Abstract:

The dental mesenchymal stem cells （DMSCs） are mesenchymal stem cells derived from the neural crest ectoderm and have exceptional self-renewal and multilineage differentiation capabilities. The DMSCs are extensively used in tissue engineering and regenerative medicine research. The DMSCs can be expanded in vitro on a large scale to meet the needs of research and therapy by three-dimensional culture technique. Compared with traditional two-dimensional cell culture techniques，three-dimensional culture more effectively simulates the structure and microenvironment that the stem cells encounter in vivo，providing simultaneous spatial and temporal regulation of the proliferation and differentiation of the stem cells. Various three-dimensional in vitro culture techniques have been developed in recent years. Hanging drop culture is straightforward， but controlling the tissue culture environment is challenging； microfluidic chips offer better control over cellular parameters， but are costly and face the technical platform challenges，so their widespread application is limited； magnetic levitation culture is cost-effective and easy to perform with the rapid cell spheroid formation， but its magnetization effect make it unsuitable for the quantitative analysis. Other three-dimensional culture methods include rotating cell culture systems， centrifugation for spheroid culture， overlay culture and artificial scaffold methods，and they all have their own advantages and limitations. This review encompasses the different methods of three-dimensional culturing of DMSCs and their applications in various tissue regeneration and disease treatment，and provide the reference for the precise regulation of the function of the DMSCs and the research of regenerative medicine.

Key words: Dental mesenchymal stem cell, Sphere culture, Stemness maintenance, Tissue engineering, Regenerative medicine