The interplay between chemical conjugation and biologic performance in the development of alginate-based 3D matrices to mimic neural microenvironments.

Biofunctionalization of polysaccharides is a widely used strategy for obtaining extracellular matrix (ECM)-mimicking biomaterials. Still, commonly employed chemistries present low reaction yields and the selection of the most adequate bioconjugation route can be challenging. Herein, we compared the performance of carbodiimide and reductive amination chemistries for the synthesis of tailored peptide-alginate hybrid hydrogels as neural tissue mimics. Reductive amination dramatically improved the peptide grafting efficiency, with yields of 50 % vs. 20 %, allowing 1.5 to 3-fold higher incorporation of cell-adhesive and matrix-metalloproteinases (MMP)-sensitive peptides, respectively. The conjugation of dual-end reactive MMP-sensitive peptides promoted a partial crosslinking, allowing adjusting gelation, stiffness, and degradability of hydrogels. Such parameters depended on the glycosidic position where the bioactive peptide binds, determined by the adopted chemical strategy, and this significantly impacted the biological response. Reductive amination provided softer (50-210 Pa) and fully degradable (60-100 % weight loss) hydrogels, depending on the amount of peptide in formulation, contrasting with the stiffer (400 Pa) and less degradable (40 % weight loss) carbodiimide-based hydrogels. Due to their opened polymer chain and increased peptide availability to cells, such hydrogels better supported the 3D culture of primary astrocytes, which present high complexity and process branching, allowing the development of improved brain ECM-mimicking systems.