Characterization of Shear Stiffness and Cell Metabolism in Chondrocyte-Seeded Self-Assembling Peptide Hydrogel Scaffolds

Self-assembling KLD12 peptide hydrogels have previously been established as suitable scaffolds for chondrocytes for the purpose of generating a three-dimensional, cartilaginous neo-tissue, which could potentially be implemented in the repair of in vivo chondral defects. Such tissue constructs have been shown to metabolically respond to dynamic compression by an increase in proteoglycan synthesis and glycosaminoglycan accumulation. For this thesis, modified chondrocyte-seeded, KLD-12 hydrogel constructs have been developed and evaluated for the characterization of age-dependent shear mechanical properties and metabolic response to shear loading. Based on previous research, a new casting frame and technique were developed to create 2 mm thick, KLD12 hydrogels for chondrocyte encapsulation. One day after casting, the total population in the hydrogel was 75-85% live, similar to the viability previously reported in 1.6 mm gels. The equilibrium and dynamic shear stiffnesses of the constructs were determined over the course of 5-35 days post-casting. Methods for the application of pure shear deformation and the measurement of load response were similar to those previously used for cartilage explants. By Day 35, the average measured equilibrium and dynamic moduli were 8.0 ± 1 kPa and 67.7 ± 21 kPa, respectively. Thus, the construct stiffnesses were -3 orders of magnitude lower than explants in shear and -1-2 orders of magnitude lower than 1.6 mm constructs in confined compression. A polysulfone (PSF) shear loading chamber was designed and fabricated to test the effect of dynamic shear deformation on metabolism of chondrocytes in 2 mm KLD-12 hydrogels. In contrast to the stimulatory result in cartilage explants, cyclic shear deformation caused a dramatic decrease in protein and proteoglycan incorporation in the KLD-12 constructs, compared to free-swell controls in culture plates. Notable increase in GAG loss in both loaded and unloaded chamber specimens suggested that chamber artifacts might have been responsible for these metabolic effects.