Effect of Various Ratios of Co‐Cultured ATDC5 Cells and Chondrocytes on the Expression of Cartilaginous Phenotype in Microcavitary Alginate Hydrogel

ABSTRACT The present study introduced a direct co‐culture of mouse ATDC5 cells and primary porcine chondrocytes into a microcavitary hydrogel, which possessed advantages in promoting the growth of chondrocytes and retaining the phenotype. These two types of cells were encapsulated with gelatin microspheres in alginate hydrogels in either of the three ratios (3:1, 1:1, or 1:3 of ATDC5 cells to chondrocytes) and cultured in chondrogenic medium for 28 days. Simultaneously, the single encapsulation of ATDC5 cells or chondrocytes was set as a control. Cell Counting Kit‐8 (CCK‐8), real‐time PCR, and immunohistochemistry staining were used to evaluate the effect of various ratios of co‐cultured ATDC5 cells and chondrocytes on the expression of the cartilaginous phenotype. The CCK‐8 data indicated that the ratio of 3:1 group had an outstanding ability of cell growth. The other results demonstrated that higher the ATDC5 ratios and longer the culture duration, greater the expression of cartilage‐specific genes (including type II collagen and aggrecan) and more the synthesized cartilaginous extracellular matrix. Also, the Western blot analysis suggested that p44/42 MAP Kinase was involved in cell proliferation. However, due to the direct co‐culture of the two cell types, the underlying mechanism necessitates further investigation. Overall, the co‐culture system in microcavitary hydrogel improved the effect of chondrogenesis and exhibited promising strategy for cartilage tissue engineering therapies. J. Cell. Biochem. 118: 3607–3615, 2017. © 2017 Wiley Periodicals, Inc.