Xenofree generation of oral keratinocytes from induced pluripotent stem cells derived from adult human fibroblasts for oral mucosal regeneration

This study investigated the potential of xenofree generation of oral keratinocytes from induced pluripotent stem cells (iPSCs) derived from adult oral and skin fibroblasts for oral mucosal regeneration. Using an extracellular matrix-based protocol, iPSCs were differentiated into cells with a phenotype and molecular profile of oral epithelium, proven through morphological and molecular analyses. Differentiation to epithelial squamous lineage was achieved in serum-containing and xenofree conditions, although the former showed higher efficiency. The differentiated cells successfully formed pluristratified squamous epithelial tissues in 3D organotypic cultures, mimicking oral mucosa. In vivo tests using an immunodeficient mouse model validated the viability of multilayered squamous epithelial-like tissues, with distinct oral-specific markers identified in tissues derived from gingival fibroblast iPSCs. These findings demonstrate the feasibility of using iPSCs to create functional oral mucosal sheets, highlighting their potential for clinical applications in regenerative therapies, though further optimization is necessary to enhance differentiation efficiency under xenofree conditions.