OBJECTIVE: To determine if the various three-dimensional structures of bioscaffolds affect wound healing by investigating the efficacy of different pomine-derived urinary bladder matrix (UBM) structures in treating murine diabetic wound healing. METHODS: The authors studied three different UBM structures: particulate (pUBM), one-layer freeze-dried sheet (fdUBM), and tree-layer laminated sheet (ImUBM). Scanning electron microscopy images of the structures were used to calculate a wound-exposed surface-area-to-volume ratio. A 1.0 x 1.0-cm full-thickness dorsal wound was excised on 93 clb/ mice. Mice were either untreated (blank, n = 15), treated with one UBM structure (pUBM, n =15; fdUBM, n = 15; ImUBM, n =15), or treated with a combination of either the one- or three-layer sheet over the particulate matrix (fdUBM + pUBM, n = 15; ImUBM + pUBM, n = 15). The authors obtained macroscopic images of the wounds and harvested tissues for analyses at multiple time points. RESULTS: The surface area available to interact with the wound was highest in the pUBM group and lowest in the ImUBM group. Greater wound bed thickness was noted in the fdUBM, fdUBM + pUBM. and ImUBM groups compared with the blank group. Cellular proliferation was significantly higher in the fdUBM and fdUBM pUBM groups than in the blank group. The ImUBM + pUBM group had the highest collagen deposition. The pUBM group induced significantly higher leukocyte infiltration compared with the ImUBM, ImUBM pUBM, and blank groups. Microvessel density was highest in the fdUBM + pUBM group. Significant differences in the wound closure rate were noted between the blank group and the fdUBM and fdUBM + pUBM groups. CONCLUSIONS: Assessment of the three UBM bioscaffold structures highlighted differences in the wound-exposed surface area. Variations in wound healing effects, including collagen deposition, cellular proliferation, and angiogenesis, were identified, with combinations of the structures displaying synergistic effects. This study serves as a platform for future scaffold design and offers promising evidence of the benefits of combining various structures of scaffolds.