Controlling medium osmolality improves the expansion of human articular chondrocytes in serum-free media

To investigate the effects of medium osmolality on the expansion of human articular chondrocytes (HACs) with serum-free media (SFM), proprietary SFM of various osmolalities (290, 320, 350, 400, and 450 mOsm/kg), supplemented with components known to enhance chondrocyte growth, were constructed by the adjustment of NaCl concentration. It was found that HACs obtained better expansion in SFM at osmolalites lower than the average osmolality (400 mOsm/kg) of human articular cartilage in vivo. SFM at 290, 320, and 350 mOsm/kg showed similar growth, attaining up to a 1.55-fold increase in the proliferation rate compared with SFM at 400 mOsm/kg. Increasing SFM osmolality to 450 mOsm/kg resulted in a proliferation rate of 0.65-fold lower than at 400 mOsm/kg. Chondrogenic capacity was also examined via three-dimensional pellet cultures in a chondrogenic medium with HACs expanded in 320 and 400 mOsm/kg SFM. Biochemical, histological, and immunohistochemical analyses revealed similar glycosaminoglycan and collagen type II contents in both groups. Taken together, these results show that the expansion of HACs in SF cultures can be improved by adjusting the medium osmolality to be within the range of 290-350 mOsm/kg and that controlling medium osmolality during monolayer cultures does not deter the tissue-forming capability of the cells.