Journal: iScience

Article Title: The m6A reader HNRNPC is a key regulator in DSS-induced colitis by modulating macrophage phenotype

doi: 10.1016/j.isci.2025.111812

Figure Lengend Snippet: HNRNPC with broad expression across diverse colonic cells downregulates in DSS-induced colitis Acute colitis was induced with 2.5% DSS (dextran sulfate sodium salt) in drinking water for 7 days and then with regular drinking water for another 3 days (A) Twenty-three major clusters of colonic cells from three healthy controls and three patients with ulcerative colitis. (B) Expression level of HNRNPC in 23 kinds of diverse colonic cells. (C) Disease activity index during acute colitis. (D) The body weight percentage (relative to the initial weight, set as 100%) during the progression of acute DSS-induced colitis. (E) Representative colonic images and colon length. (F) Representative hematoxylin and eosin (H&E) images of the colon. Scale bar: 200 μm (G) The relative mRNA expression of Tnf-α , Il1β , Claudin1 , and Zo1 . (H) The relative mRNA expression of Hnrnpc in the colon tissue. (I) The protein expression of HNRNPC in the colon tissues was determined by western blot. (J) The relative mRNA expression of Hnrnpc in the epithelial cells. Data are represented as mean ± SEM. (n = 6–7/group). Two-sided Student’s t test (E, G, H, I, and J) was performed. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. CON, control group with normal water; DSS, colitis group with dextran sulfate sodium water; Hnrnpc, heterogeneous nuclear ribonucleoprotein C; Tnfa , tumor necrosis factor α; Il1b , interleukin-1β; Zo1 , tight junction protein 1.

Article Snippet: Rabbit monoclonal anti- HNRNPC , Zen-bio , Cat# R381235; RRID: AB_ 3099728.

Techniques: Expressing, Activity Assay, Western Blot, Control

Journal: iScience

Article Title: The m6A reader HNRNPC is a key regulator in DSS-induced colitis by modulating macrophage phenotype

doi: 10.1016/j.isci.2025.111812

Figure Lengend Snippet: HNRNPC is upregulated in the macrophages of colonic lamina propria in the colitis (A) Gate strategy of flow cytometry for analyzing the expression of HNRNPC in macrophages. (B) Representative plots of ratio of the colonic CD45 + F4/80 + CD11b + macrophages in the CD45 + lymphocytes and (D) quantitative analysis. (C) Representative plots ratio of the colonic CD86 + and CD206 + macrophages and (E and F) quantitative analysis. (G) Representative histograms of ratio of HNRNPC + in the colonic macrophages and (H) quantitative analysis. (I) Representative histograms of ratio of HNRNPC + in the colonic CD86 + macrophages and (J) quantitative analysis. (K) Representative histograms of ratio of HNRNPC + in the colonic CD206 + macrophages and (L) quantitative analysis. (E and F) The cell number of HNRNPC+ macrophages, HNRNPC+ CD86 + macrophages, and HNRNPC+ CD206+ macrophages (The total cell number of the three types of macrophages is 500). Data are represented as mean ± SEM. (n = 6–7/group). Two-sided Student’s t test was performed. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. ns: not significant. FMO, Fluorescence Minus One; CON, control group with normal water; DSS, colitis group with dextran sulfate sodium water.

Article Snippet: Rabbit monoclonal anti- HNRNPC , Zen-bio , Cat# R381235; RRID: AB_ 3099728.

Techniques: Flow Cytometry, Expressing, Fluorescence, Control

Journal: iScience

Article Title: The m6A reader HNRNPC is a key regulator in DSS-induced colitis by modulating macrophage phenotype

doi: 10.1016/j.isci.2025.111812

Figure Lengend Snippet: HNRNPC knockdown relives LPS-induced cellular inflammation in vitro (A–E) The relative mRNA expression of Hnrnpc, Tnf-α , Il1β , Il18 , Inos , and Il6 after 2, 4, 6, 12, and 24 h treatment with LPS in the Raw264.7 cells. (F) The relative mRNA expression of Hnrnpc after transfected with siHNRNPC-1,2,3, or negative control for 24 h in the Raw264.7 cells. (G, H) The relative protein expression of HNRNPC after transfected with siHNRNPC-1,2,3 or negative control for 48 h in the Raw264.7 cells was determined by western blot. (I–M) The relative mRNA expression of Hnrnpc, Tnf-α , Il1β , Il18, Inos , and Il6 in the siNC + LPS and siHNRNPC + LPS groups. (After Raw264.7 cells are transfected with siRNA for 24h, LPS was added to treat the cells for another 2, 4, 6, 12, and 24 h). (N) The relative mRNA expression of HNRNPC after transfected with siHNRNPC or negative control for 24h in the THP1 cells. (O, P) The relative protein expression of HNRNPC after transfected with siHNRNPC or negative control for 48 h in the THP1 cells was determined by western blot. (Q–S) The relative mRNA expression of HNRNPC, TNF-α, IL1β, IL6, Il18, CCL2, CCL3, and CCL8 in the siNC + LPS and siHNRNPC + LPS groups. (After THP1 cells are transfected with siRNA for 24 h, 100 ng/mL or 500 ng/mL LPS was added to treat the cells for another 4 or 24 h). Data represent three independent experiments and are represented as mean ± SEM. ( n = 3/group). Two-sided Student’s t test (A–E and I–M) was performed. One-way ANOVA was applied in Figures 5 F and 5H. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. ns: not significant. Raw, Raw264.7 cells group without additional treatment; Raw+LPS, Raw264.7 cells after LPS treatment; siNC, Raw264.7, or THP1 cells with negative control siRNA treatment; siHNRNPC, Raw264.7, or THP1 cells with siHNRNPC treatment; siNC/siHNRNPC + LPS, Raw264.7, or THP1 cells with siRNA and LPS; LPS, lipopolysaccharide; Hnrnpc , heterogeneous nuclear ribonucleoprotein C; Tnf-α , tumor necrosis factor alpha; Il, interleukin; Inos , inducible nitric oxide synthase; CCL, C-C motif chemokine ligand 2.

Article Snippet: Rabbit monoclonal anti- HNRNPC , Zen-bio , Cat# R381235; RRID: AB_ 3099728.

Techniques: Knockdown, In Vitro, Expressing, Transfection, Negative Control, Western Blot

Journal: iScience

Article Title: The m6A reader HNRNPC is a key regulator in DSS-induced colitis by modulating macrophage phenotype

doi: 10.1016/j.isci.2025.111812

Figure Lengend Snippet: Downregulation of HNRNPC leads to alterations of inflammation, adhesion, and mRNA-splicing-related functions (A) A volcano plot was generated using RNA-seq data to compare HNRNPC knockeddown (KD) and wild-type (WT) Raw264.7 cells. The x axis displays the log2 fold changes (FC), and the y axis represents the −log10 p value for HNRNPC KD compared to HNRNPC WT. Downregulated genes are depicted as blue dots, and upregulated genes are shown as red dots. (B) GO enrichment analysis containing biological process, cellular component, and molecular function for HNRNPC knockdown. (C) GSEA dot plot containing activated and suppressed pathways. (D–F) Representative signaling pathways and their associated genes in the Raw264.7 cells with HNRNPC knockdown from GSEA analysis. (G–I) qPCR was used to verify the differentially expressed genes in the Raw264.7 cells with or without HNNRPC knockdown. ( n = 3/group). (J) The relative level of Itgb7 mRNA was examined by qPCR in Raw264.7 cells after HNRNPC knockdown and actinomycin D treatment (2 μg/mL) for 1, 2, and 4 h. The mRNAs at various time points were normalized to the t = 0 level (100%). (K) The relative mRNA level of Itgb7 in the siNC and siHNRNPC groups at various time points of actinomycin D treatment. Data are represented as mean ± SEM, and two-sided Student’s t test was performed. GSEA, gene set enrichment analysis; GO, Gene Ontology. siNC, Raw264.7 cells with negative control siRNA treatment; siHNRNPC, Raw264.7 cells with siHNRNPC treatment.

Article Snippet: Rabbit monoclonal anti- HNRNPC , Zen-bio , Cat# R381235; RRID: AB_ 3099728.

Techniques: Generated, RNA Sequencing, Knockdown, Protein-Protein interactions, Negative Control

Journal: iScience

Article Title: The m6A reader HNRNPC is a key regulator in DSS-induced colitis by modulating macrophage phenotype

doi: 10.1016/j.isci.2025.111812

Figure Lengend Snippet: Adoptive transfer of macrophages with HNRNPC knockdown alleviates DSS-induced colitis (A) Schematic diagram displaying the comprehensive design. Mice were treated with drinking water containing 2.5% DSS. On the day 0, 3, 5, and 7 of DSS-induced colitis, 2∗10ˆ6 cells were separately transferred to mice intravenously. (B) Disease activity index during acute colitis. (C) The body weight percentage (relative to the initial weight, set as 100%) during the progression of acute DSS-induced colitis. (D) Representative colonic images and colon length. (E) Representative hematoxylin and eosin (H&E) images of the colon. Scale bar: 100 μm (F) Gate strategy of flow cytometry for analyzing the expression of HNRNPC in macrophages in the colonic lamina propria. (G) Representative histograms of ratio of HNRNPC + in the colonic macrophages and (H) quantitative analysis. (I) The relative mRNA expression of anti-inflammatory cytokines including Il4 , Il13 , and Il10 . (J) The relative mRNA expression of pro-inflammatory cytokines including Il6 , Il18 , and Inos . (K) The relative mRNA expression of intestinal epithelial barrier makers including Zo1 , Occludin , and Claudin1 . Data are represented as mean ± SEM. (n = 5–6/group). Two-sided Student’s t test was performed. ∗ p < 0.05, ns: not significant. DSS+Raw-siNC, mice subjected to induction of colitis and the transfer of Raw264.7 cells transfected with negative control siRNA; DSS+Raw-siHN, mice subjected to induction of colitis and the transfer of Raw264.7 cells with HNRNPC knockdown; Il , interleukin; Inos , inducible nitric oxide synthase; Zo1 , tight junction protein 1.

Article Snippet: Rabbit monoclonal anti- HNRNPC , Zen-bio , Cat# R381235; RRID: AB_ 3099728.

Techniques: Adoptive Transfer Assay, Knockdown, Activity Assay, Flow Cytometry, Expressing, Transfection, Negative Control

Journal: iScience

Article Title: The m6A reader HNRNPC is a key regulator in DSS-induced colitis by modulating macrophage phenotype

doi: 10.1016/j.isci.2025.111812

Figure Lengend Snippet:

Article Snippet: Rabbit monoclonal anti- HNRNPC , Zen-bio , Cat# R381235; RRID: AB_ 3099728.

Techniques: Recombinant, Lysis, Saline, Modification, Bicinchoninic Acid Protein Assay, cDNA Synthesis, In Vitro, Sequencing, Software