Regulation of chondroprogenitor cell gene expression and migration
Cartilage, which lines joint surfaces to allow near-frictionless movement, lacks the ability to adequately repair itself and there are currently no effective, disease-modifying drugs to halt or repair the damage. Development of powerful in vitro models to investigate gene expression changes during osteoarthritis and chondrogenesis is key in understanding how the disease develops and how cartilage might attempt to repair itself. In this thesis, an enhanced model of chondrogenesis of the murine ATDC5 chondroprogenitor cell line was developed with cells cultured in micromass. Results revealed not only an increase in chondrogenesis markers, but markers of growth plate differentiation, including type X collagen, were either restricted or repressed, whilst expression of genes rich in articular cartilage were upregulated. This suggests that the enhanced ATDC5 model is more reminiscent of articular cartilage, making this model suitable for investigations into osteoarthritis - a disease of articular cartilage. The role of WNT5A signalling was then investigated (which is up-regulated in osteoarthritic cartilage) in a disease-like context by stimulating cultures with cytokines. Microarray analysis unearthed interesting and novel results, including a decrease in WNT5A signalling and in expression of members of the CCN family. Pathway analysis allowed further exploration of the interrelationship between cytokine and WNT signalling. Some changes in gene expression were reminiscent of those observed previously in in vivo models of early osteoarthritis. Finally, migration studies revealed that non-induced, undifferentiated ATDC5 cells have a migratory phenotype reminiscent of chondroprogenitor cells, which have the capacity to migrate to sites of cartilage injury in vivo. A novel model of cartilage invasion was also developed, with results suggesting WNT5A may be a potential inducer of chondroprogenitor invasion. Together, this thesis shows that the ATDC5 model is a good model for investigating articular cartilage both in a physiological and pathological setting.
Networked Digital Library of Theses and Dissertations