Extended Data Figure 6: CD36⁺ cells are stimulated by a high-fat diet or adipocyte-conditioned medium, and require the ability of CD36 to internalize fatty acids for their pro-metastatic potential.

a, Flow cytometry analysis of orthotopic transplants of Detroit-562 cells transduced with PLKO or shRNACD36#98 or #99, from mice fed with high-fat diet (HFD) or control diet (CD), analysed 4 weeks after OSCC injection. Numbers indicate CD44^bright CD36⁺, CD44^bright CD36^– and CD44^dim (differentiated) cells in the represented gate, expressed as percentages from the total GFP⁺ Lin^– OSCC parental tumour. n = 5 animals per group. b, Flow cytometry analysis of co-cultured SCC-25/OP-9, SCC-25/adipogenic OP9 or SCC-25/HNCAFS (head and neck cancer–associated fibroblasts) cells. Numbers indicate CD36⁺ cells in the represented gate, expressed as percentage. c, FACS analysis of co-cultured Detroit-562 or SCC-25 with OP9 (control) or adipogenic OP9, showing an increase in the percentage of CD36-positive cells in the adipogenic co-cultures. Numbers indicate CD44^bright CD36⁺ and CD44^bright CD36^– from the total GFP⁺CD29⁻ OSCC cells. d, CD36 mRNA relative expression levels, measured by RT–qPCR, from SCC-25 CD36^– sorted cells either co-cultured with adipogenic OP9 (Ad.OP9) cells or not, or from SCC-25 CD36⁺ sorted cells co-cultured with Ad.OP9 cells. In b–d, OSCC were co-cultured in vitro for 2 days. e, Flow cytometry analysis of OSCC cells co-cultured with adipogenic OP-9 cells or with 0.4 mM palmitic acid (PA). Histograms show the average normalized number of events as a function of CD36 and CD44 fluorescence intensity. f, cDNA and amino acid sequence of the CD36 receptor at the level of the point mutation introduced to generate the fatty acid-binding site mutant, CD36-K164A (left). Fatty acid uptake assay is shown for SCC-25 cells not transduced (as control, CT) or transduced with CD36wt (overexpressing wild-type CD36), shRNA Cd36 or CD36-K164A. g, BLI monitoring of transplants from SCC-25 cells overexpressing CD36wt (wild-type, n = 10) or CD36-K164A (n = 10). Frequency of developed tumours is expressed as percentage (*P = 0.02, Fisher exact test), and BLI signal quantification is expressed as the relative normalized photon flux (* P= 0.05, two-tailed t-test). Data are given as the mean ± s.e.m. h, FACS analysis of OSCC cells overexpressing either CD36 wild-type (wt) or mutant (Lys164mut). Histograms show the average normalized number of events as a function of CD36 and CD44 fluorescence intensity. Source data from mouse experiments are in Supplementary Information.

Source data