Blastomeres arising from the first cleavage division have distinguishable fates in normal mouse development
Karolina Piotrowska*,
Florence Wianny,
Roger A. Pedersen
and
Magdalena Zernicka-Goetz
Wellcome/CRC Institute and Department of Genetics, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK
* Permanent address: Department of Experimental Embryology, Polish Academy of Sciences, Jastrzebiec, Poland
Permanent address: Department of Obstetrics, Gynecology and Reproductive Sciences, University of California, San Francisco, CA, USA
View larger version (43K):
[in a new window]
|
Fig. 1. Clones derived from the two-cell stage tend to occupy either embryonic or abembryonic parts of the blastocyst. Blastomeres of two-cell embryos were labelled with DiI (red) or DiD (blue) and the distribution of the progeny of labelled cells were analysed at the blastocyst stage. The frequencies of the four categories of blastocyst scored are indicated below. A total of 69 blastocysts were classified according to the extent to which cells derived from one blastomere comprising mainly the embryonic part (A) crossed the embryonic-abembryonic boundary zone (consisting of a region one cell deep and parallel to the roof of the blastocyst cavity, i.e. between broken white lines) into the abembryonic part and (B) vice versa. Comparison of the data in the row A and row B indicated that cells from the abembryonic part showed a greater tendency ( 2 P<0.02, 3 d.f.) to contribute to the embryonic part than their embryonic cousins to contribute to abembryonic part. Blastocysts were scored ++ if up to two cells crossed the boundary zone (arrow). In cases where three cells crossed this boundary, blastocysts were scored +. When four cells, or five or more cells crossed the boundary the blastocyst were scored – and ––, respectively. The micrographs represent individual optical sections mid-way through the embryo to show the cavity, which occupies the lower half of each blastocyst (see Materials and Methods). The broken yellow lines show the border between clones derived from each blastomere. Scale bar: 25 µm.
|
|
View larger version (82K):
[in a new window]
|
Fig. 2. The early dividing two-cell stage blastomere contributes to the embryonic part of the blastocyst. Blastomeres of two-cell embryos were labelled with DiI or DiD (left hand micrographs; A,D). At the three-cell stage it was recorded whether the earlier dividing cell had been marked with the former (red, upper panel, B) or latter (blue, lower panel, E) dye. Embryos were then allowed to develop to the blastocyst stage (right hand micrograph; C,F). The blastomere that divided first to the four-cell stage gave rise to cells in the embryonic part in 49/56 (88%) embryos in this series of experiments. Scale bar: 25 µm.
|
|
View larger version (88K):
[in a new window]
|
Fig. 3. Confocal sections of a blastocyst that shows correspondence of the clonal border of the two-cell stage progeny with a plane separating the embryonic and abembryonic parts. Blastomeres were labelled at the two-cell stage with dyes of different colours. The boundary zone is marked with broken red lines and the border of the blastocoel was traced on a central section and is shown projected onto each of the other sections as a broken white line. (A-I) Individual optical sections at 7.5 µm intervals in the ‘z-dimension’. (J,K) The dissociated cells of this blastocyst observed by fluorescence or DIC optics, respectively.
|
|
View larger version (81K):
[in a new window]
|
Fig. 4. Confocal sections of a blastocyst in which the clonal border of the two-cell stage progeny (broken yellow line) is tilted with respect to the boundary zone between the embryonic and abembryonic parts (broken red lines). Blastomeres were labelled with dyes of two different colours at the two-cell stage. The border of the blastocoel was traced on a central section and is shown projected onto each of the other sections as a broken white line. (A-H) Individual optical sections at 7.5 µm intervals in the z-dimension. (I,J) The dissociated cells of this blastocyst observed by DIC optics or fluorescence, respectively. Note that all cells are labelled but not uniformly throughout.
|
|
View larger version (41K):
[in a new window]
|
Fig. 5. Several possible models depicting the relationship between the first cleavage plane and the boundary between embryonic and abembryonic parts of the blastocyst. (A) Model by Piotrowska and Zernicka-Goetz (Piotrowska and Zernicka-Goetz, 2001). The left-hand diagram indicates the position of the first cleavage that divides the zygote into two (orange and blue) cells. The first cleavage plane (outlined in black) is set in relation to the animal pole (marked by the polar body shown in green) and the SEP (the sperm head is shown in yellow). Depending on whether cleavage passes on one or other side of these structures, its actual position with respect to the animal pole might vary by several degrees (arrow, plane outlined in green). The two-cell stage blastomere that inherits the SEP tends to divide first to produce cells that tend to populate the embryonic part of the blastocyst (right-hand diagram). The first cleavage plane is reflected in the blastocyst as the border (black disc) between the embryonic region (Em (blue); polar trophectoderm and ICM destined to become epiblast) and the abembryonic region (Ab (orange); mural trophectoderm and ICM that will contribute cells to primitive endoderm). Blastocoel in grey. The precise position of this border may reflect variations in the first cleavage plane (arrow leading to green disc). (B) Variation on the model in A in which factors influencing both the embryonic-abembryonic axis and the first cleavage plane are inter-dependent. In this model, the first cleavage plane of the zygote, and hence the clonal border of the blastocyst stage (outlined in green), is set by the relative positions of the polar body and SEP. The morphology of the blastocyst defined by a plane separating the embryonic and abembryonic parts (outlined in black) is depicted as being independently set by factor(s) (white arc) present in the animal pole cytoplasm. (C) Variation on the model in A in which the animal pole marked by the polar body is the primary determinant of the cleavage plane of the zygote or clonal border of the blastocyst (outlined in green). The displacement of the plane separating embryonic and abembryonic parts (outlined in black) is depicted as being influenced by the variation in the position of sperm entry (arrows).
|
|
© The Company of Biologists Ltd 2001
