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

Abstract

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.

Sustained Release of Dexamethasone from 3D-Printed Scaffolds Modulates Macrophage Activation and Enhances Osteogenic Differentiation

“…For example, MSCs have been shown to switch activated M1 macrophages to an M2 phenotype via prostaglandin E2. , MSC-borne chemokines, CCL2 in particular, have been found to modulate the phenotype of macrophages . Factors secreted by pro-inflammatory and anti-inflammatory macrophages activated the immunomodulatory potential of MSCs. , Inflammation-primed hMSCs exhibited higher immunomodulatory capacity compared to nonprimed cells. , All microphage phenotypes (M0,M1,M2) were demonstrated to be potent chemotactic stimulators for MSCs . Immuno-regulatory effects of placental MSCs have been found to be mediated by soluble molecules, such as IL10 and TGF-β1, acting partially via glucocorticoid receptor and progesterone receptor .…”

“… 62 Factors secreted by pro-inflammatory and anti-inflammatory macrophages activated the immunomodulatory potential of MSCs. 63 , 64 Inflammation-primed hMSCs exhibited higher immunomodulatory capacity compared to nonprimed cells. 63 , 64 All microphage phenotypes (M0,M1,M2) were demonstrated to be potent chemotactic stimulators for MSCs.…”

“… 63 , 64 Inflammation-primed hMSCs exhibited higher immunomodulatory capacity compared to nonprimed cells. 63 , 64 All microphage phenotypes (M0,M1,M2) were demonstrated to be potent chemotactic stimulators for MSCs. 65 Immuno-regulatory effects of placental MSCs have been found to be mediated by soluble molecules, such as IL10 and TGF-β1, acting partially via glucocorticoid receptor and progesterone receptor.…”

Donor Variability in Human Mesenchymal Stem Cell Osteogenic Response as a Function of Passage Conditions and Donor Sex

“…Dry sterile scaffolds were hydrated and crosslinked prior to cell seeding as previously described [37,56,59] and all steps were done under moderate shaking. Scaffolds were first hydrated for 2 hours in ethanol at ~20°C (room temperature).…”

“…Our collaborators have shown that MSCs cultured on the mineralized collagen scaffolds promote the secretion of osteoprotegerin, which subsequently reduces osteoclast resorption in vitro [49][50][51]. Moreover, our group has used these materials to understand the antimicrobial effects of manuka honey [55], the immunomodulatory effects of placenta-derived tissues [40,44], and the crosstalk between immune cells and hMSCs [56]. Additionally, the material properties of these scaffolds are well studied and characterized in terms of stiffness (~1000 kPa dry [38,39], ~30kPa, hydrated [45]), pore size (100-200 um [39,40,42]), and pore alignment (isotropic, anisotropic [42]).…”