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. 2021 Mar 17;28(8):2536–2551. doi: 10.1038/s41418-021-00769-0

Fig. 2. FAR1 is critical for long-chain saturated fatty acid-induced ferroptosis.

Fig. 2

a Western blot analysis of HT1080 cells expressing shRNA-ctrl or shRNA-FAR1. The experiments were repeated twice, independently, with similar results. b Dose-dependent toxicity of RSL-3 induced cell death of HT1080 and 786-O cells expressing shRNA-ctrl or shRNA-FAR1. Cell viability was assessed 24 h thereafter using CCK8. c Western blot analysis of HT1080 cells expressing sg-ctrl or sg-FAR1. The experiments were repeated twice, independently, with similar results. d Cell death measurement of HT1080 cells expressing sg-ctrl or sg-FAR1 treated with erastin (10uM) and Fer-1 (1 μM) for 24 h. e Cell death measurement of HT1080 cells expressing sg-ctrl or sg-FAR1 treated with RSL3 (500 nM) and Fer-1 (1 μM) for 12 h. f Representative phase-contrast images of HT1080 cells expressing sg-ctrl or sg-FAR1 treated with RSL3 (500 nM). Dead cells were stained with Sytox Green. Scale bars, 100 µm.The experiments were repeated twice, independently, with similar results. g Lipid peroxidation measurements in HT1080 cells expressing sg-ctrl or sg-FAR1 treated with erastin (10 μM). h Western blot analysis of HT1080 cells expressing sg-ctrl or sg-FAR1 transfected with FAR1 plasmids. The experiments were repeated twice, independently, with similar results. i Cell death measurements of HT1080 cells expressing sg-FAR1 transfected with FAR1. j Western blot analysis of HT1080 GPX4−/− cells expressing sg-ctrl or sg-FAR1. The experiments were repeated twice, independently, with similar results. k Cell death measurements of HT1080 GPX4−/− cells expressing sg-ctrl or sg-FAR1 upon Fer-1 withdraw. l Cell death measurements of WT and FAR1 KO HT1080 cells were pre-incubated with PA (10 μM), SA (10 μM), 1-HE (10 μM) and 1-OE (10uM) for 12 h, and then treated with RSL3(100 nM). PA, palmitic acid. 1-HE, 1-hexadenocal. SA, stearic acid. 1-OE, 1-Octadecanol. Data and error bars are mean ± s.d., n = 3 (b, d, e, g, i, k–l) independent repeats. All P values were calculated using two-tailed unpaired Student’s t-test.