Modulating Gradients in Regulatory Signals within Mesenchymal Stem Cell Seeded Hydrogels: A Novel Strategy to Engineer Zonal Articular Cartilage

Engineering organs and tissues with the spatial composition and organisation of their native equivalents remains a major challenge. One approach to engineer such spatial complexity is to recapitulate the gradients in regulatory signals that during development and maturation are believed to drive spatial changes in stem cell differentiation. Mesenchymal stem cell (MSC) differentiation is known to be influenced by both soluble factors and mechanical cues present in the local microenvironment. The objective of this study was to engineer a cartilaginous tissue with a native zonal composition by modulating both the oxygen tension and mechanical environment thorough the depth of MSC seeded hydrogels. To this end, constructs were radially confined to half their thickness and subjected to dynamic compression (DC). Confinement reduced oxygen levels in the bottom of the construct and with the application of DC, increased strains across the top of the construct. These spatial changes correlated with increased glycosaminoglycan accumulation in the bottom of constructs, increased collagen accumulation in the top of constructs, and a suppression of hypertrophy and calcification throughout the construct. Matrix accumulation increased for higher hydrogel cell seeding densities; with DC further enhancing both glycosaminoglycan accumulation and construct stiffness. The combination of spatial confinement and DC was also found to increase proteoglycan-4 (lubricin) deposition toward the top surface of these tissues. In conclusion, by modulating the environment through the depth of developing constructs, it is possible to suppress MSC endochondral progression and to engineer tissues with zonal gradients mimicking certain aspects of articular cartilage.