Dental Pulp Stem Cell-Derived Multicellular Microtissue Spheres: A Synergistic Strategy for Dental Pulp Regeneration

Restoring pulp tissue to its natural state is the ultimate goal of pulp regeneration, during which vascularization and neuralization of regenerated pulp are key factors. Hence, this study aimed to construct multicellular microtissue spheres derived from dental pulp stem cells to promote the vascularization and neuralization of regenerated dental pulp tissue.

First, we induced dental pulp stem cells (DPSCs) to differentiate into endothelial-like cells and neural-like cells. Then, 7 types of multicellular microtissue spheres were constructed from DPSCs, endothelial-like cells and neural-like cells, and their gross morphology, tissue morphology, and endothelial cell marker and neuronal marker expression were evaluated. Additionally, the microtissue spheres were placed in rabbit molars, which were implanted subcutaneously into mice to assess dental pulp tissue regeneration.

Internally dense microtissue spheres with a stable morphology were successfully constructed via the low adhesion method. The spheres containing endothelial-like cells displayed enhanced CD31-positive expression, and those with neural-like cells showed elevated expression of the neuronal marker Nestin. Cell activity was maintained in all 7 spheres, and compared with those derived from DPSCs only, the cell survival rates in the spheres constructed from endothelial-like and neural-like cells together or separately combined with DPSCs were greater. Neovascularized pulp-like tissue with neuronal marker-positive cells appeared in the root canals of each group, indicating that these spheres promoted pulp regeneration.

The results of this study indicate that DPSC-derived multicellular microtissue spheres facilitated neovascularization and expressed neuronal markers in newly formed pulp-like tissue, providing an experimental and theoretical basis for their application in pulp regeneration.