The effect of cyclic tensile loading and growth factors on the chondrogenic differentiation of bone-marrow derived mesenchymal stem cells in a collagen-glycosaminoglycan scaffold.

The aim of this thesis was to demonstrate that the application of cyclic tensile strain to MSC-seeded collagen-GAG scaffolds regulates the chondrogenic differentiation process, in the presence of chondrogenic growth factors. A custom-designed multistation bioreactor was developed to apply cyclic uniaxial tensile strain to 3D constructs.  Firstly, the chondrogenic differentiation of the MSCs using TGF-β1 was demonstrated.  To apply cyclic tensile strain, the cell-seeded scaffolds were uniaxially clamped between stainless steel grips.  This allowed for an investigation into the importance of cell-mediated contraction for the differentiation process.  Inhibition of contraction through clamping resulted in a reduction in the rate of GAG synthesis.  The application of physiological loading of 10% strain at 1 Hz reversed this process and resulted in an increase in the rate of GAG synthesis. A computational model was developed to investigate the biophysical stimuli developed within the scaffold during cyclic loading.  The results demonstrated that the magnitudes of fluid flow and strain developed were similar to those previously reported to regulate chondrogenic differentiation, therefore indicating that these stimuli were the main regulators in the mechanoregulation of the differentiation process. Characterisation of intracellular signalling pathways involved in TGF-β1 and IGF-1-induced chondrogenic differentiation demonstrated the differential involvement of the MAPK and PI3-kinase in involved in the chondrogenic differentiation of adult MSCs, in 2D and in the 3D collagen-GAG scaffold.  These intracellular studies provide further knowledge of possible downstream pathways of mechanosensitive membrane receptors involved in the mechanotransduction and mechanoregulation of MSC differentiation.