Fig. 3: Native and infectious CJD prions drive ferroptosis sensitivity.

A FABP5 changes in PrP^C OE and empty vector control cells by immunostaining and Western with densitometry. Pictures shown are representative results of three independent repetitions performed in triplicate with similar outcomes (scale bar = 20 µm). B Relative expression of ferroptosis markers in PrP^C OE (OE) and control (C) cells detected by qPCR and Western. Lower blots display total protein loading. C Comparison of FABP5 staining intensity in iPS-derived brain organoids infected with normal brain homogenates (NBH) and sCJD prion homogenates (169 days post-infection, dpi). Pictures shown are representative results of at least three independent repetitions performed with similar outcomes of minimum 10 organoids (scale bar = 200 µm (left panels) and 50 µm (right panels)). D Western blotting and densitometry comparing FABP5 protein levels in the brain tissue of mice inoculated with normal brain homogenate (NBH; control) or RML scrapie brain homogenate and sacrificed at 80, 108, and 160 days post inoculation (dpi). E Western blotting and densitometry comparing FABP5 protein levels in the brain tissue of people who died from sporadic CJD with tissue from patients who died of a non-brain-related condition. F Lactate dehydrogenase (LDH) release level of brain organoids treated with increasing RSL3 concentrations and aToc rescue for ferroptosis specificity. G Normalized survival of brain organoids infected with sCJD (MM1) prion homogenates for 90 days compared to NBH controls against an RSL3 treatment at indicated concentrations (scale bar = 200 µm). Cell death was measured by LDH release activity (48 h, relative to DMSO). H Normalized survival of PRNP KO compared to NBH controls treated with RSL3 and measured by LDH release (48 h, relative to DMSO). Western data (A, B) is shown as mean ± SD of n = 3 technical replicates. Relative mRNA expression (B) is shown as mean ± SEM of n = 3 technical replicates. LDH release data and Western data (E) are plotted as representative mean ± SEM of n  =  3 (D, F) or n = 6 (E, G, H) biological replicates for independent experiments. Significance was determined by two-tailed t-test (A, B, F), two-tailed Welch’s t-test (E) and two-way ANOVA comparisons, Tukey post-test (D, G, H). *P  <  0.05, **P  <  0.01, ***P  <  0.001, ****P  <  0.0001. Source data are provided as a Source Data file.