Three-dimensional Sphere-forming Cells Are Unique Multipotent Cell Population in Dental Pulp Cells

Area of Science:

• Stem Cell Biology
• Tissue Engineering
• Regenerative Medicine

Background:

• Conventional mesenchymal stem cell (MSC) culture in monolayers inadequately mimics the in vivo microenvironment, leading to stemness loss.
• Three-dimensional (3D) sphere culture offers a more physiologically relevant model for stem cell research.
• The stemness and differentiation potential of spheres derived from dental pulp stem cells (DPSCs) remain underexplored.

Purpose of the Study:

• To investigate the stemness and multilineage differentiation capacity of DPSC spheres.
• To compare the in vivo tissue-forming ability of DPSC spheres versus adherent DPSC cultures.

Main Methods:

• DPSCs were cultured under nonadherent, serum-free conditions to promote sphere formation.
• Sphere and adherent cells were analyzed for pluripotency transcription factors, MSC markers, and multilineage differentiation potential (in vitro).
• In vivo mineralized tissue formation was assessed by subcutaneous transplantation in nude mice.

Main Results:

• DPSCs successfully formed spheres with robust expression of pluripotency factors and reduced MSC markers compared to monolayer cultures.
• DPSC spheres demonstrated enhanced in vitro multilineage differentiation capacity and increased expression of related genes.
• DPSC spheres exhibited superior in vivo mineralized tissue-forming ability compared to adherent DPSC cultures.

Conclusions:

• Sphere-forming cells represent a distinct multipotent cell population within DPSCs.
• DPSC spheres hold significant promise for advancing pulp tissue regeneration strategies.