Fig. 4

Loss of lipid content in cancer-associated adipocytes. Adipocytes were cocultured with melanoma cells (lines: WM1341D, A375, WM9, and Hs294T) present on Transwell inserts. A After coculture, obtained CAAs, control adipocytes, and 3T3L1 fibroblasts were seeded on coverslips, fixed, and stained with Lipid Spot 488 to visualize lipid droplets. Cell nuclei were stained using Hoechst 33,342 reagent. Scale bar: 25 μm. B To quantify the amount of neutral lipids, cells were fixed and then stained with Oil Red O, and the amount was measured spectrophotometrically. C Expression of PLIN1 (perilipin1) in 3T3L1 cells, control adipocytes, and CAAs established by real-time PCR analysis. For quantification, the samples were normalized against the expression of EEF2. For all experiments, the control constitutes normal adipocytes. In the case of quantitative analyses, the mean of at least three biological repetitions ± SD is shown. Asterisks indicate statistically significant differences between control cells and 3T3L1/CAAs. The significance levels were set at p ≤ 0.01 (**), p ≤ 0.001 (***), or p ≤ 0.0001 (****)