Figure 4: Fat tissue filtrate from mice fed a high fat diet stimulates the formation of a fibroblast-like morphology of JB6 cells growing in liquid medium. A. JB6 cells were seeded at 100 cells per well in a 6-well plate. Cells were treated with 5% FBS or fat tissue filtrate from mice fed a HFD for 7 days and stained with Giemsa. The picture shows a comparison of epithelial-like morphology and fibroblast-like morphology of JB6 cells (magnification = 400×). The fibroblast-like clones pictured were characterized as being either early- (A2), middle- (A3), or late-stage (A3) depending upon the size of the foci formed. B. JB6 cells were treated for 7 days with 5% FBS or fat tissue filtrate (50 μg protein/mL) from mice fed a HFD. E-cadherin, Twist and β-Actin levels were determined in the epithelial-like and fibroblast-like clones by western blotting. C. JB6 cells were seeded and treated as described in (A). JB6 cells treated with fat tissue filtrate (50 or 100 μg protein/mL) from mice fed a HFD had a greater ratio of clones with fibroblast-like to epithelial-like cells when compared with cells cultured with fat tissue filtrate from mice fed a LFD (*P<0.05). Arrows indicate % increase in the ratio of fibroblast-like to epithelial-like clones at the different doses. One well = 9.6 cm2. All data are mean ± SEM, Student t test (unpaired, two-tailed).