Bilayer Biomimetic Scaffolds Loaded with Mesenchymal Stem Cell Secretomes Promote Diabetic Wound Healing

Diabetic ulcers are among the most common and challenging complications of diabetes mellitus, and effective therapeutic strategies remain elusive. While stem cell secretome (SCS)-based therapy has attracted considerable attention due to its regenerative potential, its direct application is hindered by low bioavailability and rapid diffusion at the wound site. To address these limitations, we designed a bilayer bacterial cellulose-gelatin (Bi-BCG) scaffold inspired by the hierarchical structure of native skin. This scaffold features a compact bacterial cellulose (BC) upper layer with nanoscale porosity and a porous BCG lower layer with pore sizes of ~52 μm, optimized for SCS delivery. The Bi-BCG scaffold demonstrated a water vapor transmission rate of 2384 g/(m²·24 h) and exhibited significantly improved SCS retention capacity while maintaining high fluid absorption, outperforming monolayer BCG scaffolds. Functionally, human umbilical cord-derived mesenchymal stem cell (hUCMSCs)-derived secretomes significantly enhanced the proliferation (by up to 70.7%) and migration of skin fibroblasts under high-glucose conditions, promoted vascular endothelial cell proliferation (increasing Ki-67+ cells from 25.87% to 46.89%) and angiogenic network formation, and effectively suppressed macrophage-mediated inflammatory responses and oxidative stress. In vivo, the combination of SCSs with the Bi-BCG scaffold exhibited a clear synergistic effect, achieving a wound closure rate of 78.8% by day 10 and promoting superior structural restoration with well-organized collagen deposition, outperforming either treatment alone. These findings underscore the potential of the Bi-BCG scaffold combined with SCSs as an effective strategy for enhancing diabetic wound healing.