Fig. 2

Lipid loading augments hypoxia mediated secretion of pro-tumorigenic factors. a, Ultrasensitive proximity extension assay (PEA) profiling of the secreted proteome of GBM cells (U-87 MG) grown in serum free medium for 48 h at normoxic and hypoxic (1% O₂) conditions with no addition of extracellular lipid or in hypoxia with extracellular lipid (LDL, 50 μg/ml) (N = 3 independent experiments). Shown are protein identities increased by at least 1.5-fold (linear) in hypoxia vs normoxia (light gray bars). Dark blue bars show fold increase by lipid loading in hypoxic cells (Hypoxia + LDL vs Hypoxia). ELISA validation of VEGF-A (pg/ml) (b) and HGF (pg/ml) (c) in the secretome of GBM cells grown under the same conditions as in (a) for 6, 24 and 48 h. # = not detected. Western blot analysis of CA9 (d) and vimentin (e) levels in the secretome of GBM cells grown under the same conditions as in (a). Shown are representative blots (left panels) and quantifications (right panels) from at least three independent experiments. HMW: High molecular weight variant of CA9 substituted with glycosaminoglycan