Development of a cell-seeded construct for osteochondral modelling and repair

Regenerative therapeutic solutions are required to address the increasing prevalence of bone and cartilage diseases within the population. Limitations of existing treatments, such as bone graft reconstructions or biomaterial implants, suggest that osteochondral tissue constructs with the ability to support differentiation of mesenchymal stem cells into both osteoblast and chondrocyte lineages is desirable. To date, tissue-engineering approaches have focused on developing individual scaffolds for each osteochondral lineage. Constructs are combined once tissues have developed sufficiently. Unfortunately, delimitation often occurs under normal physiological loading and implants fail. The overall aim of this research project was to develop a scaffold made from a single material with the capacity to maintain osteogenic and chondrogenic cells. In this manner, it was intended to overcome issues arising from delamination and the divergent differentiation requirements for each lineage by providing scaffolds with spatially resolved environments, each supportive of one of osteochondral cell lineages. The work reported here describes a novel method to produce porous chitosan scaffolds with large pore regions (300-425 μm) to promote the osteogenic differentiation of mesenchymal stem cells, and smaller pores (180-300 μm) to encourage chondrogenesis. Porogen properties and cross-linker optimisation were fundamental for the production of a bi-layered chitosan scaffolds containing two distinct pore sizes, successfully achieved in the current project. The architecture of the chitosan scaffolds also permitted the development of a culture medium that could activate simultaneous osteogenic and chondrogenic differentiation in mesenchymal stem cells. More specifically, it was determined that 5-day transient serum treatments with fetal calf serum or human serum, allowed bone and cartilage development. Finally, a perfusion bioreactor system was used to confirm the biocompatibility and osteochondral differentiation potential of the bi-layered chitosan scaffolds