Extended Data Fig. 3: α-Toc is the primary lipid driving lipoprotein-mediated antioxidant protection.
From: Glycosaminoglycan-driven lipoprotein uptake protects tumours from ferroptosis
a. Rank of significantly altered lipid metabolites (detected in negative mode) in B16 cells treated with HDL versus LDL. b. LC/MS analysis of eight vitamin E isoforms in HeLa cells supplemented with human LDL and HDL (50 µg/mL each), shown as fold change relative to lipoprotein-depleted cells. c. LC/MS analysis of eight vitamin E isoforms in HeLa cells treated with LDL or HDL (50 µg/mL), shown as fold change relative to lipoprotein-depleted cells. d. Relative metabolite peak area for eight vitamin E isoforms in plasma from mice fed a control or Vitamin E-deficient diet. e. Proliferation (log2 doublings, 5 days) of Karpas299 in vitro untreated or treated with ML162 (125 nM) ± 50 ug/mL of human HDL, HDL isolated from mice fed a vitamin E-deficient diet, HDL isolated from mice fed a vitamin E-sufficient diet, or Fer-1 (1 μM). f. Representative tumours of Karpas299, HeLa, and B16 cells implanted subcutaneously in mice fed a control or vitamin E-deficient diet. g. Quantification of circulating plasma triglycerides (TGs, mg/dL) and HDL (mg/dL) in mice fed a control or a vitamin E-deficient diet. h. Schematic depicting generation of Ttpa-liver specific knockout mice to evaluate depletion of α-toc in driving tumour growth in mice. i. Immunoblot of TTPA in the liver of mice injected with liver-specific AAV8-sgControl or AAV8-sgTtpa. Ponceau staining is used as loading control. j. Plasma α-toc levels from Cas9 knock-in mice injected with liver-specific AAV8-sgControl or AAV8-sgTtpa 4 weeks after AAV injection. k. Quantification of 4-HNE H-scores in B16 tumours grown in control or liver-specific Ttpa knockout mice. l. Tumour weights from control and liver-specific Ttpa knockout mice implanted with B16 cells. m. Mass spectrometry analysis of vitamin E isoforms α-tocotrienol and δ-tocotrienol in plasma from mice fed a standard mouse diet or a Vitamin E-sufficient diet. Data is presented as relative metabolite peak area. b-e, g, k, m: Bars represent mean ± s.d.; j, l: Boxes represent median, first/third quartiles; whiskers are range. b, c, e, n = 3 biological replicates; d: n = 4 biological replicates; g: n = 7 biological replicates; j: n = 5 biological replicates; k: n = 14 representative fields; l: n = 10-12 biological replicates; m: n = 3-4 biological replicates. Statistics by: i) two-sided unpaired t-tests as indicated or compared to untreated cells (b, c), control-diet mice (d, g), or to ML162-treated cells (e); or ii) by one-way ANOVA followed by a Kruskal–Wallis nonparametric test (a). For gel source data, see Supplementary Fig. 1.
