In this work, cell-responsive polysaccharide hydrogels were prepared by a simple procedure based on the sequential bioconjugation and cross-linking of the polysaccharide backbone with bioactive peptides and poly(ethylene glycol)-bis(thiol) (PEG-(SH)2), respectively. Using thiol-ene reactions, we successfully functionalized hyaluronic acid (HA) and carboxymethylcellulose (CMC) with short and long peptides (5-mer and 15-mer derivatives, respectively) derived from adhesive proteins of bone extracellular matrix. The resulting HA-peptide and CMC-peptide conjugates with varying degrees of substitution were then carefully characterized by (1)H NMR spectroscopy to precisely control the peptide density into the hydrogels cross-linked with PEG-(SH)2. Preosteoblast seeded on the hydrogels with controlled identical stiffness spread in a manner that was strongly dependent on ligand density. Surprisingly, increasing the density of the adhesive peptide anchors did not result in a plateau of initial cell spreading but rather in a bell-shaped cell response that varies with the nature of both polysaccharide backbone and functional peptide. Placing the cells under optimal conditions for cell/hydrogel interaction, we showed that in HA hydrogels, the polysaccharide moiety is not solely a passive scaffold that presents the active peptides but is an active player in cell microenvironment to control and sustain cell activity.