Trophectoderm and embryoblast proliferate at slow pace in the course of embryonic diapause in the roe deer (Capreolus capreolus)

Abstract

The European roe deer (Capreolus capreolus) was the first mammal in which embryonic diapause has been described. While diapause is characterized by a complete developmental arrest in some species, roe deer blastocysts show a very slow, yet continuous growth. To date, it is neither known whether this growth is accompanied by developmental progression nor whether it is uniform in both, the trophectoderm (TE) and the inner cell mass (ICM). We collected roe deer blastocysts during the regular hunting season from September 2018 to January 2019, and quantified the fraction of cells expressing the proliferation marker Ki67 by immunofluorescence and light-sheet microscopy. We found that the cell number increased from around 300 cells in September to over 20'000 cells per blastocyst in December before elongation occured. Concurrently, we observed considerable morphological changes, i.e. cavity formation and transition to a disk-like shape of the inner cell mass. During diapause, less than 10% of all cells displayed positive Ki67 staining. Strikingly, the relative increase in cell number was lower in the ICM compared to the TE, whereas the fraction of Ki67 positive cells appeared to be lower in the TE than in the ICM. Our findings thus confirm that roe deer blastocysts display developmental progression in the course of diapause. We hypothesize that while the overall duration of the cell cycle is longer in the ICM than in the TE, the fractional distribution of cell cycle phases differs, with TE cells having a longer G1 phase than cells of the ICM.

Amino acids activate mTORC1 to release roe deer embryos from decelerated proliferation during diapause

“…Different from other known species exhibiting diapause, roe deer and nine-banded armadillo embryos continuously increase in size and cell number over the course of diapause ( 4 – 6 ). In the diapausing roe deer embryo, less than 10% of cells stain positive for the proliferation marker Ki67, and the fraction of Ki67-positive cells is slightly lower in the trophectoderm (TE) than in the inner cell mass (ICM) ( 7 ). The doubling time of the number of cells is estimated to be 18 to 21 d for TE cells and 21 to 26 d for the ICM cells ( 7 ).…”

Molecular Regulation of Paused Pluripotency in Early Mammalian Embryos and Stem Cells

“…But not all diapause embryos are truly dormant. About 10% of cells in the roe deer blastocyst proliferate throughout diapause, with the embryo expanding from 300 to 20,000 cells within 4 months (Rüegg et al, 2020). The inner cell mass proliferates less than the TE.…”

“…The inner cell mass proliferates less than the TE. The inner cell mass also undergoes morphological changes, from round to flattened to cyst to disc, indicating active restructuring and communication between the cells, even at the low rate of proliferation (Rüegg et al, 2020).…”

Embryonic diapause in mammals and dormancy in embryonic stem cells with the European roe deer as experimental model

“…Strikingly, development is not completely halted, as shown in many other species. Rather, it features gradual proliferation at a considerably slower pace (Rüegg et al 2020). The diameter of the blastocyst increases from 0.5 mm in September to approximately 4.0 mm in December ( Fig.…”

“…The diameter of the blastocyst increases from 0.5 mm in September to approximately 4.0 mm in December ( Fig. 1; Drews et al 2019;Rüegg et al 2020;Ulbrich 2020a, 2020b). The end of diapause in the roe deer is marked by a rapid resumption of developmental pace, which morphologically coincides with embryo elongation, a feature shared across ungulates.…”