Fig. 5

Beige adipocytes released FFAs are essential for CRC proliferation, invasion, and survival. a A flowchart illustrating the two-step CM preparation and treatment of human or mouse parental CRC cell lines. 1: CM preparation from MIIP-decreased or control cells; 2: Treatment of mature adipocytes with the corresponding CM; 3: After exposure to CRC CM for 24 h to induce their browning, the adipocytes were washed, and the medium was refreshed; 4: After another 24 h, the adipocytes CM were collected and applied to fresh cultures of parental CRC cell lines. b Parental HCT116 cells treated with the two-step CM as described in a, and cell viability was measured with CCK-8 (7 biological replicates per group). c Parental HCT116 cells treated with the two-step CM as described in a, and cell viability was measured with CCK-8, 7ACC1 (10 μM), or SSO (20 μM) were added at the indicated time point, respectively (7 biological replicates per group). d–e Parental HCT116 cells treated with the two-step CM as described in a, and the invasive ability was evaluated by trans-well invasion assay (40 × for images with a 50 μm scale bar. 7ACC1: 10 μM; SSO: 20 μM. 6 biological replicates per group). f–g Parental HCT116 cells treated with the two-step CM as described in a, and cell apoptosis induced by oxaliplatin was determined by PI-Annexin V staining f followed by flow cytometric analysis and quantification g, 7ACC1: 5 μM, SSO: 10 μM (5 biological replicates per group). h RT-qPCR-determined expression of genes related to FFAs transport (CD36 and FABP4), FAO, lactate transport (MCT1 and MCT4), and catabolism (LDHA and LDHB) in parental HCT116 cells treated with the two-step CM as described in a (6 biological replicates per group). i Oxygen consumption rates (OCR) were measured by Seahorse XF analysis in parental HCT116 cells at 24 h after exposure to the indicated CM. Arrows indicate the time when Etomoxir (Eto, 50 μM), and rotenone (Ron, 0.2 μM)/antimycin A (AA, 0.2 μM) were added (7 biological replicates per group). j The amount of OCR derived from FAO of HCT116 cells treated with the indicated CM (1 technical replicate of 7 biological replicates per group). k Stable CD36-knockdown or scramble HCT116 cells treated with the two-step CM as described in a, and cell viability was measured with CCK-8 (7 biological replicates per group). Adi or Adipo: mature adipocytes. All data are presented as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001