(761f) Microenvironment Formation and Enhancement of hMSC Osteogenesis In Hydroxyapatite-Chitosan-Gelatin Hydrogel

Tissue engineered bone grafts require both an osteogenic cell source and scaffolds that can support the regeneration of healthy bone.  Hydroxyapatite (HA), chitosan (C), and gelatin (G) have chemical similarity to the structural components of natural bone and have been frequently used as bone scaffolds.  Human mesenchymal stem cells (hMSCs) are inducible osteoprogenitors and a promising cell source for bone regeneration.  In this study, the formation of extracellular microenvironment (ECM) in the composite hydroxyapatite-chitosan-gelatin (HCG) hydrogel and its impact on hMSC fate are investigated.  The association of hydroxyapatite, chitosan, and gelatin forms a biodegradable hydrogel with 60 wt% water and an initial stiffness of ~ 20 kPa.  Preconditioning in serum containing media results in the formation of micro/nanopores in the HCG gels, and increased ECM protein adsorption.  Expression of integrin α₂β₁ and α₅β₁ coincides with ECM enrichment, suggesting the enhanced cell-ECM interactions that contribute to hMSC migration and proliferation.  The elevated expression of bone marker proteins and genes in the absence of chemical induction further demonstrates HCG’s intrinsic osteogenic inductive properties.  The results show that ECM proteins can effectively functionalize HCG hydrogel and form an instructive microenvironment that promotes bone construct development from hMSCs.