class i hdac inhibitors  (Huntsman International LLC)

Journal: Cell Death & Disease

Article Title: Insulin enables acquisition of the IL7R + memory phenotype in PD1 + T cells in RA tissues

doi: 10.1038/s41419-026-08916-6

Figure Lengend Snippet: A Glucose metabolism. Studied metabolic pathways are in bold. B Box plots of standardized expression sum (SES), by RNA-seq, of genes within metabolic pathways in CD4 + cells of RA ( n = 11), healthy controls (HC, n = 57), and of CD4 + cells split by mean expression of BIRC5 gene in high (B5 hi ) and low (B5 lo ) ( n = 24). Boxes indicate IQR and whiskers indicate the minimum and maximum values. GO-terms used for pathway annotation: Glycolysis (GO:0006096); oxidative phosphorylation (OxPhos, GO:0006119 and GO:0022900); pentose phosphate pathway (PPP, GO:0006098); tricarboxylic acid cycle (TCA, GO:0006099). P -values are calculated by Wilcoxon unpaired test. C Heatmap of Spearman’s correlation between HAT, HDAC and metabolic pathways as above. GO-terms used for annotation HAT (GO:0000123) and HDAC (GO:0000118). Asterisks indicate p -values. * < 0.05, ** < 0.01, *** <0.001. D Heatmap of Spearman’s correlation between HAT and HDAC and insulin signaling (IS). Asterisks indicate p -values * < 0.05, ** < 0.01, *** <0.001. E Scatter plot of Spearman’s correlation between plasma insulin levels and SES of insulin signaling. F Uniform manifold approximation and projection (UMAP) map of scaled expression intensity of BIRC5 , IFNG , and TNF in T cells of RA synovial tissue (ST), by single cell transcriptome. BIRC5 Hi CD4 + clusters are indicated. G Heatmap of scaled expression intensity of metabolic pathway (annotated as above) in peripheral blood (PB), synovial fluid (SF), and ST of RA patients, by scRNA-seq. H Violin plot of insulin signaling by SES of INSR, IRS1, IRS2 and IGF1R genes in BIRC5 Hi CD4 + clusters. P-value was calculated by chi-square test. I Violin plot of metabolic pathways SES in cells with high and low IS in BIRC5 Hi CD4 + clusters. P -values are calculated by Wilcoxon unpaired test. Asterisks indicate * < 0.05, ** < 0.01, *** <0.001, **** <0.0001 J . Box plot of BIRC5, IFNG , and TNF expression in Tph cells with high and low IS. Boxes indicate IQR and whiskers indicate the minimum and maximum values. P -values are calculated by Wilcoxon unpaired test. K Heatmap of expression for IL7R-signaling targets in Tph cells high and low IS. Genes regulated by survivin-H3K27ac are in bold. P -values are calculated by Wilcoxon unpaired test. Asterisks indicate * < 0.05, ** < 0.01, *** <0.001, **** <0.0001.

Article Snippet: Cells were stimulated with concanavalin A (ConA, 0.625 μg/mL, MP Biomedicals), and lipopolysaccharide (LPS, 5 μg/mL, Sigma-Aldrich) for 48 h. For immunophenotyping, the cultures were supplemented with insulin (Humalog 100 U/mL, Eli Lilly, Indianapolis, IN, USA) 25 nM and/or the class I HDAC inhibitor (HDACi) valproate, 50 μg/mL (stock 200 mg/mL, Ergenyl, Sanofi, Paris, France) after 24 h. Supernatants were collected for cytokine measures and cells were analyzed by flow cytometry.

Techniques: Expressing, RNA Sequencing, Phospho-proteomics, Clinical Proteomics, Single Cell

Journal: Cell Death & Disease

Article Title: Insulin enables acquisition of the IL7R + memory phenotype in PD1 + T cells in RA tissues

doi: 10.1038/s41419-026-08916-6

Figure Lengend Snippet: A Confocal microscopy images of THP1 cells stained with antibodies to H3K27ac (red) in nucleus (blue) after stimulation with insulin (top), and HDAC-inhibitor (bottom) for 24 h. Images are acquired with 40× magnification with additional digital magnification 1.5×. B Scatter plot of nuclear H3K27ac staining intensity after stimulation with insulin (top) and HDAC-inhibitor (bottom). P -values are calculated by Wilcoxon unpaired test. Asterisks indicate * < 0.05, ** < 0.01, *** <0.001, **** <0.0001. C Histogram of phosphorylated serine 473 (p)AKT1 mean fluorescence intensity in THP1 cells stimulated with 10 nM insulin, by flow cytometry. D Histogram of H3K27ac mean fluorescence intensity in THP1 cells stimulated with increasing concentrations of insulin for 24 h, by flow cytometry. E Histogram of pAKT1 mean fluorescence intensity in human lymphocytes stimulated with increasing concentrations of insulin for 30 min, by flow cytometry.

Article Snippet: Cells were stimulated with concanavalin A (ConA, 0.625 μg/mL, MP Biomedicals), and lipopolysaccharide (LPS, 5 μg/mL, Sigma-Aldrich) for 48 h. For immunophenotyping, the cultures were supplemented with insulin (Humalog 100 U/mL, Eli Lilly, Indianapolis, IN, USA) 25 nM and/or the class I HDAC inhibitor (HDACi) valproate, 50 μg/mL (stock 200 mg/mL, Ergenyl, Sanofi, Paris, France) after 24 h. Supernatants were collected for cytokine measures and cells were analyzed by flow cytometry.

Techniques: Confocal Microscopy, Staining, Fluorescence, Flow Cytometry

Journal: Cell Death & Disease

Article Title: Insulin enables acquisition of the IL7R + memory phenotype in PD1 + T cells in RA tissues

doi: 10.1038/s41419-026-08916-6

Figure Lengend Snippet: A Analysis strategy of survivin and H3K27ac deposition, by ChIP-seq, in cis-regulatory elements ( cis -RE) and connected genes in human CD4 + T cells. B Venn diagram of insulin-responsive (InsResp) genes connected to cis -RE with survivin-H3K27ac co-deposition. Genes are identified by DESeq2 analysis of CD4 + cells transcriptome after a) direct insulin stimulation (InsStim) and b) in regression to corresponding plasma insulin levels, by RNA-seq. C Scatter plot of pathway enrichment of insulin-responsive genes, by GSEA GO-terms. Circle size indicates the number of genes in the pathway, and color intensity indicates p -value. D Venn diagram of insulin-responsive (InsResp) genes connected to cis -RE with H3K27ac deposition. Genes are identified by DESeq2 analysis of CD4 + cells transcriptome after a) direct insulin stimulation (InsStim) and b) HDAC inhibition. E Scatter plot of protein subunits with survivin-H3K27ac in HAT (E1) and HDAC (E3) complexes, annotated by STRING database. Circle size represents number of cis- RE with survivin-H3K27ac connected to the gene. Circle color represents beta-coefficient of expression difference by DESeq2 analysis of CD4 + cell transcriptome in regression to corresponding plasma insulin levels. (E2 and E4) Heatmap of transcription difference, by log2FoldChange (FC), in CD4 + cells in regression to plasma insulin levels (InsReg), and after HDAC inhibitor treatment in vitro. F . Bar plot of percentage of H3K27ac and survivin-H3K27ac connected genes in each metabolic pathway. GO-terms used for pathway annotation as in Fig. . G Scatter plot of expression difference of survivin-H3K27ac connected genes in transcriptome of CD4 + cells in regression to plasma insulin levels and after HDACi treatment, by RNA-seq. Expression difference is calculated by DESeq2 analysis. Colour fill corresponds to metabolic pathway annotation of the gene.

Article Snippet: Cells were stimulated with concanavalin A (ConA, 0.625 μg/mL, MP Biomedicals), and lipopolysaccharide (LPS, 5 μg/mL, Sigma-Aldrich) for 48 h. For immunophenotyping, the cultures were supplemented with insulin (Humalog 100 U/mL, Eli Lilly, Indianapolis, IN, USA) 25 nM and/or the class I HDAC inhibitor (HDACi) valproate, 50 μg/mL (stock 200 mg/mL, Ergenyl, Sanofi, Paris, France) after 24 h. Supernatants were collected for cytokine measures and cells were analyzed by flow cytometry.

Techniques: ChIP-sequencing, Clinical Proteomics, RNA Sequencing, Inhibition, Expressing, In Vitro

Journal: Cell Death & Disease

Article Title: Insulin enables acquisition of the IL7R + memory phenotype in PD1 + T cells in RA tissues

doi: 10.1038/s41419-026-08916-6

Figure Lengend Snippet: A Transition model of PD1 hi Tph cells to the IL7R + T memory cells, influenced by insulin and HDAC inhibition. B Box plot of protein cytokine levels in supernatants of CD4 + cells ( n = 12) stimulated with insulin (25 nM) and HDACi (50 µg/mL) for 24 h, by ELISA. Boxes indicate IQR and whiskers indicate the minimum and maximum values. P -values are calculated by Wilcoxon Sign rank test. C Gating strategy to CD4 + T cell subset analysis by flow cytometry. D Box plot of PD1 and CD27 expression in CD4 + cells ( n = 8) stimulated by insulin and HDACi, by mean fluorescence intensity (MFI). Boxes indicate IQR and whiskers indicate the minimum and maximum values. P -values are calculated by Wilcoxon Sign rank paired test. E Heatmap of expression difference of IL7R-related signaling molecules in CD4 + cells after in vitro stimulation with insulin (25 nM), HDACi (50 µg/mL), IFNγ (50 ng/ml), and survivin inhibitor YM155 (10 μM), by RNA-seq. Genes connected to cis -RE containing survivin-H3K27ac co-deposition are indicated bold. Asterisks indicate RNA-seq p -values * < 0.05, ** < 0.01, *** <0.001, **** <0.0001. F Heatmap showing expression difference, by log2FC, of cluster markers of Tph cluster after insulin stimulation, HDAC inhibition. In bold are the genes supervised by a cis-RE containing survivin-H3K27ac deposition. P -values are calculated by DESeq2 analysis. Asterisks indicate nominal p -values * < 0.05, ** < 0.01, *** <0.001, **** <0.0001.

Article Snippet: Cells were stimulated with concanavalin A (ConA, 0.625 μg/mL, MP Biomedicals), and lipopolysaccharide (LPS, 5 μg/mL, Sigma-Aldrich) for 48 h. For immunophenotyping, the cultures were supplemented with insulin (Humalog 100 U/mL, Eli Lilly, Indianapolis, IN, USA) 25 nM and/or the class I HDAC inhibitor (HDACi) valproate, 50 μg/mL (stock 200 mg/mL, Ergenyl, Sanofi, Paris, France) after 24 h. Supernatants were collected for cytokine measures and cells were analyzed by flow cytometry.

Techniques: Inhibition, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Expressing, Fluorescence, In Vitro, RNA Sequencing

Journal: eNeuro

Article Title: Evidence That Dmrta2 Acts through Repression of Pax6 in Cortical Patterning and Identification of a Mutation Impairing DNA Recognition Associated with Microcephaly in Human

doi: 10.1523/ENEURO.0377-24.2025

Figure Lengend Snippet: The Dmrta2-Pax6 interaction requires the recruitment of the HDAC-NuRD complex by Zfp423. A , B , Coimmunoprecipitation assays as indicated with Zfp423, Flag-Dmrta2, and a Flag-Dmrta2 mutant lacking the DM domain (Flag-Dmrta2 ΔDM) overexpressed in HEK293T cells as indicated. Note in A that Flag-Dmrta2 pulled down Zpf423 (line 1–2) and that, conversely, Zpf423 coprecipitated Flag-Dmrta2 (line 3–4). Note in B that Flag-Dmrta2 ΔDM does not pull down Zfp423 (line 4). n = 3. Segments from the same blot have been spliced together to show side by side the results of the WT and the ΔDM Dmrta2 mutant. C , Full-length mouse Myc-tagged Zfp423 was synthesized in vitro and incubated with GST alone or with GST fusion proteins bound to glutathione-agarose beads as indicated. Bound Myc-tagged Zfp423 was detected by immunoblot with an anti-Myc antibody. A 2% input sample was loaded for comparison. The corresponding Coomassie-stained gel is shown. n = 2. D , Coimmunoprecipitation assays as indicated with HEK293T cells transfected with a Flag -Dmrta2 expression construct, alone or together with increasing doses of a Zfp423 expression construct. Note that Dmrta2 immunoprecipitates some NuRD subunits and that the binding of Zfp423 to Dmrta2 increases the amount of coimmunoprecipitated HDAC1/2 and MBD3. Densitometric quantification of the western blot results is shown in Extended Data . n = 3. E , Reporter assays in P19 cells transfected with a Pax6 E60 tk-luc reporter vector, or an “empty” tk-luc reporter vector as indicated, together with a pCS2Myc -Pax6 expression vector and/or a pCS2Flag -Dmrta2 expression vector as indicated, in the absence (white bars) or presence of increasing doses of the HDAC1 inhibitor romidepsin (gray bars). Note that romidepsin leads to a stronger increase of luciferase activity reaching significance in the presence of Dmrta2 but not in its absence. The mean activity of the Pax6 E60 enhancer reporter construct with cotransfected Pax6 is set to 1. NS, not significant. * p < 0.05, one-way ANOVA test. Results of similar reporter assays performed in P19 cells, in the presence or absence of Zfp423 are presented in Extended Data . F , Reporter assays in HEK293T cells show that both Gal4-Dmrta2 and the Gal4-Dmrta2 (126–531) fusion construct lacking the DM domain required for Zfp423 interaction has strong repression activity on the 5XUAS-tk-luc reporter construct and that Zfp423 slightly increases the repressive activity of Gal4-Dmrta2 but not of the Gal4-Dmrta2 (126–531) construct. In each condition, 200 ng of the 5XUAS-tk-luc reporter was transfected, together with 25 ng of the pCMV-Gal4-Dmrta2 or the pCMV-Gal4-Dmrta2 (126–531) and different doses (200, 400 and 600 ng) of pCDNA3-Myc -Zfp423 expression plasmids. Values represent the mean ± SD of one transfection done in triplicate. A Western blot showing the expression levels of the overexpressed factors is shown below. Reporter assays showing that the Gal4-Dmrta2 fusion protein represses in a UAS-dependent manner the activity of the 5XUAS-tk-luc reporter are presented in Extended Data . Reporter assays in HEK293T cells showing that Zfp423 does not increase the modest repression observed when an expression vector encoding the Gal4 DNA-binding domain alone is cotransfected with the 5XUAS-tk-luc reporter are presented in Extended Data .

Article Snippet: The HDAC class I inhibitor Romidepsin (MedChemExpress, catalog #HY-15149) was used at 0.005–0.01 μm.

Techniques: Mutagenesis, Synthesized, In Vitro, Incubation, Western Blot, Comparison, Staining, Transfection, Expressing, Construct, Binding Assay, Plasmid Preparation, Luciferase, Activity Assay

Journal: bioRxiv

Article Title: Short chain fatty acids regulate the chromatin landscape and distinct gene expression changes in human colorectal cancer cells

doi: 10.1101/2025.05.07.652677

Figure Lengend Snippet: (A) Schematic of experimental set-up. Cells were treated with either vehicle (DMSO), 5 mM butyrate, 1 μM Entinostat, 1 μM A485, or A485 with butyrate for 24 hours in HPLM media. Image was created with Biorender.com. (B) Levels of select histone marks were analyzed by immunoblotting with the different treatments. H3 serves as a loading control and representative blot is shown. (C) Quantitative analysis of signal intensity for the histone marks normalized to total histone H3 levels. Immunoblot signal intensities were quantified using Image Lab 6.1, and the bar graphs represent the mean and s.e.m. for three independent experiments (n = 3). Statistical significance was evaluated using one-way ANOVA with Dunnett’s test for multiple comparisons. (D) Heatmap of differential gene expression. Z-scores of rlog(counts) are depicted of significant genes following Likelihood Ratio Test (padj < 0.05). (E) Heatmap and hierarchical clustering of differential gene expression after removing Entinostat-dependent genes. Entinostat-dependent genes were defined as padj <0.05 in DMSO vs HDACi. Z-scores of rlog(counts) are depicted of significant genes following Likelihood Ratio Test (padj < 0.05). (F) Gene ontology analysis of p300/CBP dependent genes that are independent of HDAC inhibition. GO ontology of significantly changing genes in clusters 2 and 4 from Figure 4e are shown.

Article Snippet: On the following day, the culture medium was replaced with treatment medium containing: 5 mM of individual SCFAs; 1 µM of A485, a p300/CBP inhibitor (MedChemExpress, HY-107455), a combination of 1 µM A485 and 5 mM butyrate; or 1 µM of Entinostat (ES), a class I HDAC inhibitor (MedChemExpress, HY-12163).

Techniques: Western Blot, Control, Gene Expression, Inhibition