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, 12 (3), e0175032
eCollection

Mouse Blastomeres Acquire Ability to Divide Asymmetrically Before Compaction

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Mouse Blastomeres Acquire Ability to Divide Asymmetrically Before Compaction

Monika Humięcka et al. PLoS One.

Abstract

The mouse preimplantation embryo generates the precursors of trophectoderm (TE) and inner cell mass (ICM) during the 8- to 16-cell stage transition, when the apico-basal polarized blastomeres undergo divisions that give rise to cells with different fate. Asymmetric segregation of polar domain at 8-16 cell division generate two cell types, polar cells which adopt an outer position and develop in TE and apolar cells which are allocated to inner position as the precursors of ICM. It is still not know when the blastomeres of 8-cell stage start to be determined to undergo asymmetric division. Here, we analyze the frequency of symmetric and asymmetric divisions of blastomeres isolated from 8-cell stage embryo before and after compaction. Using p-Ezrin as the polarity marker we found that size of blastomeres in 2/16 pairs cannot be used as a criterion for distinguishing symmetric and asymmetric divisions. Our results showed that at early 8-cell stage, before any visible signs of cortical polarity, a subset of blastomeres had been already predestined to divide asymmetrically. We also showed that almost all of 8-cell stage blastomeres isolated from compacted embryo divide asymmetrically, whereas in intact embryos, the frequency of asymmetric divisions is significantly lower. Therefore we conclude that in intact embryo the frequency of symmetric and asymmetric division is regulated by cell-cell interactions.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Fig 1
Fig 1. Pattern of divisions of blastomeres isolated from uncompacted and compacted embryos and intact embryos.
(A) p-Ezrin uniformly distributed in their cortex of cells of uncompacted eight-cell embryo and restricted to the apical domain of cells in compacted embryo. (B) Symmetric and asymmetric inheritance of p-Ezrin during divisions of 1/8 blastomeres isolated from uncompacted and compacted embryos. (C) symmetric and asymmetric divisions in group of equally sized blastomeres in 2/16 pair. a) p-Ezrin uniformly distributed in cortex of blastomeres before compaction. b) p-Ezrin restricted to the apical domain of blastomeres after compaction. (D) symmetric and asymmetric divisions in group of unequally sized blastomeres in 2/16 pair. a) p-Ezrin uniformly distributed in cortex of blastomeres before compaction; b) p-Ezrin restricted to the apical domain of blastomeres after compaction. Scale bar 10μm. (E,F,G,H) Analysis of symmetric/asymmetric and equal/unequal divisions of 1/8 blastomeres isolated from uncompacted (n = 80), compacted 8-cell stage embryos (n = 78) and blastomeres of intact embryos (n = 90). The distribution of p-Ezrin was estimated by immunofluorescence after fixation of 2/16 pairs of blastomeres.
Fig 2
Fig 2. Expression of CDX2 in embryo fragments after equal and unequal divisions of blastomeres isolated from uncompacted and compacted embryos.
(A) Equal division of 1/8 blastomeres—1) 2/16 pairs of blastomeres before compaction. 2,3) 2/16 pairs of blastomeres after compaction. The borders of cells are visualized by actin staining with phalloidin (white). 4,5,6) Embryo fragments composed from 4 cells generated after the next round of division with different number of CDX2-positive cells—4/4, 3/4, 2/4 (number of CDX2-positive cells/number of cells). All images are merged single optical images of chromatin (red) and CDX2 (yellow) staining. Scale bar 10μm. (B) Unequal division of 1/8 blastomeres—1) 2/16 pairs of blastomeres before compaction, 2,3) 2/16 pairs of blastomeres after compaction. The borders of cells are visualized by actin staining with phalloidin (white). 4,5,6) Embryo fragments composed from 4 cells generated after the next round of division with different number of CDX2-positive cells (4/4, 3/4, 2/4). All images are merged single optical images of chromatin (red) and CDX2 (yellow) staining. Scale bar 10μm. (C) Frequency of embryo fragments with different number of CDX2-positive cells (4/4, 3/4, 2/4) developed from 1/8 blastomeres of uncompacted and uncompacted embryos. (D) Frequency of embryo fragments with different number of CDX2-positive cells developed from 1/8 blastomeres of uncompacted and uncompacted embryos after equal and unequal divisions.
Fig 3
Fig 3. The model of cell division of 2/16 pair of blastomeres developed in symmetric/asymmetric and equal/unequal divisions of 1/8 blastomeres.
Fig 4
Fig 4. Cell-cell interactions in the 2/16 pair of blastomeres.
(A) 1. Symmetric equal division–p-Ezrin in both blastomeres. 2. Asymmetric equal division–p-Ezrin in one of the blastomere. 3. Asymmetric unequal division–p-Ezrin in the bigger blastomere. (B) Asymmetric unequal division. 1,2,3 Sequence of enveloping the small apolar blastomere by the polar bigger one. Scale bar 10μm.

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Grant support

The author(s) received no specific funding for this work.
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