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. 2022 Dec 19;26(1):12–26. doi: 10.1038/s41593-022-01221-3

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© The Author(s) 2022

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 3 — a, Schema for co-cultures ± microglia. Illustrations were created with BioRender. b, Draq7⁺ death kinetics in tri-cultures exposed to 1,600 µM iron + 1 µM RSL3 ± microglia (n = 3 biologically independent experiments). Two-way ANOVA, Dunnett post hoc. P values are indicated in the graph. c, AUC of death kinetics (n = 3 biologically independent experiments). Two-way ANOVA, Dunnett post hoc. P values are indicated in the graph. Error bars represent s.e.m. d, Representative images of cultures ± microglia under vehicle or ferroptotic conditions. βIII-tubulin⁺ (yellow) neurons are dying and have increased lipid peroxidation (green) in cultures with IBA1⁺ (red) microglia (n = 4 biologically independent experiments). Scale bar, 100 µm. e, Average neuronal surface area in cultures ± microglia (n = 3 for no iron + RSL3; n = 4 biologically independent experiments for all other conditions). Two-way ANOVA, Sidak post hoc. P values are indicated in the graph. Error bars represent s.e.m. f, Lipid peroxidation in neurons (n = 3 for no iron + RSL3; n = 4 biologically independent experiments for all other conditions). Log-transformed, two-way ANOVA, Dunnett post hoc. P values are indicated in the graph. Error bars represent s.e.m. g, Correlation of neuronal surface area and neuronal lipid peroxidation (n = 33 wells examined over three biologically independent experiments). Two-tailed simple linear regression. h, Correlation of microglia number and neuronal surface area (n = 48 wells examined over four biologically independent experiments). Two-tailed simple linear regression. i, Correlation of microglia number and neuronal lipid peroxidation (n = 33 wells examined over three biologically independent experiments). Two-tailed simple linear regression. j,k, IL-8 (j) and IL-1β (k) production in supernatants from co-cultures ± microglia ± ferroptosis treatment (n = 4 biologically independent experiments). One-way ANOVA, Dunnett post hoc. P values are indicated in the graph. Error bars represent s.e.m. D, day.