Early stages of implantation as revealed by an in vitro model

Abstract

Our limited understanding of the processes underlying steroid hormonal control of human endometrial receptivity is largely due to the lack of a relevant model system. To overcome scarcity of material, we have developed a model in which mouse embryos attach to human Ishikawa cells, which express functional steroid hormone receptors. Blastocysts flushed from day 4 pregnant superovulated mice were transferred to confluent Ishikawa cell monolayers. After 48 h of co-culture, 85% of the blastocysts had attached loosely, but only 40% attached stably to the epithelial cell surface. In contrast, 95% of the embryos attached stably to tissue culture plastic. Thus, weak attachment of a majority of the embryos was followed by stronger adhesion of a smaller proportion. Seventeen percent of the transferred blastocysts modified the epithelial cell surface with loss of MUC1 at the attachment site, extending variably to adjacent epithelial cells. Initially, stable attachment occurred without disruption to the integrity of the epithelial monolayer, but at later stages after the embryo had spread laterally, displacement of subjacent cells was observed. A modest increase in stable attachment, but no changes to MUC1 clearance, was observed after assisted hatching. After 24 h priming of Ishikawa cells by 17b-oestradiol (OE 2 ) followed by 72-h incubation with medroxyprogesterone acetate and OE 2 , stable attachment increased from 40 to 70%. Initial attachment is efficient either in the presence or in the absence of hormone; steroid treatment increased the incidence of stable attachment. Implantation failure is predicted to occur in this model when embryos fail to progress from initial to stable attachment.

Embryo–epithelium interactions during implantation at a glance

“…Present knowledge relies partly on in vitro models in which embryos, or trophoblastic spheroids derived from cell lines, primary tissue or embryonic stem cells, attach to 2D or 3D cultures of endometrial epithelium from either primary tissue or cell lines (Bentin-Ley et al, 1994;Boggavarapu et al, 2016;Hannan et al, 2009;Kang et al, 2015Kang et al, , 2014Lalitkumar et al, 2013;Lee et al, 2015;Singh and Aplin, 2015;Singh et al, 2010;Weimar et al, 2013). Recent observations of embryos attaching directly to cell-free surfaces demonstrate their self-organising potential (Deglincerti et al, 2016;Shahbazi et al, 2016).…”

Molecular mechanisms of membrane interaction at implantation

“…In vitro studies on implantation have shown that MUC1 is down-regulated specifically in the region of the implanting blastocyst in both humans and mice Singh et al, 2010). Low levels of MUC1 in the region of the implanting blastocyst during the window of implantation is hypothesized to be an important factor for successful embryo-endometrium interactions during implantation, since MUC1 otherwise conceals the expression of cell adhesion molecules that are important for blastocyst attachment (Fig.…”

“…Low levels of MUC1 in the region of the implanting blastocyst during the window of implantation is hypothesized to be an important factor for successful embryo-endometrium interactions during implantation, since MUC1 otherwise conceals the expression of cell adhesion molecules that are important for blastocyst attachment (Fig. 3) (Horne et al, 2005;Singh et al, 2010, Bastu et al, 2015. Moreover, paracrine signals from embryos appear to regulate MUC1 to alter local epithelial phenotypes, possibly via growth factors, including insulin-like growth factor (IGF) (Fluhr et al, 2008) and heparin-binding epidermal growth factor (HB-EGF) (Paria et al, 2001;Pochampalli et al, 2007;Lim and Dey, 2009).…”

“…Investigations have shown that Ishikawa cells are an exciting model for investigating endometrial integrins (Lessey et al, 1992;Castelbaum et al, 1997). They grow stably, form a monolayer, and maintain epithelial polarity, however, can exhibit some morphological changes following long periods of culture (Singh et al, 2010).…”

Endometrial receptivity in women with disoders in reproductive system

“…Исследователи выявили, что среди женщин с бесплодием отмечен низкий уровень муцина-1 в период «окна имплантации» [39,40]. В то же время лабораторные исследования де-монстрируют, что экспрессия муцина-1 должна снижаться непосредственно в месте импланта-ции бластоцисты [41]. Локальное падение син-теза муцина-1 «критично» для успешной им-плантации, что опосредовано модификацией клеточных мембран непосредственно в месте эмбрион-эндометриального диалога под влия-нием паракринных механизмов [42].…”