Epithelial integration in tissue-engineered oral mucosa using RGDTFI-presenting micropatterned collagen scaffolds

Abstract

Objective To determine whether presentation of the integrin αvβ3/αvβ5-binding peptide motif RGDTFI on micropatterned collagen scaffolds improves epithelial coverage and stabilizes the epithelial–scaffold interface during fabrication of tissue-engineered oral mucosa (TEOM). Methods TEOMs were fabricated on micropatterned type I collagen scaffolds with or without RGDTFI presentation, pre-soaked with type IV collagen, laminin-511, or D-PBS. Epithelial coverage and interface integrity were evaluated by gross observation, histology, immunofluorescence analysis for laminin γ2 and integrins α6, β4, and β1, Ki-67 immunostaining, and ultrastructural analysis. Epithelial coverage was compared between the four scaffold conditions, and Ki-67 labeling indices were analyzed. Results RGDTFI presentation was associated with greater epithelial coverage and a more stable epithelial–scaffold interface on micropatterned collagen scaffolds, whereas non-modified scaffolds frequently had micro-gaps at the interface. TEOMs fabricated on RGDTFI-presenting scaffolds had stronger laminin γ2 deposition and integrin α6β4 expression in basal cells. Ultrastructural analysis revealed denser hemidesmosome-like electron-dense plaques, particularly in scaffolds pre-soaked with type IV collagen or laminin-511. Ki-67-positive basal cells were more frequent in rete ridge regions than in inter-ridge regions, and integrin β1 staining extended into suprabasal layers. Conclusions Presentation of RGDTFI on micropatterned collagen scaffolds was associated with improved epithelial coverage and a more stable epithelial–scaffold interface in TEOMs, together with altered basement membrane-associated staining and integrin expression. These findings suggest that this strategy may assist in improving the stability and reproducibility of TEOMs in in vitro oral mucosa studies.