Fig. 6. Schematic illustration of the Wnt5a mimetic peptide–conjugated, porous HA hydrogels promoting the osteogenesis of seeded hMSCs by activating RhoA signaling and elevating the intracellular calcium level.

Presentation of the Wnt5a mimetic Foxy5 peptide by the hydrogel scaffold to Wnt receptors (Frizzled) and coreceptors on the membrane of seeded MSCs activates noncanonical Wnt signaling, leading to the activation of Dvl2 and downstream RhoA-ROCK2 signaling. The activation of RhoA signaling leads to enhanced cytoskeletal stability and actomyosin contractility (increased NMII) and therefore boosts mechanotransduction and the formation of focal adhesions (elevated vinculin expression). Meanwhile, the presentation of Foxy5 peptide mediates elevated intracellular calcium levels, contributing to the amplification of mechanosensing and osteogenic lineage commitment. The amplified mechanosensing of hMSCs residing in Foxy5 peptide–functionalized hydrogels is demonstrated by the increased YAP nuclear localization and up-regulated RhoA and ROCK2 expression, and these events promote the commitment of MSCs to the osteogenic lineage in the synthetic osteogenic niche microenvironment.