Journal: Journal of Neurochemistry
Article Title: Neuronal Damage Induced by Gradual Oxidative Stress in i PSC ‐Derived Neurons: Implications for Ferroptosis Involvement and ALS Drug Evaluation
doi: 10.1111/jnc.70246
Figure Lengend Snippet: Protection of neurons from oxidative stress‐induced damage by ferroptosis inhibitors. (a–h) Time course changes in the total neurite length (mm/mm 2 ) of motor neurons (iCell Motor neurons) cultured in DMEM/F12 with antioxidants (+AO) or without antioxidants (−AO). Z‐VAD‐FMK, necrostatin‐1, necrosulfonamide, ferrostatin‐1, liproxstatin‐1, UAMC‐3203, RIPA‐56, or GSK‐872 was added under the −AO condition. The lines represent the average of three experiments with independent cell culture preparations. Each experiment consists of six wells for the −AO and +AO groups, and two wells for compound treatment. Error bars indicate SEM. Significant effects increasing AUC and H50 are indicated by # and *, respectively, as determined using two‐way ANOVA followed by Dunnett's multiple comparison test. # p = 0.0061, Z‐VAD‐FMK at 20 μmol/L; p < 0.0001, necrostatin‐1 at 2.2 and 20 μmol/L; p < 0.0001, ferrostatin‐1 at 0.11 and 1.0 μmol/L; p < 0.0001, liproxstatin‐1 at 0.11 and 1.0 μmol/L; p < 0.0001, UAMC‐3203 at 0.012, 0.11, and 1.0 μmol/L. * p < 0.0001, Z‐VAD‐FMK at 20 μmol/L; p < 0.0001, necrostatin‐1 at 2.2 and 20 μmol/L; p < 0.0001, ferrostatin‐1 at 0.11 and 1.0 μmol/L; p < 0.0001, liproxstatin‐1 at 0.11 and 1.0 μmol/L; p < 0.0001, UAMC‐3203 at 0.012, 0.11, and 1.0 μmol/L. See Tables and for detailed statistical analysis. Note that these compound assays were conducted using the same plate sets, but the results are presented in separate graphs for clarity. Therefore, panels a–h share the same control data (+AO and −AO). (i) Time course of changes in the total neurite length (mm/mm 2 ) of motor neurons cultured in +AO, −AO, or with deferoxamine under the −AO condition. The lines represent the average of three experiments with independent cell culture preparations. Error bars indicate SEM. Significant effects increasing AUC and H50 are indicated by # and *, respectively, as determined using two‐way ANOVA followed by Dunnett's multiple comparison test (# and * P < 0.0001 at 5.0 μmol/L). See Tables and for detailed statistical analysis. DMSO at 0.5% was included as the vehicle. (j) Flow cytometry analysis of BODIPY 581/591 C11 oxidation in motor neurons cultured in +AO, −AO, with ferrostatin‐1 (1.0 μmol/L) or AY 9944 (0.1 μmol/L) under the −AO condition for 5 days. (k) Representative images at 168 h of iCell Motor neurons and 312 h of axoCells Human iPSC‐Derived Motor Neurons cultured in DMEM/F12 with (+AO) or without AO (−AO), or with ferrostatin‐1 (1.0 μmol/L) under the −AO condition. Scale bar: 100 μm.
Article Snippet: Human iPSC‐derived motor neurons were purchased from FUJIFILM Cellular Dynamics (iCell Motor Neurons‐01279, cat. no. C1048) and Axol Bioscience (axoCells Human iPSC‐Derived Motor Neurons, cat. no. AX0078). iCell Motor neurons were primarily used as motor neurons, unless otherwise specified.
Techniques: Cell Culture, Comparison, Control, Flow Cytometry, Derivative Assay
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