Journal: BioMed Research International

Article Title: IL-4 Alleviates Ischaemia-Reperfusion Injury by Inducing Kupffer Cells M2 Polarization via STAT6-JMJD3 Pathway after Rat Liver Transplantation

doi: 10.1155/2020/2953068

Figure Lengend Snippet: Liver IRI inhibition induced by IL-4 is due to KCs M2 polarization. KCs in the donor livers were depleted by clodronate liposome treatment, or donor liver JMJD3 was inhibited by GSK-J4 before liver transplantation. (a, b) Representative images of haematoxylin and eosin staining of liver graft at 6 h after liver transplantation (original magnification, ×200) and Suzuki's histological grading of liver IRI ( n = 6/group). (c, d) Levels of sAST and sALT were measured at 6 h after liver transplantation ( n = 6/group). The data are shown as mean ± SD, ∗ p < 0.05 vs. the Sham group, # p < 0.05 vs. the LT + IL-4 group.

Article Snippet: In the LT + GSK-J4 group, the donor rats were injected intraperitoneally with 5 mg/kg GSK-J4 (MCE, USA) for 3 consecutive days before liver transplantation.

Techniques: Inhibition, Transplantation Assay, Staining

Journal: Science advances

Article Title: KDM6B drives epigenetic reprogramming associated with lymphoid stromal cell early commitment and immune properties.

doi: 10.1126/sciadv.adh2708

Figure Lengend Snippet: Fig. 3. KDM6B inhibition partially impairs immunofibroblast commitment in vitro. (A) Heatmap visualization of the expression values of genes up-regulated by 6 hours (H6) of tumor necrosis factor–α (TNF)/lymphotoxin α1β2 (LT) treatment (n = 4, adjusted P < 0.05, fold change >2, DESeq2 analysis) in the presence of the KDM6 inhibitor GSK-J4 or dimethyl sulfoxide (DMSO) as a control. Z score is visualized through a color code. (B) Table of the 72 KDM6-specific genes repressed in the presence of GSK-J4. (C) Heatmap of a selection of lymphoid stromal cell (LSC)/immunofibroblast gene expression as defined by RNA sequencing (RNA-seq) analysis at H6 after TNF/LT treatment in the presence of the KDM6 inhibitor GSK-J4 or DMSO as a control (n = 4). Z score is visualized through a color code. (D) gp38/podoplanin (PDPN) cell surface protein expression quantified by flow cytometry on adipose-derived stromal cells (ASCs) treated or not with TNF/LT for 3 days (D3), in the presence or not of the KDM6 inhibitor GSK-J4 (n = 9). (Values are means ± SD, ****P < 0.0001). (E) IL-8 concentration was assessed by Luminex in the supernatants of ASC treated or not with TNF/LT for D3, in the presence or not of the KDM6 inhibitor GSK-J4 (n = 6). Values are means ± SD, P = ns). (F) Expression of LSC/immunofibroblast genes in ASC treated or not with TNF/LT for D3, in the presence or not of the KDM6 inhibitor GSK-J4 (n = 5 for IL1B, CCL5, and TNFRSF9, and n = 9 for the others, values are mean, FDR-adjusted *P < 0.05, **P < 0.01, ***P < 0.001). (D to F) Statistical analyses were performed using Mann-Whitney tests.

Article Snippet: When indicated, the KDM6 inhibitor GSK-J4 (20 μM) (Tocris) or dimethyl sulfoxide as a control were added each day.

Techniques: Inhibition, In Vitro, Expressing, Control, Selection, Gene Expression, RNA Sequencing, Flow Cytometry, Derivative Assay, Concentration Assay, Luminex, MANN-WHITNEY

Journal: Science advances

Article Title: KDM6B drives epigenetic reprogramming associated with lymphoid stromal cell early commitment and immune properties.

doi: 10.1126/sciadv.adh2708

Figure Lengend Snippet: Fig. 4. KDM6B supports immunofi- broblast commitment in vivo. (A) Seurat score enrichment of the KDM6- specific signature (right) in mouse steady-state (ss) fibroblast single-cell RNA sequencing (scRNAseq) atlas (32) reanalyzed by Uniform Manifold Ap- proximation and Projection (UMAP) (left). (B) RNA sequencing (RNA-seq) analysis of Pdpn+ fibroblasts purified from murine salivary glands at different time points before (H0) or after [3 hours (H3), 1 day (D1), 3 days (D3); 8 days (D8) and 23 days (D23)] tertiary lymphoid structure (TLS) induction (n = 3). Differentially ex- pressed genes (adjusted P < 0.05, fold change >2 or < −2, DESeq2 analysis) were represented through an heatmap, with z score visualized through a color code. (C) Heatmap showing the normal- ized counts of selected genes in Pdpn+

Article Snippet: When indicated, the KDM6 inhibitor GSK-J4 (20 μM) (Tocris) or dimethyl sulfoxide as a control were added each day.

Techniques: In Vivo, RNA Sequencing, Purification

Journal: Science advances

Article Title: KDM6B drives epigenetic reprogramming associated with lymphoid stromal cell early commitment and immune properties.

doi: 10.1126/sciadv.adh2708

Figure Lengend Snippet: Fig. 5. Early activation of KDM6B affects immunofibroblast immune properties. (A) KDM6B enrichment on CCL2 and CCL5 regulatory elements obtained by CUT&RUN quantitative polymerase chain reaction (qPCR) in adipose-derived stromal cells (ASCs) treated or not (CTR) with tumor necrosis factor–α (TNF)/lymphotoxin α1β2 (LT) for 6 hours (H6) (n = 5). Immunoglobulin G (IgG) was used as negative control. (B) H3K27me2 enrichment on CCL2 and CCL5 regulatory elements obtained by CUT&RUN qPCR in ASCs treated or not with TNF/LT for 3 days (D3) (n = 5). Enrichment was normalized to a positive control region (MYT1). (C) H3K27ac enrichment on CCL2 and CCL5 regulatory elements obtained by CUT&RUN qPCR in ASCs treated or not (CTR) with TNF/LT for D3, in the presence or not of the KDM6 inhibitor GSK-J4 (n = 5). Enrichment was normalized to a positive control region (ACTB). (D) RT-qPCR of CCL2 (n = 9) and CCL5 (n = 5 in ASC treated or not (CTR) with TNF/LT for D3 in the presence or not of the KDM6 inhibitor GSK-J4. (E) CCL2 and CCL5 concentration were assessed by Luminex in supernatants of ASC treated or not (CTR) with TNF/LT for D3, in the presence or not of the KDM6 inhibitor GSK-J4 (n = 6). (F) Migration of monocytes purified from healthy donor peripheral blood in response to supernatants collected from ASCs treated or not (CTR) with TNF/LT for D3, in the presence or not of the KDM6 inhibitor GSK-J4. When indicated, a specific CCR2 antagonist was added. Cell migration index is calculated as the number of DAPI−CD14+ viable monocytes migrating in response to cell supernatants divided by their number in response to migration medium (n = 7). (A) to (F) Statistical analyses were performed using Mann-Whitney tests or Wilcoxon matched-pairs signed rank tests (values are means ± SD, *P < 0.05, **P < 0.01, ****P < 0.0001).

Article Snippet: When indicated, the KDM6 inhibitor GSK-J4 (20 μM) (Tocris) or dimethyl sulfoxide as a control were added each day.

Techniques: Activation Assay, Real-time Polymerase Chain Reaction, Derivative Assay, Negative Control, Positive Control, Quantitative RT-PCR, Concentration Assay, Luminex, Migration, Purification, MANN-WHITNEY

Journal: Clinical immunology (Orlando, Fla.)

Article Title: LL37/self-DNA complexes mediate monocyte reprogramming.

doi: 10.1016/j.clim.2024.110287

Figure Lengend Snippet: Fig. 2. LL37 alone and LL37/self-DNA-mediated epigenetic modifications and associated signaling in HD monocytes. (A) Assessment of KDM6A/B enzymatic activity in HD monocytes 24 h and 4 days post-initial stimulation with LL37 alone and LL37/self-DNA complexes by ELISA (n = 5). (B) Western blot analysis depicting H3K27me3 and H3K4me3 marks in HD monocytes treated with LL37 alone and LL37/self-DNA complex for 24 h and following 4 days' rest (n = 4). (C) Comparison of epigenetic modifications in LL37/self-DNA-treated HD monocytes, demonstrating significantly decreased H3K27me3 marks and increased H3K4me3 marks at the promoter regions of IL6, IL8, and TNF genes compared to untreated HD monocytes 24 h and (D) 4 days post-initial stimulation (n = 5). (E) Western blot analysis depicting activation p-STAT3, STAT3, p-IκB-α and IκB-α in HD monocytes treated with LL37 alone and LL37/self-DNA complexes for 24 h and 4 days of rest (n = 4). (F) Western blot analysis of HD monocytes from untreated or treated with LL37/self-DNA complexes for 24 h in the presence of 2-DG or AA (Antimycin A) and Rot (Rotenone) inhibitors (Error bars show means ± SEM, P values were calculated using Wilcoxon signed rank test and one-way ANOVA followed by Dunnett's multiple comparisons test.).

Article Snippet: GSK-J4, a selective KDM6A/ B inhibitor, was obtained from MedChemExpress (Cat# HY-15648F).

Techniques: Activity Assay, Enzyme-linked Immunosorbent Assay, Western Blot, Comparison, Activation Assay

Journal: Molecular Brain

Article Title: Histone modification of pain-related gene expression in spinal cord neurons under a persistent postsurgical pain-like state by electrocautery

doi: 10.1186/s13041-021-00854-y

Figure Lengend Snippet: Effect of an H3K27 demethylase inhibitor on electrocautery-induced allodynia. A Schematic diagram of the mechanism of action of GSK-J4. B Protocol for measurement of the pain threshold and drug administration. C Changes in the pain threshold as measured by the von Frey test (0.16 g). Each point represents the mean ± S.E.M. of 5 mice (Two-way repeated measures ANOVA with post-hoc Bonferroni test, *p < 0.05, **p < 0.01, ***p < 0.001 vs. Sham-Vehicle group, # p < 0.05 vs. Electrocautery-Vehicle group). D , E The mRNA levels of Sprr1a D or Anxa10 E in the ipsilateral side of the spinal cord were measured at 18 days after surgery. Each column represents the mean ± S.E.M. of 5 independent experiments (One-way ANOVA with post-hoc Bonferroni test, *p < 0.05, ***p < 0.001 vs. Sham-Vehicle group, # p < 0.05, ### p < 0.001 vs. Electrocautery-Vehicle group)

Article Snippet: GSK-J4 was prepared daily, in saline (0.9% NaCl) containing 1% DMSO (FUJIFILM Wako Pure Chemical Co. LTD, Osaka, Japan), and mixed at room temperature for at least 1 h before use.

Techniques: