Figure 2. Silencing EWS/FLI results in functional metabolic changes.
A & B. Glycolysis stress tests demonstrate an increased extracellular acidification rate (ECAR) in response to a glucose bolus in shEF vs. shLUC A673 cells. No difference was seen in oxygen consumption rate in the glycolysis stress test. C & D. Basal ECAR is higher in shEF vs. shLUC cells, a reflection of different basal glucose concentrations in different test media. Mito stress tests reveal increased oxygen consumption rate (OCR) after treatment with the FCCP mitochondrial uncoupler, indicating increased maximal respiration in shEF vs. shLUC A673 cells. Basal respiration was similar between groups. E. Hexokinase 1 (HK1) was increased in A673 and UTES-14-01872 cells after EF knockdown. F. Uptake of the fluorescent glucose molecule 2-NBDG was increased in shEF vs. shLUC cells, as measured by flow cytometry. G. Mitotracker red signal was elevated in shEF vs. shLUC cells, suggesting increased mitochondrial membrane potential. A representative image from A673 cells is shown, and quantification was done on 3 cells from each of 5 high-power field images, shown as the corrected total cell fluorescence (CTCF) ratio of mitotracker red / mitotracker green (mean ± standard deviation).