Transplantation in the rat of enzymatically and genipin-modified decellularized uterine tissue after mesenchymal stem cell recellularization; a proof-of-concept study.

Uterus transplantation can restore fertility in women with uterine factor infertility, but donor shortage, surgical complexity, and immunosuppression limit its wider use. Bioengineered uterine grafts may offer an alternative, yet animal studies have shown that large decellularized uterine scaffolds often lack sufficient mechanical durability and may not support complete tissue regeneration. This study used the rat model to investigate whether enzyme-treated genipin-crosslinked decellularized uterine patches could support long-term repair after full-thickness uterine wall injury. The patches were transplanted either alone or after mesenchymal stem cell recellularization, with particular focus on identifying the molecular profile during tissue remodeling and regeneration one and four months after transplantation. Methods included histology, collagen quantification, immunohistochemistry, biocompatibility testing, and digital droplet polymerase chain reaction analysis of genes linked to uterine identity, angiogenesis, proliferation, extracellular matrix turnover, fibrosis, and immune response. Genipin treatment increased scaffold stiffness and improved surgical handling without impairing scaffold biocompatibility or recellularization. Regeneration improved over time in recellularized grafts, but glandular regeneration remained absent and cellularity was sparse in parts of the graft. Gene expression indicated a continued remodeling state rather than mature uterine restoration. Reduced epithelial cadherin expression, increased mesenchymal vimentin expression, persistent transforming growth factor beta upregulation, and elevated proliferation and matrix-remodeling markers suggested an ongoing epithelial-to-mesenchymal transition repair process that may explain the incomplete epithelial and glandular regeneration. Immune-related gene expression further suggested that recellularized grafts showed signs of improved immune resolution. These findings suggest that genipin-crosslinked uterine scaffolds provide a robust structural platform for large uterine wall repair, but functional regeneration may be limited by a delayed epithelial maturation. By identifying the incomplete mesenchymal-to-epithelial transition and the glandular lineage commitment as key barriers, this study defines actionable targets for the next generation of bioengineered uterine grafts.