Differentiation of Human Endometrial Mesenchymal Cells to Epithelial and Stromal Cells by Seeding on the Decellularized Endometrial Scaffold

This study aimed to construct the endometrial-like structure by co-culturing of human mesenchymal endometrial cells and uterine smooth muscle cells in the decellularized scaffold. After decellularization of the human endometrium, cell seeding was performed by centrifugation of human mesenchymal endometrial cells with different speeds and times in 15 experimental subgroups. Analysis of residual cell count in suspension was done in all subgroups and the method with the lower number of suspended cells was selected for subsequent study. Then, the human endometrial mesenchymal cells and the myometrial muscle cells were seeded on the decellularized tissue and cultured for 1 wk; then, differentiation of the seeded cells was assessed by morphological and gene expression analysis. The cell seeding method by centrifuging at 6020 g for 2 min showed the highest number of seeded cells and the lowest number of residual cells in suspension. In the recellularized scaffold, the endometrial-like was seen with some protrusions on their surface and the stromal cells had shown spindle and polyhedral morphology. The myometrial cells almost were homed at the periphery of the scaffold and mesenchymal cells penetrated in deeper parts similar to their arrangement in the native uterus. The more expression of endometrial-related genes such as SPP1, MMP2, ZO-1, LAMA2, and COL4A1 and low-level expression of the OCT4 gene as a pluripotency marker confirmed the differentiation of seeded cells. Endometrial-like structures were formed by the co-culturing of human endometrial mesenchymal cells and smooth muscle cells on the decellularized endometrium.