Do factors secreted from synovial fibroblasts affect the differentiation of C2C12 cells?
McCabe, Emma Louise (2011)
M.Res. thesis, University of Birmingham.
The Wnt signalling pathway plays a key role within muscle differentiation. Wnt3a, Wnt5a, and DKK1 all have pivotal roles within this pathway. It’s hypothesised that due to their role within the Wnt pathway, Wnt3a, Wnt5a, and DKK1 will affect the differentiation of muscle, demonstrated by the murine C2C12 cell line.
Differentiation of the C2C12 cells was induced by adding DMEM differentiation media at day 0. Treatments (Wnt3a, Wnt5a, DKK1, control conditioned media, TNF-α conditioned media, and dexamethasone conditioned media) were added day 0, and mRNA levels of differentiation markers, MyoD, myogenin, α-actinin, and 11βHSD1 were measured using RT-PCR at days 1, 3 and 6. Wnt3a and control conditioned media gave no significant change in differentiation. Wnt5a, DKK1, TNF-α conditioned media and dexamethasone conditioned media gave significant decreases in differentiation. DKK1 inhibitor was tested on cells treated with TNF-α conditioned media, resulting in the decrease in the differentiation no longer being significant. 11βHSD1 enzyme activity assays were carried out to test Wnt3a, Wnt5a, DKK1, and DKK1 inhibitor effects, the results followed the trend of the mRNA data, however were not statistically significant. The results suggest that factors secreted from synovial fibroblasts during inflammation affect muscle differentiation.
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