Journal: Cell Death Discovery

Article Title: Rottlerin triggers dual degradation of SLC7A11 and GPX4 to drive ferroptosis and chemosensitization in hepatocellular carcinoma

doi: 10.1038/s41420-026-02942-1

Figure Lengend Snippet: A HLE and LM3 cells were treated with 2 μM rottlerin or in combination with 2 μM Fer-1. The cells were stained with crystal violet and then dissolved with 10% glacial acetic acid. The absorbance at 595 nm was measured and calculated. Data were represented as mean ± SD ( n = 3) and analyzed using Two-way ANOVA, *** P < 0.001. B LM3 cells were treated with 0.5 μM rottlerin with or without 2 μM Fer-1 for 14 days and then treated with crystal violet stain and photographed. Scale bar = 1 cm. C HLE and LM3 cells were treated with 5 μM rottlerin with or without 2 μM Fer-1 for 72 h and then imaged. Scale bar = 100 μm. D HLE and LM3 cells were exposed to the indicated doses of rottlerin with or without 2 μM Fer-1 for 48 h. Cell viability was assessed using the CCK-8 assay. Data were represented as mean ± SD ( n = 3) and analyzed using two-way ANOVA, *** P < 0.001. E HLE and LM3 cells were treated with 5 μM rottlerin with or without 2 μM Fer-1 for 72 h, then stained with PI for flow cytometric analysis. The representative images (top) and cell death rates (bottom) were displayed. Data were represented as mean ± SD ( n = 3) and analyzed using Student’s t -test; ns means not significant, *** P < 0.001. F HLE and LM3 cells were treated with 5 μM rottlerin for 60 h, then stained with the lipid ROS probe BODIPY 581/591 C11 for flow cytometric analysis. The representative images (top) and relative lipid ROS level (bottom) were displayed. Data were represented as mean ± SD ( n = 3) and analyzed using Student’s t -test, *** P < 0.001. G The morphological changes of mitochondria were detected by transmission electron microscopy (TEM) in HLE cells incubated with or without 5 μM rottlerin for 48 h. Scale bar, 500 nm. H Measurement of malondialdehyde (MDA) levels in HLE cells treated with 5 μM rottlerin for 48 h. Data were represented as mean ± SD ( n = 4) and analyzed using Student’s t -test, * P < 0.05. I Measurement of GSH levels in HLE cells treated with 5 μM rottlerin for 48 h. Data were represented as mean ± SD ( n = 3) and analyzed using Student’s t -test, *** P < 0.001.

Article Snippet: Following incubation in the dark for the specified duration, the medium was replaced with PBS containing the lipid ROS probe BODIPY 581/591 C11 (MCE, Cat# HY-D1691, China) or Fe 2+ probe RhoNox-1 (TargetMol, Cat# T87330 , China), and the cells were incubated at 37 °C for 30 min.

Techniques: Staining, CCK-8 Assay, Transmission Assay, Electron Microscopy, Incubation