Articular cartilage lesions cause pain, morbidity and may progress to osteoarthritis, which in terms of medical care, has been approximated to cost 1-2.5% of the gross national product of USA, UK, France and other countries. Current repair strategies have considerable limitations and have prompted the development of cartilage tissue engineering (CTE) approaches. One of the challenges of CTE is the expansion and differentiation of adult stem cells in vitro, into functional articular chondrocytes whilst avoiding hypertrophy. Hypoxia is an important environmental factor required for cartilage development, for stimulating articular chondrogenesis and ECM formation. There remains, however, a number of questions regarding the in vitro role of hypoxia and HIF stabilization in the development of CTE strategies. These include: a) the appropriate level of hypoxia required for chondrogenesis, b) determining if artificial HIF stimulation has advantages over physiological hypoxia, c) which of the commonly-used HIF-stimulating compounds most potently induces HIF-mediated articular chondrogenesis and d) if there exists a relationship between mechanostransduction and HIF during chondrogenesis. Compared to normoxia, hypoxia (2%O2 and 5%O2) induced the expression HIF target genes (including VEGFA, PGK1 and EGLN) but only 5% inhibited hypertrophic collagen type X expression. Artificial stimulation of HIF-1a by DMOG induced greater expression of HIF chondrogenic targets (SOX9 and collagen-modifying enzymes) than other compounds used (CoCl2 and DFX) and physiological hypoxia. DMOG also reduced collagen type X at the mRNA level compared to the other HIF stabilising compounds. In terms of the effect of hypoxia on mechano-signalling during chondrogenesis, exposure to 2%O2 induced ROCK activity, actin re-organisation and SOX9 expression during BM-MSC chondrogenesis on soft polyacrylamide gels. No such changes were induced on stiff substrates. This suggests the existence of specific crosstalk between HIF and stiffness-sensing pathways, which may inform CTE strategies in which hypoxia-mediated chondrogenesis of BM-MSCs is conducted within biomaterial scaffolds of a defined stiffness.
|Date of Award||2018|
|Supervisor||Eileen Gentleman (Supervisor)|