Three-Dimensional Biofabrication Models of Endometriosis and the Endometriotic Microenvironment

Abstract

Endometriosis occurs when endometrial-like tissue grows outside the uterine cavity, leading to pelvic pain, infertility, and increased risk of ovarian cancer. The present study describes the optimization and characterization of cellular spheroids as building blocks for Kenzan scaffold-free method biofabrication and proof-of-concept models of endometriosis and the endometriotic microenvironment. The spheroid building blocks must be a specific diameter (~500 m), compact, round, and smooth to withstand Kenzan biofabrication. Under optimized spheroid conditions for biofabrication, the endometriotic epithelial-like cell line, 12Z, expressed high levels of estrogen-related genes and secreted high amounts of endometriotic inflammatory factors that were independent of TNF stimulation. Heterotypic spheroids, composed of 12Z and T-HESC, an immortalized endometrial stromal cell line, self-assembled into a biologically relevant pattern, consisting of epithelial cells on the outside of the spheroids and stromal cells in the core. 12Z spheroids were biofabricated into large three-dimensional constructs alone, with HEYA8 spheroids, or as heterotypic spheroids with T-HESC. These three-dimensional biofabricated constructs containing multiple monotypic or heterotypic spheroids represent the first scaffold-free biofabricated in vitro models of endometriosis and the endometriotic microenvironment. These efficient and innovative models will allow us to study the complex interactions of multiple cell types within a biologically relevant microenvironment.

Endometriotic Organoids: A Novel In Vitro Model of Endometriotic Lesion Development

“…Interestingly, both of these culture models formed self-organizing spheroid structures, but separate epithelial and stromal cell compartments were evident only in ultra-low attachment conditions (27); indicating that ultra-low attachment culture might be more suitable for developing organoids with multiple cell types.…”

“…Therefore, inclusion of both cell types more closely resembles the pathophysiology of endometriosis and advances the potential for in vitro discovery. Ultra-low attachment plates and hanging drop culture have been used to generate spheroids with 12Z cells, immortalized stromal cells or primary ectopic stromal cells, or a combination of epithelial and stromal cells to investigate endometriosis (27,28).…”

“…Therefore, inclusion of both cell types more closely resembles the pathophysiology of endometriosis and advances the potential for in vitro discovery. Ultra-low attachment plates and hanging drop culture have been used to generate spheroids with 12Z cells, immortalized stromal cells or primary ectopic stromal cells, or a combination of epithelial and stromal cells to investigate endometriosis ( 27, 28 ). Interestingly, both of these culture models formed self-organizing spheroid structures, but separate epithelial and stromal cell compartments were evident only in ultra-low attachment conditions ( 27 ); indicating that ultra-low attachment culture might be more suitable for developing organoids with multiple cell types.…”

Three-dimensional culture models of human endometrium for studying trophoblast-endometrium interaction during implantation

“…The assembloids could capture the unique endometrial epithelial and stromal cell interactions with proper polarity and expression of steroids receptors, and could be used for drug screening. Another heterotypic assembloid model using the endometriotic cell line TZ2 and the human endometrial stromal cell line (T-HESC) was developed to study endometriosis [99]. The model enabled bio-fabrication of endometriosis tissue without the use of scaffold.…”

Endometriosis, the Silent Disease: Molecular Targets, Active Principles, and Drug Delivery Systems

“…The pathophysiology of endometriosis is multifactorial; a selection of pathways is reported below. The historical lack of endometriosis models makes the identification of new targets and therapeutic agents challenging; however, in vitro [21–28] and in vivo [29–31] models of endometriosis are emerging. Table 1 shows a selection of active principles in different stages of clinical and preclinical development for endometriosis treatment, some of which were reviewed recently [42,43] .…”