Decellularization and enzymatic preconditioning of bovine uterus for improved recellularization

Abstract

Background Uterus tissue engineering aims to repair a dysfunctional uterus that causes infertility, e.g., after significant scarring from benign or malign resection procedures. Decellularized uterine tissue provided regenerative support in several animal models as a biocompatible natural extracellular matrix (ECM) derived scaffold after uterine damage. However, variations in decellularization protocols and species used limit conclusive evidence and translational progress. Hence, a species-independent decellularization protocol could facilitate preclinical research. Therefore, we investigated if our developed sheep uterus decellularization protocol was species-independent and effective for the significantly larger bovine uterus. We further assessed if there were any negative post transplantation immunological consequences from the metalloproteinases 2 and 9 (MMP 2, MMP 9) treatment that was used as a preconditioning treatment to significantly improve scaffold recellularization after decellularization. Methods Bovine uterus was decellularized using sodium deoxycholate, and the remaining ECM was quantitatively assessed for DNA, protein, and ECM components. The morphology and physical attributes were examined by immunohistochemistry, electron microscopy, and mechanical tests. Scaffold biocompatibility, bioactivity, and angiogenic properties were assessed with the chorioallantoic membrane assay (CAM) and the immune response following transplantation of MMP treated scaffolds was compared with untreated scaffolds in a rat model. The in vitro recellularization efficiency of the scaffolds was also assessed. Results The decellularization protocol was effective for bovine uterus. The MMP treatment did not negatively affect scaffold immunogenicity in vivo, while the treatment potentiated mesenchymal stem cell recellularization in vitro. Furthermore, the decellularization protocol generated biocompatible and angiogenic uterine scaffolds. Conclusion Bovine uterus was successfully decellularized using previously established protocols. These results confirm earlier findings in the sheep model and further indicate that MMP treatment may be beneficial. The results further conclude the development of a species-independent, reproducible, and biocompatible scaffold generation protocol that can provide an important element for successful translational research.