Fig. 5: β-hydroxybutyrate induces ferroptosis of pancreatic cancer cells by downregulating CAV1.

A Western blotting of CAV1 and α-Tubulin after treatment by a gradient concentration (0, 1, 2, and 5 mM) of Na-OHB. B qPCR of CAV1 after treatment by a gradient concentration (0, 1, 2, and 5 mM) of Na-OHB. C MIA PaCa-2 cells were treated with a gradient concentration of Na-OHB, and dead cells were stained with SYTOX Orange. The bar graph (left) shows the cell death rate of MIA PaCa-2 cells following Na-OHB treatment, while the bar graph (right) presents the cell death rate of PANC1 cells after Na-OHB treatment. D Apoptosis and dead cells were detected using flow cytometry. E Cells were treated with Na-OHB with or without Fer-1, and dead cells were stained with SYTOX Orange. F Lipid ROS levels within cells were measured by flow cytometry. G GSH/GSSG ratio of MIA PaCa-2 and PANC1 cells after being treated with Na-OHB/Fer-1. H Intracellular Fe²⁺ levels were assessed using flow cytometry. I Intracellular cystine was assessed using flow cytometry. J CAV1 overexpression cells were treated with 0 mM or 2 mM Na-OHB, and dead cells were stained by SYTOX Orange. K CAV1 overexpression cells were treated with 0 or 2 mM Na-OHB, and lipid ROS levels within cells were measured by flow cytometry. L CAV1 overexpression cells were treated with 0 or 2 mM Na-OHB, and intracellular Fe²⁺ levels were assessed using flow cytometry. A Data were shown as the mean ± SD, n = 3, two-tailed t-test or one-way ANOVA with Dunnett t-test. B, C, E–L Data were shown as the mean ± SEM, n = 3, two-tailed t-test or one-way ANOVA with Dunnett t-test. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001.