Tuneable collagen I and poly(N-isopropylacrylamide) based hydrogels for tissue engineering applications of mesenchymal stem cells

Due to their multipotency, mesenchymal stem cells (MSCs) are used to model and regenerate many connective tissues. Their behaviour varies with the environment they are maintained within, therefore different tissue scaffolds are required for each application to suitably mimic the tissue being engineered. Although hydrogels used as cell scaffolds are able to mimic certain tissue properties, this is often at the expense of others. Collagen for example is cell adhesive, but lacks the mechanical properties of native tissues. One method to address this challenge is to create hybrid scaffolds, combining materials that separately mimic one, or more of the desired tissue properties, resulting in an optimal scaffold. Furthermore by adjusting the composition of the components within such a gel, a panel of scaffolds can be designed with variable properties for multiple MSC applications. Both collagen and poly(NIPAM-co-styrene-graft-NVP) (NSN) hydrogels undergo gelation at physiological temperatures, enabling cell encapsulation, but individually are less than ideal as cell scaffolds. In this thesis it was demonstrated that combining these two materials produced hydrogels with the biocompatibility and cell adhesiveness of collagen and the mechanical strength of NSN. Through varying the composition of these bio-synthetic hydrogels, a repertoire of materials was produced with a range of mechanical properties and porosities. Hydrogels with low concentrations of NSN, contained fibrillar collagen networks, with no evidence of syneresis whilst gels with higher concentrations of NSN had superior mechanical properties, but were prone to syneresis. Two applications of MSCs were examined using the collagen-NSN gels, the development of an artificial lymph node model and injectable scaffolds for cartilage repair. The backbone of the lymph node consists of a heterogeneous network of stromal cells, which are mesenchymal. Through treating MSCs with the factors involved in lymph node organogenesis, they were induced into a lymphoid mesenchyme phenotype. These were seeded hybrid gels with low concentrations of NSN to develop 3D networks. Increasing the NSN concentration perturbed the morphology and migration of the MSCs within the hydrogels, and produced a scaffold with mechanical properties that mimic those of the lymph node. The tougher hydrogels were also able to support MSCs in 3D. Although the mechanical properties of these gels approached native cartilage, the chondrogenic capacity of the encapsulated cells was reduced compared with MSCs cultured in cell-only 3D pellets. The gels require further optimising for use as scaffolds for cartilage repair. The novel collagen-NSN hydrogels presented in this thesis, have potential as scaffolds for numerous applications as the exact composition of the gels can be tuned to alter their physical properties and therefore direct cell fate.