Novel 3D embryo implantation model within macroporous alginate scaffolds

Abstract

Background: Implantation failure remains an unsolved obstacle in reproductive medicine. Previous studies have indicated that estrogen responsiveness, specifically by estrogen receptor alpha (ERα), is crucial for proper implantation. There is an utmost need for a reliable in vitro model that mimics the events in the uterine wall during the implantation process for studying the regulatory mechanisms governing the process. The current twodimensional and hydrogel-based in vitro models provide only short-term endometrial cell culture with partial functionality. Results: Endometrial biopsies showed an increase in E-cadherin expression on the typical window of implantation of fertile women, compared to negligible expression in recurrent implantation failure (RIF) patients. These clinical results indicated E-cadherin as a marker for receptivity. Three-dimensional (3D) macroporous alginate scaffolds were the base for epithelial endometrial cell-seeding and long-term culture under hormone treatment that mimicked a typical menstrual cycle. The RL95-2 epithelial cell culture in macroporous scaffolds was viable for 3 weeks and showed increased E-cadherin levels in response to estrogen. Human choriocarcinoma (JAR) spheroids were used as embryo models, seeded onto cell constructs and successfully adhered to the RL95-2 cell culture. Moreover, a second model of HEC-1A with low ERα levels, showed lower E-cadherin expression and no JAR attachment. Ecadherin expression and JAR attachment were recovered in HEC-1A cells that were transfected with ERα plasmid. Conclusions: We present a novel model that enables culturing endometrial cells on a 3D matrix for 3 weeks under hormonal treatment. It confirmed the importance of ERα function and E-cadherin for proper implantation. This platform may serve to elucidate the regulatory mechanisms controlling the implantation process, and for screening and evaluating potential novel therapeutic strategies for RIF.

TNF-α Regulated Endometrial Stroma Secretome Promotes Trophoblast Invasion

“…Thus, there is a need to establish multicellular models that mimic the process of implantation. To overcome these limitations, several investigators have developed in vitro models of embryo/endometrium interaction where two or more cellular components are evaluated (69,(78)(79)(80)(81). We established a 3D in vitro model of the human endometrium consisting of hESC within an extracellular matrix and trophoblast cells as the main component of a BLS that lacks the inner cell layer (19,33).…”

Opportunities involving microfluidics and 3D culture systems to the in vitro embryo production

“…A recent study used a porous alginate scaffold for 3D culture of human epithelial endometrial cells, in which cells remained viable and hormone-responsive for three weeks under hormonal treatment, creating a receptive and non-receptive environment for embryo implantation. This model could potentially use patients' cells to investigate individual mechanisms that may lead to pregnancy failure and enable personalized therapies ( Stern-Tal et al, 2020 ).…”

Novel microarchitecture of human endometrial glands: implications in endometrial regeneration and pathologies

“…Owing to both technical and ethical limitations, it is not possible to study human embryo implantation in vivo. Current knowledge about implantation has mainly been obtained from animal models and in vitro 2D monolayer culture systems, which fail to represent the 3D architecture and physiology of the human process ( Cha et al , 2012 ; Stern-Tal et al , 2020 ). Recent data revealing the novel 3D architecture of the endometrial epithelial compartment has now shed light on possible new explanations for the pathogenesis of many poorly understood endometrium-related conditions, including RPL and RIF ( Tempest et al , 2020 ; Yamaguchi et al , 2021 ).…”