Figure 1. MSF over expression confers the cancer-associated fibroblast phenotype, characterized by the expression of myo fibroblast markers and activated TGF-β signaling. (A) To assess the functional role of MSF stromal expression in tumor growth, we stably overexpressed the MSF protein in immortalized human fibroblast cell line (hTERT-BJ1 cells). Successful overexpression of the MSF protein was verified by immunoblot analysis. Empty vector control (Lv-105) or MSF fibroblasts were also subjected to immunoblot analysis with different myofibroblast marker proteins. Equal protein loading was assessed by immunoblotting with β-actin. Note that MSF overexpression strongly induces the expression of myofibroblast markers such as α-SMA, calponin 1 and 3 and fibronectin, while no increases in vimentin protein expression were observed in the fibroblasts. (B) MSF overexpression induces the activation of TGF-β signaling. Note that immunoblot analysis of control and MSF fibroblasts reveals that TGF-β ligand protein expression is significantly increased in MSF fibroblasts. Conversely, MSF fibroblasts are characterized by a reduction in the protein expression of TGFβ-RI (type I receptor), as compared with control fibroblasts.