Optimization of Extracellular Matrix for Culture of Vascularized Endometrial Triple-Cell-Type Organoids

Abstract:\nThe regeneration of the vascular system is crucial for the success of bioengineered implants. In this study, we aimed to optimize the extracellular matrix (ECM) for the culture of endometrial triple-cell-type organoids. We incorporated human umbilical vein endothelial cells (HUVECs), endometrial stromal cells (ESCs), and endometrial epithelial organoids (EEOs) into a hydrogel containing fibrin and matrigel. We found that the proportion of fibrin in the hydrogel influenced the formation of vascular networks by HUVECs and the adhesion of EEOs. We also investigated the stiffness and pore size of the hydrogel scaffold and determined that a hybrid hydrogel with a 1:1 ratio of fibrin to matrigel provided an optimal physical environment for organoid growth and vascular network establishment. Furthermore, we developed a dynamic 3D culture system using a microfluidic chip, which allowed for the formation of highly interconnected networks of organoids. The triple-cell-type endometrial organoids exhibited a capillary network-like structure and complex cellular interactions. Transcriptome sequencing confirmed the similarity between the organoids and in vivo endometrial tissues. Additionally, the organoids responded to sex hormones and induced endometrial regeneration in a mouse model. These findings demonstrate the potential of our optimized ECM and dynamic culture system for the study of endometrial biology and tissue engineering applications.