Figure 3

Semi-quantitative real time PCR showed IGF-1 and latent TGF-β1 resulted in changes in marker gene expression consistent with EMT in MCF-7 cells. MCF-7 cells treated with (i) IGF-1, (ii) IGF-1+latent TGF-β1, (iii) IGF-1+latent TGF-β1+PI3K inhibitor, (iv) IGF-1+latent TGF-β1+MAPK inhibitor, (v) IGF-1atent TGF-β1+TGF-β1 inhibitor or (vi) IGF-1+latent TGF-β1+MMP inhibitor were analyzed for levels of marker genes associated with EMT. E-cadherin, and occludin levels decreased by 21 and 6 fold respectively while N-cadherin and vimentin levels increased by 24 and 8 fold respectively when treated with IGF1-+ latent TGF-β1 as compared to latent TGF-β1 treatment alone. As there was no significant difference between vehicle and latent TGF-β1 treatment, latent TGF-β1 treatment alone was used for comparison with other treatments to further highlight changes associates with the activation of TGF-β1 and EMT. The addition of PI3K, MAPK, MMP or TGF-β1 inhibitors attenuated these changes in marker gene expression. With the exception of vimentin, IGF-1 treatment alone caused changes in marker gene expression similar to that seen with IGF-1+ TGF-β1+inhibitors. All data are mean ± s.e.m. (n = 3, three experimental repeats).