X-irradiated at the two-cell stage and cultured to Day 5. An analysis of DNA by comet assay demonstrated that 1.5 Gy X-irradiation directly induced damage in cultured two-cell mouse embryos; this was correlated with retarded development to blastocyst stage and increased apoptosis at the blastocyst stage but not prior to this. Trp53 null embryos developed to blastocysts at a higher frequency and with higher cell numbers than wild-type embryos. Trp53 also mediates apoptosis in conditions of low levels of DNA damage, in vivo or in vitro in the absence of irradiation. However, following DNA damage induced by X-irradiation, apoptosis is induced by Trp53 independent as well as dependent mechanisms. These data suggest that Trp53 and apoptosis play important roles in normal mouse embryonic development both in vitro and in vivo and in response to DNA damage. Therefore, clinical ART practices that alter apoptosis in human embryos and/or select embryos for transfer, which potentially lack a functional Trp53 gene, need to be carefully considered.

Development of single mouse blastomeres enlarged to zygote size in conditions of nucleo-cytoplasmic synchrony

Zygote ◽  
2002 ◽  
Vol 10 (4) ◽  
pp. 283-290 ◽  
Author(s):  
Jacek A. Modliński ◽  
Jean-Pierre Ozil ◽  
Marta K. Modliński ◽  
Alina Szarska ◽  
Michael A. Reed ◽  
...  
Keyword(s):  
Mouse Strain ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Blastocyst Formation ◽  
Cell Stage ◽  
Cell Numbers ◽  
Insufficient Number

The following blastomeres were enlarged to the size of the zygote by one, two or three rounds of blastomere enucleation and electrofusion: (1) from the 2-cell stage (referred to as 2/1 embryos), (2) from the 4-cell stage (referred to as 4/1 embryos), (3) from the 8-cell stage (referred to as 8/1 embryos). Such single enlarged blastomeres developed into blastocysts in vivo in 55.5% (2/1), 28% (4/1) and 6.6% (8/1) of cases. Their mean cell numbers were 45.3, 24.5 and 13.0 in 2/1, 4/1 and 8/1 embryos, respectively. When a blastomere nucleus from another mouse strain (heterologous nucleus) was substituted for a blastomere's own (homologous) one, then fewer blastocysts were formed from 2/1 embryos (34.6%), but not from 4/1 and 8/1 embryos. Five young (10.4%) were born from 2/1 embryos with a homologous nucleus, and nine (8.3%) from 2/1 embryos with heterologous nuclei. Four young (7.1%) were born from 4/1 embryos with heterologous nuclei. No young were obtained from 8/1 embryos. Incorrect cavitation resulting in trophoblastic vesicles and false blastocyst formation was common in 4/1 embryos (18.7% of those with homologous nuclei and 41.3% with heterologous nuclei) and in 8/1 embryos (53.3% and 43.7%, respectively). The results show that neither enlargement to zygote size nor nucleo-cytoplasmic synchrony improve postimplantation development of 4- and 8-cell stage blastomeres when compared with less enlarged non-synchronous ones; therefore, it appears that an insufficient number of inner cell mass cells in blastocysts and not too small a size of isolated blastomeres precludes their postimplantation development.

scholarly journals Totipotency of mouse zygotes extends to single blastomeres of embryos at the four-cell stage

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Marino Maemura ◽  
Hiroaki Taketsuru ◽  
Yuki Nakajima ◽  
Ruiqi Shao ◽  
Ayaka Kakihara ◽  
...  
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Mouse Embryos ◽  
Primitive Endoderm ◽  
Cell Stage ◽  
Supportive Environment ◽  
Mouse Zygotes ◽  
Single Blastomeres ◽  
Multicellular Organisms

AbstractIn multicellular organisms, oocytes and sperm undergo fusion during fertilization and the resulting zygote gives rise to a new individual. The ability of zygotes to produce a fully formed individual from a single cell when placed in a supportive environment is known as totipotency. Given that totipotent cells are the source of all multicellular organisms, a better understanding of totipotency may have a wide-ranging impact on biology. The precise delineation of totipotent cells in mammals has remained elusive, however, although zygotes and single blastomeres of embryos at the two-cell stage have been thought to be the only totipotent cells in mice. We now show that a single blastomere of two- or four-cell mouse embryos can give rise to a fertile adult when placed in a uterus, even though blastomere isolation disturbs the transcriptome of derived embryos. Single blastomeres isolated from embryos at the eight-cell or morula stages and cultured in vitro manifested pronounced defects in the formation of epiblast and primitive endoderm by the inner cell mass and in the development of blastocysts, respectively. Our results thus indicate that totipotency of mouse zygotes extends to single blastomeres of embryos at the four-cell stage.

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