Trophectoderm regeneration to support full-term development in the inner cell mass isolated from bovine blastocyst

Nanami Kohri,Hiroki Akizawa,Sakie Iisaka,Hanako Bai,Yojiro Yanagawa,Masashi Takahashi,Masaya Komatsu,Masahito Kawai,Masashi Nagano,Manabu Kawahara

doi:10.1074/jbc.ra119.010746

Abstract

Which comes first: tissue structure or cell differentiation? Although different cell types establish distinct structures delineating the inside and outside of an embryo, they progressively become specified by the blastocyst stage, when two types of cell lineages are formed: the inner cell mass (ICM) and the trophectoderm (TE). This inside-outside aspect can be experimentally converted by the isolation of the ICM from a blastocyst, leading to a posteriori externalization of the blastomeres composing the outermost layer of the ICM. Here, we investigated the totipotency of isolated mouse and bovine ICMs to determine whether they are competent for TE regeneration. Surprisingly, a calf was generated from the bovine isolated ICM with re-formed blastocoel (re-iICM), but no mouse re-iICMs developed to term. To further explore the cause of difference in developmental competency between the mouse and bovine re-iICMs, we investigated the SOX17 protein expression that is a representative molecular marker of primitive endoderm. The localization pattern of SOX17 was totally different between mouse and bovine embryos. Particularly, the ectopic SOX17 localization in the TE might be associated with lethality of mouse re-iICMs. Meanwhile, transcriptome sequencing revealed that some of the bovine re-iICMs showed transcriptional patterns of TE-specific genes similar to those of whole blastocysts. Our findings suggest that TE regeneration competency is maintained longer in bovine ICMs than in mouse ICMs and provide evidence that the ICM/TE cell fate decision is influenced by structural determinants, including positional information of each blastomere in mammalian embryos.

Highlights

Which comes first: tissue structure or cell differentiation? different cell types establish distinct structures delineating the inside and outside of an embryo, they progressively become specified by the blastocyst stage, when two types of cell lineages are formed: the inner cell mass (ICM) and the trophectoderm (TE)
The purity of the isolated ICM (iICM) was confirmed on the basis of CDX2 fluorescent signals that characterize TE, and no TE carryover was noted in both mouse and bovine iICMs (Fig. 1, B and C)
When the iICMs were cultured in the embryo-cultivation medium for 24 h, the formation of morphologically clear blastocoels was induced in both mouse and bovine iICMs (Fig. 1A)

Summary

Introduction

Which comes first: tissue structure or cell differentiation? different cell types establish distinct structures delineating the inside and outside of an embryo, they progressively become specified by the blastocyst stage, when two types of cell lineages are formed: the inner cell mass (ICM) and the trophectoderm (TE). To compare the localization patterns of YAP1 and CDX2 between bovine and mouse re-iICMs, we performed dual immunostaining (Fig. 2, A and B).

Results

Conclusion