Osmolarity effects on bovine articular chondrocytes during three-dimensional culture in alginate beads

Summary Objective With the development of engineered cartilage, the determination of the appropriate culture conditions is vital in order to maximize extracellular matrix synthesis. As osmolarity could affect the fate of chondrocytes, the purpose of this study was to determine the effects of osmolarity on chondrocytes during relatively long-term culture. Design Bovine articular chondrocytes were cultured in alginate beads in a biocarbonate free system at 280, 380 and 550 mOsm at pH 7.4 for up to 12 days, respectively. Cell volume, intracellular pH (pHi ), cell number, glucosaminoglycan (GAG) and collagen retention were measured at day 5 and 12. Cell viability and volume were monitored over the 12 days of culture. Results By day 5 and 12, compared to the cell volume at 380 mOsm, around 20% ( P < 0.01) swelling and 15% ( P < 0.05) shrinkage were observed when the cells were cultured at 280 and 550 mOsm. The pHi over the 12 days of culture varied with osmolarity of the culture medium. In comparison with fresh cells, pHi became slightly more acidic by 0.15 pH units at 280 mOsm at day 5. However, by day 12, an alkalization of pHi , by 0.2 pH units, was noted. A higher proliferation rate was seen at 280 mOsm than at other osmolarities while less GAG was produced. Conclusions Chronic exposure to anisotonic conditions results in cell swelling at 280 mOsm and shrinkage at 550 mOsm. The osmolarity of 280 mOsm appears to encourage proliferation of chondrocytes, but inhibits matrix production.