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Plant virus nanoparticle (VNP) is a useful platform with biocompatibility and versatile monodisperse protein structures that can be engineered with bioactive cues, offering opportunities to functionalize bioinert hydrogels for tissue engineering. The hypothesis in this study was that osteogenesis of human mesenchymal stem cells (hMSCs) and biomineralization could be enhanced by incorporating VNPs, which were engineered with osteogenesis-associated or cell-adhesive peptides, into cell-laden hydrogels. Cellular responses to VNPs were examined in both 2D and 3D cultures, including VNP-cell distribution, cell attachment, morphology, and osteogenesis. VNP-laden agarose or agarose-collagen hydrogels were characterized in terms of release rate, mineralization effect, mechanical properties, and usage as bioink. The results revealed enhanced osteogenic differentiation when cells were cultured on VNP-coated surfaces, and attachment of VNPs to cells as well as at least 84 % of VNP retention were observed in hydrogels. Mineralization effect was pronounced in VNP-laden hydrogels, which also demonstrated the superiority of the enriched peptides on VNPs over free peptides and VNPs with fewer peptides. Finally, VNP-laden hydrogels showed good bioprinting reproducibility. Prospectively, VNPs could be conjugated with vasculogenesis-inducing factors, thereby potentially induce more effectively 3D-printed in vitro pre-vascularization in hydrogels.
