Abstract:

BACKGROUND: Implantation failure remains an unsolved obstacle in reproductive medicine. Previous studies have indicated that estrogen responsiveness, specifically by estrogen receptor alpha (ER$\alpha$), 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 two-dimensional 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 ER$\alpha$ levels, showed lower E-cadherin expression and no JAR attachment. E-cadherin expression and JAR attachment were recovered in HEC-1A cells that were transfected with ER$\alpha$ 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 ER$\alpha$ 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.