Hyaluronic Acid Dictates Chondrocyte Morphology and Migration in Composite Gels

Tissue equivalent collagen–hyaluronic acid-based hydrogels are widely used for cartilage tissue engineering; however, not much importance has been given to investigate how cellular responses are altered with varying concentrations of hyaluronic acid in gels. In this study, different concentrations of hyaluronic acid dialdehyde (HAD) were combined with collagen to fabricate collagen–HAD composite (CH) gels, and the influence of HAD on cell shape, migration, viability, cytoskeletal organization, and gel contraction was examined. The microstructure and the mechanical strength of the composite gels were altered by varying HAD concentrations. Morphology of chondrocytes cultured on CH gels showed a significant increase in their aspect ratio and decrease in number of cell protrusions with increase in concentration of HAD. The organization of the cytoskeleton at the cellular protrusions was vimentin localized at the base, microtubules at the tip, and actin localized throughout the cell body. Changes in HAD concentrations altered hydrogel mechanical strength, cytoskeletal organization, and formation of cellular protrusions, all of which contributed to changes in cell morphology and migration. These changes were more evident in 3D cell-encapsulated gels than chondrocytes cultured over the 2D gels. However, viability of cells and matrix contraction, staining for adhesion protein vinculin, and hyaluronic acid receptor CD44 remained similar in all CH compositions. The changes in cell responses further influenced extracellular matrix deposition during in vitro culture. Cell responses in low HAD gels mimic the cellular behavior in damaged cartilage, whereas those in high HAD gels resembled the behavior in healthy cartilage tissue. Our study illustrates the importance of careful formulations of hydrogel compositions in designing biomimetic matrices that are used as in vitro models to study chondrocyte behavior.