Figure 4.
Atg7 deficiency triggers microglia ferroptosis. (A) Immunoblots for ATG7, chaperone protein BiP, and UPR pathway components (Xbp1s, ATF4, CHOP, and ATF6) on primary microglia cell lysates. Numbers below each blot represent the protein level relative to WT. Each blot represents two to four biological replicates. (B) mRNA levels of Hspa5 (BiP), Ddit3 (CHOP), and Xbp1 in primary microglia treated with 40-μM Aβ42 peptide for 48 h. Each data point represents a technical repeat. *, P < 0.05; **, P < 0.01 by two-tailed unpaired t test. Data are presented as mean ± SEM. (C) Protein synthesis rates of primary microglia, quantified by the amount of fluorescent-labeled nascent peptides. Each data point represents a technical repeat. *, P < 0.05 by two-tailed unpaired t test. Data are presented as mean ± SEM. (D) Representative confocal images showing the ferritin+ microglia (stained with antibody-targeting FTL) in subiculum areas from 8-mo-old mice. Scale bar = 100 μm. (E) Quantification of the percentage of FTL+ microglia in the subiculum of 5-, 8-, and 12-mo-old mice. Each point represents data from one mouse, with two technical replicates. *, P < 0.05; **, P < 0.01 by two-tailed unpaired t test. Data are presented as mean ± SEM. (F) Representative FACS plots (left) and quantification (right) of Annexin V and 7AAD staining on primary microglia after 48 h treatment of 40-μM Aβ42 peptides. Each data point represents a technical replicate. **, P < 0.01 by unpaired t test. Data are presented as mean ± SEM. (G) Quantification of the percentage of C11-Bodipy+ primary microglia cells after 48 h incubation with or without 40-μM Aβ42 peptides. Each data point represents a technical replicate. *, P < 0.05 by unpaired t test. Data are presented as mean ± SEM. (H) Quantification of Mitotracker Green gMFI in primary microglia cells after 48 h incubation with or without 40-μM Aβ42 peptides. Each data point represents a technical replicate. *, P < 0.05; ***, P < 0.001 by unpaired t test. Data are presented as mean ± SEM. Source data are available for this figure: SourceData F4. Refer to the image caption for details.

Atg7 deficiency triggers microglia ferroptosis. (A) Immunoblots for ATG7, chaperone protein BiP, and UPR pathway components (Xbp1s, ATF4, CHOP, and ATF6) on primary microglia cell lysates. Numbers below each blot represent the protein level relative to WT. Each blot represents two to four biological replicates. (B) mRNA levels of Hspa5 (BiP), Ddit3 (CHOP), and Xbp1 in primary microglia treated with 40-μM Aβ42 peptide for 48 h. Each data point represents a technical repeat. *, P < 0.05; **, P < 0.01 by two-tailed unpaired t test. Data are presented as mean ± SEM. (C) Protein synthesis rates of primary microglia, quantified by the amount of fluorescent-labeled nascent peptides. Each data point represents a technical repeat. *, P < 0.05 by two-tailed unpaired t test. Data are presented as mean ± SEM. (D) Representative confocal images showing the ferritin+ microglia (stained with antibody-targeting FTL) in subiculum areas from 8-mo-old mice. Scale bar = 100 μm. (E) Quantification of the percentage of FTL+ microglia in the subiculum of 5-, 8-, and 12-mo-old mice. Each point represents data from one mouse, with two technical replicates. *, P < 0.05; **, P < 0.01 by two-tailed unpaired t test. Data are presented as mean ± SEM. (F) Representative FACS plots (left) and quantification (right) of Annexin V and 7AAD staining on primary microglia after 48 h treatment of 40-μM Aβ42 peptides. Each data point represents a technical replicate. **, P < 0.01 by unpaired t test. Data are presented as mean ± SEM. (G) Quantification of the percentage of C11-Bodipy+ primary microglia cells after 48 h incubation with or without 40-μM Aβ42 peptides. Each data point represents a technical replicate. *, P < 0.05 by unpaired t test. Data are presented as mean ± SEM. (H) Quantification of Mitotracker Green gMFI in primary microglia cells after 48 h incubation with or without 40-μM Aβ42 peptides. Each data point represents a technical replicate. *, P < 0.05; ***, P < 0.001 by unpaired t test. Data are presented as mean ± SEM. Source data are available for this figure: SourceData F4.

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