Isolation and Cultivation of Human Endometrial Epithelial Cells

Objectives: This work aimed to develop and optimise methods for obtaining and culturing human endometrial epithelial cells from a non-invasive source: menstrual blood. The study aims to create personalized in vitro endometrial models to study embryo implantation mechanisms, search for markers of endometrial receptivity, and develop new approaches to treat gynecological diseases, including infertility. Methods: Menstrual blood collected from healthy donors was used to obtain endometrial cells. Endometrial fragments were isolated by filtration and enzymatic treatment. Cell culture was carried out under various conditions: as a monolayer (2D), tissue culture and organoids (3D). Immunocytochemical staining for the markers cytokeratin and E-cadherin was used to confirm the epithelial phenotype. To investigate endocytosis of the epidermal growth factor receptor (EGFR) in tissue culture, fluorescence-labeled EGF was used. Results: Menstrual blood is an available source for producing viable endometrial cells. The resulting endometrial tissue culture preserves tissue architecture and can serve as a model for the study of endocytosis of the epidermal growth factor receptor (EGFR). Cultivation of endometrial epithelium cells in the form of organoids made it possible to preserve the epithelial phenotype and proliferative activity of cells for a long time. Organoids demonstrated the ability to self-organize and form single-layer cell structures, which confirms their suitability for modeling the processes occurring in the endometrium in vivo. At the same time, cultivation under 2D conditions led to rapid aging of cells and loss of their functional properties. Conclusion: The developed methods for culturing endometrial epithelial cells in the form of tissue culture and organoids open up new possibilities for studying the mechanisms of embryo implantation and searching for markers of endometrial receptivity. The findings have important implications for the development of personalized medicine, in particular for improving the effectiveness of assisted reproductive technology (ART) programs and developing new therapeutic approaches for the treatment of gynecological diseases.