Human Periodontal Ligament- and Gingiva-derived Mesenchymal Stem Cells Promote Nerve Regeneration When Encapsulated in Alginate/hyaluronic Acid 3D Scaffold

Repair or regeneration of damaged nerves is still a challenging clinical task in reconstructive surgeries and regenerative medicine. Here, we demonstrate that periodontal ligament stem cells (PDLSCs) and gingival mesenchymal stem cells (GMSCs) isolated from adult human periodontal and gingival tissues assume neuronal phenotype in vitro and in vivo via a subcutaneous transplantation model in nude mice. PDLSCs and GMSCs were encapsulated in a three-dimensional scaffold based on alginate and hyaluronic acid hydrogels capable of sustained release of human nerve growth factor (NGF). We demonstrate that, the elasticity of the hydrogels affected the proliferation and differentiation of encapsulated MSCs within scaffolds. Moreover, we observed that PDLSCs and GMSCs were stained positive for βIII-tubulin, while exhibiting high levels of gene expression related to neurogenic differentiation (βIII-tubulin and GFAP) via qPCR. Western blot analysis showed the importance of the elasticity of the matrix and the presence of NGF in the neurogenic differentiation of encapsulated MSCs. In vivo, immunofluorescence staining for neurogenic specific protein markers confirmed islands of dense positively stained structures inside transplanted hydrogels. To our knowledge, this study is the first demonstration of the application of PDLSCs and GMSCs as promising cell therapy candidates for nerve regeneration.