Journal: Molecular Brain

Article Title: Ferroptosis participates in neuron damage in experimental cerebral malaria and is partially induced by activated CD8 + T cells

doi: 10.1186/s13041-022-00942-7

Figure Lengend Snippet: CD8 + T cells infiltrate the cerebrum of ECM mice and induce ferroptotic damage to primary neurons. a CD8 + T cells infiltrating the cerebrum of ECM mice were co-labeled with CD3 (green) and CD8 (red), and detected by immunofluorescence staining. The nucleus was stained with DAPI (blue). b Morphological changes in primary neurons were observed by microscopy (200 × magnification). c Expression of APP and MAP2 in primary neurons, as measured by western blotting, after co-culture with naïve or activated CD8 + T cells. d Expression of APP and MAP2 in primary neurons, as measured by western blotting after co-culture with activated CD8 + T cells with or without Fer-1. e Relative mitochondrial membrane potential (MMP) levels were calculated based on the optical density value and shown as the fold change relative to the blank value. Data are reported as mean ± standard deviation. Each point represents one sample. * p < 0.05 and *** p < 0.001 vs. control or activated CD8 + T cells. f Expression of TfR1 and GPX4 in primary neurons, measured by western blotting after co-culture with activated CD8 + T cells for 2–24 h. APP amyloid-beta precursor protein, ECM experimental cerebral malaria, GPX4 glutathione peroxidase 4, MAP2 microtubule-associated protein 2, TfR1 transferrin receptor 1

Article Snippet: Then the membranes were blocked with 5% skim milk for 1 h at room temperature, followed by incubation with the primary antibody against TfR1 (ET1702-06, HUABIO, China), ACSL4 (sc-271800, Santa Cruz, USA), GPX4 (ET1706-45, HUABIO, China), APP (25524-1-AP, Proteintech, China), MAP2 (17490-1-AP, Proteintech, China), and β-actin (66009-1-Ig, Proteintech, China) overnight at 4 °C, and then the secondary antibody DyLight 680-labbed Goat Anti-Mouse IgG and DyLight 800-labbed Goat Anti-Rabbit IgG (A23910, A23920, KPL, USA) for 1 h at room temperature avoiding light, and scanned at 700 nm and 800 nm using Odyssey Clx Image.

Techniques: Labeling, Immunofluorescence, Staining, Microscopy, Expressing, Western Blot, Co-Culture Assay, Membrane, Standard Deviation, Control

Journal: The American journal of pathology

Article Title: Molecular and functional alterations in a mouse cardiac model of Friedreich ataxia: activation of the integrated stress response, eIF2α phosphorylation, and the induction of downstream targets.

doi: 10.1016/j.ajpath.2013.05.032

Figure Lengend Snippet: Figure 6 Proposed pathogenesis of heart failure in FA. Frataxin defi- ciency results in the early activation of the ISR via phosphorylation (P) of eIF2a and induction of its downstream stress-inducible molecules, including Atf4 and Chop. The precise stress stimulus for the ISR activation is unclear, but may involve early and pronounced perturbation in heme synthesis (markedly reduced Fech expression) acting through Hri in KO mice, compared to WT mice. In addition, the later involvement of ER stress (up-regulation of Bip and cleaved caspase-12) could potentiate the prolonged activation of the ISR pathway. ISR activation subsequently enhances autophagy (up- regulation of Atg3, LC3-II, p62, and Fundc1) and apoptosis (increased expression of Chop, Bax and Bcl-2, and cleaved caspase-12), leading to cardiomyocyte death. Histological alterations are first apparent from 5 weeks of age with iron accumulation, followed by fibrosis and cardiomyocyte degeneration from 6 weeks of age. Severe progressive reduction in cardiac function ensues, resulting in heart failure. Casp12, caspase-12.

Article Snippet: Protein isolation and Western blot analysis were performed using established techniques.3 The primary antibodies used were against TfR1 (13-6890; Life Technologies), Atf4 (sc-200; SantaCruzBiotechnology,SantaCruz,CA),Mthfd2 (ab37840; Abcam, Cambridge, UK), Asns (1732-1; Epitomics, Burlingame,CA), Chop (SC-575; Santa Cruz), p-eIF2a (Ser51; 3398; Cell Signaling Technology, Danvers, MA), eIF2a (2103; Cell Signaling Technology), Atg3 (3415; Cell Signaling Technology), LC3 (PD014; MBL International, Woburn, MA), p62 (GP62-C; Progen Biotechnik, Heidelberg, Germany), caspase12 (2202; Cell Signaling Technology), Bax (2772; Cell Signaling Technology), Bcl-2 (2870; Cell Signaling Technology), Itgb1bp3 (K0099-3; MBL International), p-Perk (Thr980; 3179; Cell Signaling Technology), Perk (3192; Cell Signaling Technology), Bip (3177; Cell Signaling Technology), Hri (07-728; Millipore, Sydney, Australia), Fech (from Harry Dailey, University of Georgia30), Fundc1 (ARP53280_P050; Aviva Systems Biology, San Diego, CA), and Gapdh (SC-25778; Santa Cruz Biotechnology).

Techniques: Activation Assay, Phospho-proteomics, Expressing