Additional data for "Selenium drives a transcriptional adaptive program to block ferroptosis and treat stroke" by Alim et al.
Paper Title: Selenium drives a transcriptional adaptive program to block ferroptosis and treat stroke These are additional data sets from the related article. 1) Scatter plots for all behavioral data from the paper. GPX4 overexpression scatter plot: (A-B) Forced expression of GPX4 in vivo improves outcome following ICH. Scatterplot of behavioral outcomes in spatial neglect task (n=16; scatter plot) and sensory neglect task (n=16; scatter plot) for up to 14 days post ICH in mice overexpressing GFP or GPX4 in striatum. Selenium treatment scatter plots: (A-B) Scatterplot of behavioral outcomes including spatial neglect task (n=16) and sensory neglect task (n=16) up to 7 days post ICH with or without ICV injection of Se (2.5µM). (C-D) Tat SelPep improves outcomes when given systemically 2h after ICH. Scatterplot of behavioral outcomes in sensory neglect task (n=16) and spatial neglect task (n=16) for up to 14 days post ICH. (E-F) Tat SelPep improves behavioral outcomes with a therapeutic window up to 6 hours. Scatterplot of behavioral outcomes in spatial neglect task ( n=16) and sensory neglect task (n=16) for up to 14 days post ICH in mice given 6 hour post injury IP injections and every day for 7 days of saline, Tat or Tat SelPep (12µg/g). 2) Blots have not been modified with the exceptions of the labels. The file names on the blots correspond to the blots in the figures from the paper. 3) Molecular depletion and validation of exon 5-6 (A-C) and exon 4 (D-F) targeted siRNA of GPX4. Raw blot images are included. 4) (A-D) Differential Interference Contrast (DIC) time-lapse video examples of primary cortical neurons treated with (A) vehicle control, (B) Erastin (5µM), (C) Hemin (80µM), and (D) Hemin+Se (1µM Se). Yellow time bar shows relative hours passed from time of cotreatment, red arrows indicate primary cortical neurons destined to die. Scale bar 100µm. 5) (A-E) ICH modeling in vitro or in vivo shows TUNEL labeling, one feature of ferroptosis. (A-C) HT22 hippocampal neuroblast cells exposed to (A) 100µM Hemin followed by TUNEL labeling, (B)Vehicle control followed by TUNEL labeling, and (C) Hemin without TdT nucleotide treatment followed by TUNEL labeling. White boxes show magnified image and arrows point to TUNEL positive cells. Scale bar 50µM. (D) TUNEL staining (green) of representative brain section from a mouse 7 days after post-saline volume control injection. Inserts represent magnified images with ToPro (blue) and/or TUNEL staining (green). Scale bar 50µM. (E) TUNEL staining (green) of brain section from a mouse 7 days after collagenase injection into the striatum (lCH model). Insert shows ToPro (blue) and/or TUNEL positive cells (green). Scale bar 50µM 6) All hubs from WGCNA data are shown based on RNA-seq conducted in the paper. 7) Caspase inhibitor Z-VAD-FMK or Se protect against ER stress.