Influence of hyaluronic acid modification on CD44 binding towards the design of hydrogel biomaterials

Hyaluronic acid (HA) is a linear polysaccharide of d-glucuronic acid and N-acetyl-d-glucosamine that is native to many tissues and interacts with cells via cell-surface receptors (e.g., CD44). HA has been extensively explored as a chemically-modified macromer for crosslinking into biomaterials, such as hydrogels and macroporous scaffolds. However, the influence of the extent and type of HA modification on its binding to CD44 is not well understood or quantified. To address this, we modified HA at either the carboxylic acid or the primary alcohol with various chemical groups (e.g., norbornenes, methacrylates) and magnitudes (~10, 20, or 40% of disaccharides) and then characterized binding in both soluble and hydrogel forms. HA binding to CD44 immobilized on plates or presented by cells was influenced by the extent and type of its modification, where increased modification (i.e., ~40%) generally decreased binding. The adhesion of CD44-modified beads to hydrogels as measured by atomic force microscopy revealed a similar trend, particularly with decreased adhesion with hydrophobic modifications to the carboxylic acid. Further, the chondrogenesis of mesenchymal stromal cells when encapsulated in hydrogels fabricated from modified HA macromers was reduced at high modification, behaving similarly to inert hydrogel controls. This work suggests that the types and extents of modification of polysaccharides are important factors that should be considered in preserving their biological function when processed as hydrogels.