Trophoblast differentiation, invasion and hormone secretion in a three-dimensional in vitro implantation model with rhesus monkey embryos

T Arthur Chang,Maureen Durning,Jessica G Drenzek,Behzad Gerami-Naini,Thaddeus G Golos,Gennadiy I Bondarenko,Jenna Kropp Schmidt,Mark A Garthwaite

doi:10.1186/s12958-018-0340-3

Abstract

BackgroundThe initiation of primate embryo invasion into the endometrium and the formation of the placenta from trophoblasts, fetal mesenchyme, and vascular components are essential for the establishment of a successful pregnancy. The mechanisms which direct morphogenesis of the chorionic villi, and the interactions between trophectoderm-derived trophoblasts and the fetal mesenchyme to direct these processes during placentation are not well understood due to a dearth of systems to examine and manipulate real-time primate implantation. Here we describe an in vitro three-dimensional (3-D) model to study implantation which utilized IVF-generated rhesus monkey embryos cultured in a Matrigel explant system.MethodsBlastocyst stage embryos were embedded in a 3-D microenvironment of a Matrigel carrier and co-cultured with a feeder layer of cells generating conditioned medium. Throughout the course of embryo co-culture embryo growth and secretions were monitored. Embedded embryos were then sectioned and stained for markers of trophoblast function and differentiation.ResultsSigns of implantation were observed including enlargement of the embryo mass, and invasion and proliferation of trophoblast outgrowths. Expression of chorionic gonadotropin defined by immunohistochemical staining, and secretion of chorionic gonadotropin and progesterone coincident with the appearance of trophoblast outgrowths, supported the conclusion that a trophoblast cell lineage formed from implanted embryos. Positive staining for selected markers including Ki67, MHC class I, NeuN, CD31, vonWillebrand Factor and Vimentin, suggest growth and differentiation of the embryo following embedding.ConclusionsThis 3-D in vitro system will facilitate further study of primate embryo biology, with potential to provide a platform for study of genes related to implantation defects and trophoblast differentiation.

Highlights

The initiation of primate embryo invasion into the endometrium and the formation of the placenta from trophoblasts, fetal mesenchyme, and vascular components are essential for the establishment of a successful pregnancy
Fertilized embryos were cultured to the 8-cell stage at 37 °C in 5% CO2, 5% O2, and 90% N2 in chemically defined, protein-free hamster embryo culture medium-9 (HECM-9) media [38, 41, 42] supplemented with amino acids
Overall, the results reported here establish a platform for assessing primate implantation in vitro

Summary

Introduction

The initiation of primate embryo invasion into the endometrium and the formation of the placenta from trophoblasts, fetal mesenchyme, and vascular components are essential for the establishment of a successful pregnancy. Embryo surrogates, such as trophoblast spheroids, derived from immortal cell lines have been utilized to evaluate attachment, invasion and migration [6, 7, 11, 12] These models are limited to studying a specific event during implantation and lack a complete representation of all cell types present at the maternal-fetal interface, including immune cells, further development of in vitro implantation models that better simulate in vivo implantation are needed

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