Integration of layered chondrocyte-seeded alginate hydrogel scaffolds

Motivated by the necessity to engineer appropriately stratified cartilage, the shear mechanics of layered, bovine chondrocyte-seeded 20 mg/mL alginate scaffolds were investigated and related to the structure and biochemical composition. Chondrocyte-seeded alginate scaffolds were exposed to a calcium-chelating solution, layered, crosslinked in CaCl2, and cultured for 10 weeks. The shear mechanical properties of the layered gels were statistically similar to those of the non-layered controls. Shear modulus of layered gels increased by approximately six-fold while toughness and shear strength increased by more than two-fold during the culture period. Hydroxyproline content in both layered gels and controls had statistically significant increases after 6 weeks. Glycosaminoglycan (GAG) content of controls increased throughout culture while GAG content in layered gels leveled off after 4 weeks. Hematoxylin and eosin histological staining showed tissue growth at the interface over the first 4 weeks. Shear mechanical properties in the engineered tissues showed significant correlations to hydroxyproline content. Dependence of interfacial mechanical properties on hydroxyproline content was most evident for layered gels when compared to controls, especially for toughness and shear strength. Additionally, interfacial properties showed almost no dependence on GAG content. These findings demonstrate the feasibility of creating stratified engineered tissues through layering and that collagen deposition is necessary for interfacial integrity.