Modulation of connective tissue stromal cell plasticity to generate cartilaginous phenotypes

Stem cells are unspecialised cells found in the body which possess the ability to self renew and can be induced to proliferate and differentiate into more specialised cells. MSCs are adult stem cells that are capable of leaving the bone marrow and travelling in the bloodstream to a different site, where they may perform repair or regeneration processes of various mesenchymal tissues such as cartilage, bone and fat. Due to these properties of MSCs, it proves to be a useful source for the repair and regeneration of cartilage. The aims of this study were to: 1) characterise primary bovine MSCs by assessing the gene expression of hyaline cartilage-specific genes (SOX-9, Aggrecan and Collagen type II) and non-specific genes (Collagen type I and type X) 2) investigate the effects of culture medium supplemented with FGF-2 or with the addition of an extra growth factor, TGF-p2, on bovine MSCs in a two dimensional culture system 3) characterise the phenotypes of MSCs tissue grafts produced using MSCs pre-cultured in culture medium supplemented with FGF-2 or with the addition of TGF- p2 in a three dimension culture system (Transwell) seeded at high (6xl06 cells) and low (0.5 × 106 cells) cell density. Our study showed that different bone chamber size and thickness influenced the amount of marrow and cells harvested without interfering with the fibroblastic-shaped cell morphology and their adherence ability. The presence of stem/progenitor cell features is present in undifferentiated P0 BMSCs and PI and P2 BMSCs cultured in FGF-2 or with the addition of TGF-p2. Gene expression analyses on undifferentiated P0 BMSCs and PI and P2 BMSCs cultured in FGF-2 or with the addition of TGF-p2 suggested that the MSCs contain both fibroblastic and chondrogenic features. This implies that the MSCs are not fully committed towards a chondrogenic lineage. We have demonstrated that it is possible to generate a tissue graft using passaged BMSCs in a Transwell culture system but seeding density is important. The ideal seeding cell density is 0.5x106 cells/well using P2 and P3 BMSCs pre-cultured in either FGF-2 or with the addition of TGF-P2. The tissue graft produced has a high expression level of aggrecan, collagens type I and II, which suggested that it has a fibro-cartilage phenotype