Journal: CNS Neuroscience & Therapeutics

Article Title: Targeting HDAC3 Suppresses Ferroptosis and Demyelination in White Matter Injury by Restoring PDK4‐Mediated Iron Homeostasis

doi: 10.1111/cns.70471

Figure Lengend Snippet: Inhibition of HDAC3 regulates iron metabolism in HDAC3‐PDK4 axis and improves ferroptosis by lipid peroxidation. (A) Representative western blot bands of ACSL4, xCT and GPX4 in sham, CPZ and RGFP966 intervention group. (B) Quantification of hepcidin antimicrobial peptide ( Hamp ) mRNA expression examined by quantitative real‐time PCR (qPCR) in white matter after RGFP966 intervention. (C) Fe 2+ expression in white matter after RGFP966 intervention. (D) Quantification of the transferrin (TF) levels by Enzyme‐Linked Immunosorbent Assay (ELISA) at 5 weeks CPZ‐modeling in CC region. n = 3–7. (E–G) Quantitative analysis of protein expression levels of ACSL4, xCT and GPX4 in CC region. n = 3–6. (H) Malondialdehyde (MDA) was detected in the CC region of mice. (I) Quantitative analysis of GSH expression levels in white matter of mouse in each group; (J) The activity level of superoxide dismutase (SOD) in the white matter of mouse in each group. n = 3–5. (K) CUT&Tag sequencing Reads enrichment map. The graph above shows the gene region in horizontal coordinates and the signal enrichment intensity in vertical coordinates. The following is a heat map. The horizontal coordinate represents 5 kb upstream and 5 kb downstream of the transcription initiation site (TSS), the vertical coordinate represents genes, and the color represents the coverage of reads at corresponding locations. The redder the color, the higher the coverage of reads at that location. (L) In the KEGG enrichment results of peak‐related genes, select the pathway with the most significant enrichment and show it in the figure. The enrichment of KEGG Pathway was measured by RichFactor, Qvalue and the number of genes enriched in this pathway. (M) Peak GO enrichment analysis of CPZ group and RGFP966 group. (N, O) KEGG analysis diagram and GO enrichment analysis of NCBI dataset GSE151306 . (P, Q) The top five genes with high logFC values among 38 intersection genes in NCBI dataset and ferroptosis gene dataset. (R–V) Quantification of PDK4, ENPP2, MLLT1, CDH1 and CDO1 mRNA expression examined by qPCR in the white matter of mouse after 5 weeks of demyelination. n = 3. (W–X) Quantification of CDH1 and PDK4 mRNA expression examined by qPCR in the CC region of mouse after 1 week of intervention with RGFP966. (Y) Reads expression of PDK4 in IGV visual CUT&Tag sequencing data. (Z) Quantification of the binding effect of HDAC3 and PDK4 expression examined by CUT&Tag‐qPCR after demyelination for 1 week. n = 3.

Article Snippet: Mouse transferrin (TF) levels were measured using a commercial enzyme‐linked immunosorbent assay (ELISA) kit (Elabscience, E‐EL‐M1184, Elabscience Biotechnology Co. Ltd., Wuhan, Hubei, China).

Techniques: Inhibition, Western Blot, Expressing, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Activity Assay, Sequencing, Binding Assay

Journal: Journal of Nanobiotechnology

Article Title: Macrophage-derived extracellular vesicles represent a promising endogenous iron-chelating therapy for iron overload and cardiac injury in myocardial infarction

doi: 10.1186/s12951-024-02800-1

Figure Lengend Snippet: Iron overload and cell ferroptosis contributes to MI-induced cardiac injury. a Quantitative analysis of serum iron level in sham and MI mice. b Quantitative analysis of cardiac iron level in sham and MI mice. c , d Representative western blotting images and quantitative analysis of transferrin and FTH1 expression in sham and MI mice. e Quantitative analysis of MDA level in sham and MI mice. f Quantitative analysis of GSH in sham and MI mice. g , h Representative western blotting images and quantitative analysis of GPX4 expression in sham and MI mice. i , j Representative immunohistochemical images and quantitative analysis of 4-HNE expression in sham and MI mice. Scale bar = 30 μm. N = 6 each group. Data are expressed as Mean ± SEM

Article Snippet: The measurement of transferrin level was performed with Transferrin ELISA kit (Elabscience, E-EL-M1184c) following manufacturer’s instructions.

Techniques: Western Blot, Expressing, Immunohistochemical staining

Journal: Journal of Nanobiotechnology

Article Title: Macrophage-derived extracellular vesicles represent a promising endogenous iron-chelating therapy for iron overload and cardiac injury in myocardial infarction

doi: 10.1186/s12951-024-02800-1

Figure Lengend Snippet: Macrophage-derived EVs attenuated myocardial injury by preventing iron overload in post-MI heart. a Diagram of MI construction and EVs injection protocol. b Quantitative analysis of serum iron level in mice. c Quantitative analysis of cardiac iron level in mice. d , e Representative western blotting images and quantitative analysis of transferrin and FTH1 expression in hearts of mice. f Quantitative analysis of MDA level in hearts of mice. g Representative DHE staining images of hearts of mice. Scale bar = 50 μm. h , i Representative western blotting images and quantitative analysis of GPX4 expression in hearts of mice. j Representative immunohistochemical images of 4-HNE expression in hearts of mice. Scale bar = 50 μm. k , l Representative TTC staining images and quantitative analysis of infarcted area of hearts. m Representative M-mode echocardiography images. n , o Quantitative analysis of LVEF and LVFS. p , q Quantitative analysis of serum CKMB and LDH1 in mice. N = 6 each group. Data are expressed as Mean ± SEM

Article Snippet: The measurement of transferrin level was performed with Transferrin ELISA kit (Elabscience, E-EL-M1184c) following manufacturer’s instructions.

Techniques: Derivative Assay, Injection, Western Blot, Expressing, Staining, Immunohistochemical staining

Journal: Journal of Nanobiotechnology

Article Title: Macrophage-derived extracellular vesicles represent a promising endogenous iron-chelating therapy for iron overload and cardiac injury in myocardial infarction

doi: 10.1186/s12951-024-02800-1

Figure Lengend Snippet: TfR was responsible for the iron-chelating capacity of macrophages-derived EVs. a Quantitative analysis of iron level in iron solution after incubation with PBS or EVs. b Representative western blotting images of TfR expression in macrophages-derived EVs. c Representative immunogold TEM images of macrophages-derived EVs incubated with IgG or anti-TfR. Scale bar = 100 nm. d Schematic diagram of the experiment protocol to determine the origin of TfR on EVs. e Quantitative analysis of fluorescence intensity of culture medium. f , g Representative flow cytometer results and quantitative analysis of fluorescence intensity of EVs released by macrophages transfected with empty-EGFP or TfR-EGFP plasmid. h Schematic diagram of the experiment protocol to determine whether TfR on EVs could bind with transferrin. i Representative flow cytometer results of EVs released by macrophages transfected with TfR-EGFP plasmid after incubation with cell transfected with transferrin-mCherry or empty-mCherry plasmid. j-m Quantitative analysis of transferrin level of myocardium lysate, serum, cardiomyocytes lysate, and culture medium of cardiomyocytes after incubation with macrophages-derived EVs. N = 6 each group. Data are expressed as Mean ± SEM

Article Snippet: The measurement of transferrin level was performed with Transferrin ELISA kit (Elabscience, E-EL-M1184c) following manufacturer’s instructions.

Techniques: Derivative Assay, Incubation, Western Blot, Expressing, Fluorescence, Flow Cytometry, Transfection, Plasmid Preparation

Journal: Journal of Nanobiotechnology

Article Title: Macrophage-derived extracellular vesicles represent a promising endogenous iron-chelating therapy for iron overload and cardiac injury in myocardial infarction

doi: 10.1186/s12951-024-02800-1

Figure Lengend Snippet: EVs released by macrophages with TfR deficiency exhibited reduced cardiac protective effect against MI. a Representative western blotting images of TfR expression in macrophages transfected with siTfR and EVs released by such macrophages. b Quantitative analysis of serum iron level of mice. c Quantitative analysis of cardiac iron level of mice. d , e Representative western blotting images and quantitative analysis of transferrin and FTH1 expression in hearts of mice. f Quantitative analysis of MDA level of mice. g Representative DHE staining images of hearts of mice. Scale bar = 50 μm. h , i Representative western blotting images and quantitative analysis of GPX4 expression in hearts of mice. j Representative immunohistochemical images of 4-HNE expression in hearts of mice. Scale bar = 50 μm. k , l Representative TTC staining images and quantitative analysis of infarcted area of hearts. m Representative M-mode echocardiography images. n , o Quantitative analysis of LVEF and LVFS. p , q Quantitative analysis of serum CKMB and LDH1 in mice. N = 6 each group. Data are expressed as Mean ± SEM

Article Snippet: The measurement of transferrin level was performed with Transferrin ELISA kit (Elabscience, E-EL-M1184c) following manufacturer’s instructions.

Techniques: Western Blot, Expressing, Transfection, Staining, Immunohistochemical staining

Journal: Journal of Nanobiotechnology

Article Title: Macrophage-derived extracellular vesicles represent a promising endogenous iron-chelating therapy for iron overload and cardiac injury in myocardial infarction

doi: 10.1186/s12951-024-02800-1

Figure Lengend Snippet: Schematic figure illustrating the mechanism underlying the therapeutic effect of macrophage-derived EVs on MI. Myocardial iron overload, cardiomyocytes ferroptosis, and cardiac function impairment were detected in the hearts post MI, which contributed to the poor prognosis of MI. This work identified that macrophage-derived EVs exhibited protective effect on reducing myocardial iron overload, alleviating cardiomyocytes ferroptosis, and improving cardiac function post MI. Mechanistically, TfR on macrophage-derived EVs was the key player responsible for binding with transferrin and removing protein-bound iron. These iron-chelating EVs were ultimately captured and processed by macrophages in the liver

Article Snippet: The measurement of transferrin level was performed with Transferrin ELISA kit (Elabscience, E-EL-M1184c) following manufacturer’s instructions.

Techniques: Derivative Assay, Binding Assay