Inflammatory Licensed hMSCs Exhibit Enhanced Immunomodulatory Capacity in a Biomaterial Mediated Manner

Craniomaxillofacial (CMF) bone injuries represent particularly challenging environments for regenerative healing due to their large sizes, irregular and unique defect shapes, angiogenic requirements, and mechanical stabilization needs. These defects also exhibit a heightened inflammatory environment that can complicate the healing process. This study investigates the influence of the initial inflammatory stance of human mesenchymal stem cells (hMSCs) on key osteogenic, angiogenic, and immunomodulatory criteria when cultured in a class of mineralized collagen scaffolds under development for CMF bone repair. We previously showed that changes in scaffold pore anisotropy and glycosaminoglycan content can significantly alter the regenerative activity of both MSCs and macrophages. While MSCs are known to adopt an immunomodulatory phenotype in response to inflammatory stimuli, here, we define the nature and persistence of MSC osteogenic, angiogenic, and immunomodulatory phenotypes in a 3D mineralized collagen environment, and further, whether changes to scaffold architecture and organic composition can blunt or accentuate this response as a function of inflammatory licensing. Notably, we found that a one-time licensing treatment of MSCs induced higher immunomodulatory potential compared to basal MSCs as observed by sustained immunomodulatory gene expression throughout the first 7 days as well as an increase in immunomodulatory cytokine (PGE₂ and IL-6) expression throughout a 21-day culture period. Further, heparin scaffolds facilitated higher osteogenic cytokine secretion but lower immunomodulatory cytokine secretion compared to chondroitin-6-sulfate scaffolds. Anisotropic scaffolds facilitated higher secretion of both osteogenic protein OPG and immunomodulatory cytokines (PGE₂ and IL-6) compared to isotropic scaffolds. These results highlight the importance of scaffold properties on the sustained kinetics of cell response to an inflammatory stimulus. The development of a biomaterial scaffold capable of interfacing with hMSCs to facilitate both immunomodulatory and osteogenic responses is an essential next step to determining the quality and kinetics of craniofacial bone repair.