Facile Decellularization of Stem Cell Spheroids by Integrating Bioactive Nanofibers for Enhanced Wound Healing

Abstract Decellularized 3D spheroids of stem cells may address challenges associated with the direct transplantation of cellular components; however, decellularization is inefficient in large spheroids due to the dense cellular architecture, which restricts detergent infiltration and the preservation of biomolecules. In this study, a strategy incorporating anti‐oxidative polydopamine (PD)‐coated nanofibers into human adipose‐derived stem cell (hADSC) spheroids is developed to facilitate solution infiltration and improve the retention of decellularized residual biomolecules for wound healing applications. Compared with cell‐only spheroids, this approach achieves >99% deoxyribonucleic acid (DNA) removal while substantially preserving key extracellular matrix proteins. Consequently, the decellularized spheroids with PD‐coated nanofibers and preserved residual proteins (dPS) reduce intracellular reactive oxygen species, suppress pro‐inflammatory M1 macrophage polarization, promote M2 macrophage polarization, and enhance endothelial cell adhesion with tubule formation. In two types of murine models, dPS implantation attenuates immune activation, enhances tissue integration, and significantly accelerates wound healing, with pronounced neovascularization and hair follicle formation. This nanofiber‐incorporated decellularized spheroid platform can offer a facile but robust method for advanced wound repair.