（1.School of Life Sciences, Shanghai University, Shanghai 200444, China) （2.Suzhou Institute of NanoTech and NanoBionics, Chinese Academy of Sciences, Suzhou 215123, China) （3. School of NanoTech and NanoBionics, University of Science and Technology of China, Hefei 230026, China）
As an important branch of stem cell, mesenchymal stem cells (MSCs) are derived from the mesoderm at early embryonic development, which have shown great potentials in tissue engineering and regenerative medicine applications due to their extensive sources, multilineage differentiation potential, low immunogenicity and selfrenewal ability. However, MSCs are still confronting challenges in clinical applications due to requirement of large quantity of cells for therapy, heterogeneity of cell quality, low survival ability after transplantation, the attenuation of MSCs characteristics in twodimensional (2D) culture and ambiguous mechanism of MSCs function for disease therapy. Threedimensional (3D) spheroid culture is widely regarded as a more preferable and closer physiological microenvironment for cells growth. Studies reported that 3D spheroid culture enhanced the characteristics of MSCs on cell survival, factor secretion, stemness maintenance, migration, antisenescence invitro and enhanced the capacities of antiinflammatory, angiogenesis, tissue repair and regeneration invivo. Therefore, 3D culture has broad application prospects in clinical application. In view of this, this article summarized the methods of 3D spheroid culture in vitro, and the spheroid biological characteristics and application briefly. Meanwhile, the prospects in future also have been discussed.