Endometrial organoid and stromal cultures demonstrate donor-derived cellular origin of the endometrium after uterus transplantation.

The human endometrium undergoes cyclical regeneration during the menstrual cycle, a process thought to be driven primarily by multipotent stem cells located within the basalis layer. Previous studies have proposed that bone marrow-derived cells (BMDCs) may contribute to endometrial regeneration; however, this hypothesis remains controversial due to methodological limitations in distinguishing hematopoietic cells from resident endometrial epithelial or stromal populations. To investigate the potential contribution of BMDCs to endometrial regeneration, we used uterus transplantation (UTx) as a unique donor-recipient model and established patient-derived 3D endometrial organoid (PDO) and 2D stromal cell cultures from UTx recipients. These culture systems enable selective expansion of epithelial and stromal compartments while excluding hematopoietic cells, thereby allowing accurate assessment of cellular origin through donor-recipient genotyping. Samples were collected from four UTx cases spanning up to 7 years, including biopsies obtained before and after pregnancy as well as tissue collected at uterus explantation. Short tandem repeat (STR) profiling was performed on genomic DNA isolated from donor and recipient blood, endometrial organoids, and stromal cell cultures to determine cellular genotype. Endometrial epithelial organoids derived from all UTx cases exhibited consistent morphological features comparable to healthy endometrium and demonstrated long-term expandability. STR analysis revealed complete concordance between the donor genotype and both epithelial and stromal cells of the transplanted uteri, with no evidence of recipient-derived cells or BMDC contribution to the endometrium. These findings indicate that the cellular composition of the endometrium in transplanted uteri remains donor-derived over extended follow-up, including after pregnancy. Together, these results provide strong evidence against a substantial contribution of BMDCs to endometrial regeneration and establish patient-derived endometrial organoid and stromal cultures as powerful model systems for investigating endometrial biology and regenerative mechanisms.