Journal: Biomolecules

Article Title: Notch Signaling Exacerbates Pulmonary Fibrosis by Regulating the Differentiation of CD4 + Tissue-Resident Memory T Cells

doi: 10.3390/biom16020328

Figure Lengend Snippet: Enhanced pro-inflammatory and pro-fibrotic function of lung CD4 + T RM Cells in PF. ( A ) Gene set enrichment analysis (GSEA) plot of the upregulated signature of T cell activation and cytokine production in lung CD4 + T RM compared to CD4 + non-T RM (using scRNA-seq data derived from GSE122960 ). ( B – I ) Male C57BL/6J mice (6–8 weeks old) were randomized to receive a single intratracheal dose of BLM (2 mg/kg) or sterile PBS. On day 14, lungs were collected for single-cell suspension preparation and flow cytometry analysis after euthanasia by anesthetic overdose. n = 8 in BLM group, and n = 4 in PBS group. ( B , C ) IL-17A expression in CD4 + CD69 + CD103 + T RM cells, CD4 + CD69 + CD103 − T cells, and CD4 + CD69 − CD103 − T cells from mice with BLM-induced PF (BLM group) was measured by flow cytometry. ( D , E ) Granzyme B (GZMB) expression in CD4 + CD69 + CD103 + T RM cells, CD4 + CD69 + CD103 − T cells, and CD4 + CD69 − CD103 − T cells from mice with BLM-induced PF (BLM group) was measured by flow cytometry. ( F , G ) IL-17A expression in CD4 + CD69 + CD103 + T RM cells from mice with or without BLM-induced PF was measured by flow cytometry. ( H , I ) GZMB expression in CD4 + CD69 + CD103 + T RM cells from mice with or without BLM-induced PF was measured by flow cytometry. ( J ) Schematic of CD4 + T RM and fibroblast co-culture. CD4 + T cells isolated from mouse spleens were stimulated with anti-CD3 (5 µg/mL), anti-CD28 (2 µg/mL), with or without TGF-β (10 ng/mL). After 3 days, TGF-β-induced CD4 + T RM and CD4 + non-T RM cells were co-cultured for 24 h with BLM (200 ng/mL)-pre-stimulated mouse pulmonary fibroblasts. ( K – M ) Fibrosis marker expressions were detected by qPCR. n = 4 for each group. Data were expressed as mean ± SEM. One-way ANOVA in panel ( C , E , K , L , M ), and Student’s t test in panel ( G , I ). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. ns: not significant.

Article Snippet: CD4 + T RM cell differentiation was induced by culturing activated cells in complete medium with 10 ng/mL TGF-β (Sino Biological, Beijing, China, Cat# 80116-RNAH-5) over a 3-day period, following previously reported methods [ ].

Techniques: Activation Assay, Derivative Assay, Sterility, Single Cell, Suspension, Flow Cytometry, Expressing, Co-Culture Assay, Isolation, Cell Culture, Marker

Journal: Biomolecules

Article Title: Notch Signaling Exacerbates Pulmonary Fibrosis by Regulating the Differentiation of CD4 + Tissue-Resident Memory T Cells

doi: 10.3390/biom16020328

Figure Lengend Snippet: Activation of Notch signaling in CD4 + T RM cells from fibrotic lungs. ( A , B ) scRNA-seq analysis of lung tissue from donors and PF patients ( GSE122960 ). ( A ) GSEA revealed enrichment of the positive regulation of Notch signaling in CD4 + T RM cells vs. CD4 + non-T RM cells ( p -value determined by clusterProfiler GSEA). ( B ) GSEA revealed enrichment in the Notch related-signaling in PF CD4 + T RM cells vs. donor CD4 + T RM cells ( p -value determined by clusterProfiler GSEA). ( C – H ) CD4 + T cells from control and BLM-treated mouse splenocytes were stimulated for 3 days with anti-CD3 (5 µg/mL), anti-CD28 (2 µg/mL), and TGF-β (10 ng/mL). n = 4 for each group. ( C , D ) Representative flow cytometry plots ( C ) and quantitative statistics ( D ) of Notch1 mean fluorescence intensity (MFI) in CD4 + T RM cells differentiated in vitro from splenic CD4 + T cells of the two mouse groups. ( E , F ) Representative flow cytometry plots ( E ) and quantitative statistics (F) of Notch1 MFI in in vitro-differentiated CD4 + CD69 + CD103 + T RM , CD4 + CD69 − CD103 − , and CD4 + CD69 + CD103 − T cell subsets from control mouse splenic CD4 + T cells. ( G , H ) Representative flow cytometry plots ( G ) and quantitative statistics ( H ) of Notch1 MFI in in vitro-differentiated CD4 + CD69 + CD103 + T RM , CD4 + CD69 − CD103 − , and CD4 + CD69 + CD103 − T cell subsets from BLM-induced PF mouse splenic CD4 + T cells. ( I , J ) Representative flow cytometry plots (I) and quantitative statistics ( J ) of Notch1 MFI in CD4 + CD69 + CD103 + T RM , CD4 + CD69 − CD103 − , and CD4 + CD69 + CD103 − T cell subsets from BLM-treated mouse lung tissues. n = 9 for each group. Data were expressed as mean ± SEM. Student’s t test in panel ( C ), and One-way ANOVA in panel ( F , H , J ). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. The grey line in panels ( C , E , G , I ) indicates the blank control in flow cytometry.

Article Snippet: CD4 + T RM cell differentiation was induced by culturing activated cells in complete medium with 10 ng/mL TGF-β (Sino Biological, Beijing, China, Cat# 80116-RNAH-5) over a 3-day period, following previously reported methods [ ].

Techniques: Activation Assay, Control, Flow Cytometry, Fluorescence, In Vitro

Journal: Molecules

Article Title: Abietane-Type Diterpenoids from the Resin of Pinus yunnanensis and Their Potential Anti-Renal Fibrosis Activities

doi: 10.3390/molecules31040659

Figure Lengend Snippet: Anti-renal fibrosis activity of compounds 1 – 10 in NRK-52E and NRK-49F cells. ( A ) The cell viability of NRK-52E and NRK-49F cells after treatment with compounds 1 – 10 at 40 μM was measured by the CCK-8 assay ( n = 3). ( B , C ) Representative Western blot bands showing the effects of the compounds on the expression of α -SMA, fibronectin, and collagen I at 40 μM in NRK-52E and NRK-49F cells ( n = 3). GW788388 (GW) was used as the positive control. Column graphs represent the quantification of the corresponding western blot bands ( n = 3). Quantification analysis was performed by ImageJ (v1.51p) and GraphPad Prism (v9.5.0). Data are presented as mean ± SEM from three independent experiments. Statistical analysis: ## p < 0.01 versus Control (Ctrl) group; ** p < 0.01 versus TGF- β 1-treated group (TGF- β 1 group). ns: no significance.

Article Snippet: After incubation for 24 h, cells were serum-starved for 6 h and then incubated with recombinant human TGF- β 1 (10 ng/mL; HY- P70543 , MedChemExpress, San Jose, CA, USA) for 48 h with or without different concentrations of compounds.

Techniques: Activity Assay, CCK-8 Assay, Western Blot, Expressing, Positive Control, Control

Journal: Molecules

Article Title: Abietane-Type Diterpenoids from the Resin of Pinus yunnanensis and Their Potential Anti-Renal Fibrosis Activities

doi: 10.3390/molecules31040659

Figure Lengend Snippet: The effects of compounds 1 , 3 , 6 – 8 , and 10 on p-Smad2/3 in NRK-52E cells. Representative Western blot bands showing the effects of compounds 1 , 3 , 6 – 8 , and 10 on the expression of Smad2/3 and p-Smad2/3 at 40 μM in NRK-52E cells ( n = 3). GW was used as the positive control. Column graphs represent the quantitative analysis data of the corresponding Western blot bands ( n = 3). Quantification analysis was performed by ImageJ (v1.51p) and GraphPad Prism (v9.5.0). Data are presented as mean ± SEM from three independent experiments. Statistical analysis: ## p < 0.01 versus Ctrl group; ** p < 0.01 versus TGF- β 1 group. ns: no significance.

Article Snippet: After incubation for 24 h, cells were serum-starved for 6 h and then incubated with recombinant human TGF- β 1 (10 ng/mL; HY- P70543 , MedChemExpress, San Jose, CA, USA) for 48 h with or without different concentrations of compounds.

Techniques: Western Blot, Expressing, Positive Control

Journal: Frontiers in Pharmacology

Article Title: RP-182 alleviated obstruction-induced renal fibrosis by reprogramming CD206 + macrophages

doi: 10.3389/fphar.2026.1739457

Figure Lengend Snippet: RP-182 peptide inhibited the MMT process. (A) The schematic of the experimental design. (B) qPCR analysis for Col1a1 , Acta2 , and Fn expression in M0-BMDMs treated with PBS or TGF-β. N = 6. (C) The schematic of the experimental design. (D–F) qPCR analysis for Col1a1 , Acta2 , and Fn expression in M1-BMDMs treated with PBS, TGF-β, or TGF-β + RP-182 peptide. N = 6. (G) The schematic of the experimental design. (H–J) qPCR analysis for Col1a1 , Acta2 , and Fn expression in M2-BMDMs treated with PBS, TGF-β, or TGF-β + RP-182. N = 6. The results represent mean ± SEM. *p < 0.05, ***p < 0.001, NS no significance.

Article Snippet: To induce macrophage-to-myofibroblast transition (MMT), recombinant TGF-β (MCE, HY-P7118, 10 ng/mL) was used.

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: Latilactobacillus curvatus DCF0620 and postbiotics derived from soybean germ reduce colitis severity by modulating fibrosis and gut dysbiosis

doi: 10.3389/fimmu.2025.1726298

Figure Lengend Snippet: L. paracasei DCF0420, L. plantarum DCF0514, and L. curvatus DCF0620 improve inflammation and fibrosis in vitro . (A) Splenocytes from C57BL/6 mice (n = 4-6) were stimulated with each strain separately (at 0.1, 1, or 10 μg/mL) or vehicle (saline) for 2 h and then cultured with anti-CD3 Ab (2 μg/mL) or LPS (500 ng/mL) for 3 days. The levels of IFN-γ, IL-17 and IL-10 in the culture supernatant were determined via ELISA. (B) CCD-18Co cells were treated with each strain separately (at 10 μg/mL) or vehicle (saline) in the presence of TGF-β (10 ng/mL) for 24 h. The levels of fibronectin were assessed via immunoblotting. Values are presented as means ± SDs. Data are representative of two independent experiments.

Article Snippet: The sections were incubated with primary antibodies (Abs) against TNF-α (#ab6671; Abcam, Cambridge, UK), IL-1β (#ab9722; Abcam), IL-6 (#ab7737; Abcam), IL-17 (#ab79056; Abcam), transforming growth factor (TGF)-β (#BS-0086R; Bioss, Woburn, MA, USA), type I collagen (Col1) (#ab6308; Abcam), and α-smooth muscle actin (α-SMA) (#ab7817; Abcam) for 2 h at room temperature.

Techniques: In Vitro, Saline, Cell Culture, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Frontiers in Immunology

Article Title: Latilactobacillus curvatus DCF0620 and postbiotics derived from soybean germ reduce colitis severity by modulating fibrosis and gut dysbiosis

doi: 10.3389/fimmu.2025.1726298

Figure Lengend Snippet: L. paracasei DCF0420, L. plantarum DCF0514, and L. curvatus DCF0620 reduce colonic fibrosis in mice with colitis. Acute colitis was induced in C57BL/6 mice by oral administration of 2% DSS. Each strain was orally administered to the mice from 7 days before DSS injection until the end of the experiment (n = 4-5/group, normal control n = 3). On day 11, colon tissue sections were stained with Masson’s trichrome and Abs against TGF-β, Col1, and α-SMA. Representative images are shown (original magnification: 400×, Scale bar: 100 µm). Graphs display the numbers of Ab-positive cells per field. Data are expressed as mean ± SDs. Statistical analysis was performed using One-way ANOVA followed by Tukey’s test. *P < 0.05, ****P < 0.0001.

Article Snippet: The sections were incubated with primary antibodies (Abs) against TNF-α (#ab6671; Abcam, Cambridge, UK), IL-1β (#ab9722; Abcam), IL-6 (#ab7737; Abcam), IL-17 (#ab79056; Abcam), transforming growth factor (TGF)-β (#BS-0086R; Bioss, Woburn, MA, USA), type I collagen (Col1) (#ab6308; Abcam), and α-smooth muscle actin (α-SMA) (#ab7817; Abcam) for 2 h at room temperature.

Techniques: Injection, Control, Staining

Journal: Frontiers in Immunology

Article Title: Latilactobacillus curvatus DCF0620 and postbiotics derived from soybean germ reduce colitis severity by modulating fibrosis and gut dysbiosis

doi: 10.3389/fimmu.2025.1726298

Figure Lengend Snippet: Postbiotics from L. curvatus DCF0620 reduce colonic fibrosis and inflammation in mice with colitis. Acute colitis was induced in C57BL/6 mice by oral administration of 2% DSS. Then, 200 μL of postbiotics (100 mg/mL in PBS) was orally administered to the mice from 7 days before DSS injection until the end of the experiment (n = 4-5/group, normal control n = 3). (A) On day 12, colon tissue sections were stained with Abs against TGF-β, Col1, and α-SMA. (B) On day 12, the sections were stained with Abs against TNF-α, IL-1β, IL-6, and IL-17. Representative images are shown (original magnification: 400×, Scale bar: 100 µm). Graphs display the numbers of Ab-positive cells per field. Data are expressed as mean ± SDs. Statistical analysis was performed using One-way ANOVA followed by Tukey’s test. ****P < 0.0001.

Article Snippet: The sections were incubated with primary antibodies (Abs) against TNF-α (#ab6671; Abcam, Cambridge, UK), IL-1β (#ab9722; Abcam), IL-6 (#ab7737; Abcam), IL-17 (#ab79056; Abcam), transforming growth factor (TGF)-β (#BS-0086R; Bioss, Woburn, MA, USA), type I collagen (Col1) (#ab6308; Abcam), and α-smooth muscle actin (α-SMA) (#ab7817; Abcam) for 2 h at room temperature.

Techniques: Injection, Control, Staining

Journal: Cell Communication and Signaling : CCS

Article Title: LDOC1 connects histone H2B monoubiquitination to tumor cell plasticity in non-small cell lung cancer

doi: 10.1186/s12964-025-02607-z

Figure Lengend Snippet: LDOC1 loss alters TGF-β–driven EMT programs and motility through H2Bub1 dysregulation. a Confocal images of A549 sublines treated with TGF-β (10 ng/mL) for the indicated days and stained with FITC–anti-α-tubulin (DM1A) and DAPI. Scale bars, 20 μm. b Time-course qRT–PCR of EMT- and cytoskeleton-related genes in A549-shCtrl-1 and A549-shLDOC1-1 cells after TGF-β stimulation; expression was normalized to GAPDH (mean ± SEM, n = 3). c–e Transwell migration of A549-shCtrl-1/-2 and A549-shLDOC1-1/-2 cells (c) and H1299 cells transfected with vector (V) or LDOC1 ( d) , with quantitative summary in ( e ) (mean ± SEM, n = 3). f–h Matrigel invasion assays for the same A549 (f) and H1299 (g) panels, with quantification in (h) (mean ± SEM, n = 3). i Incucyte wound-healing assays showing wound confluence over time in A549-shCtrl and A549-shLDOC1 sublines with or without TGF-β; representative images at 0 h and 48 h are shown. j Immunoblot of H2Bub1 in A549-shLDOC1-1 cells expressing H2BK120R mutants (clones B10 and C9); Lamin B1, loading control; V, vector. k–m Matrigel invasion ( k ) with quantification ( l ) and wound-healing analysis (m) of A549-shLDOC1-1 cells expressing V or H2BK120R mutants (B10, C9) (mean ± SEM, n = 3). * P < 0.05, ** P < 0.01 (two-tailed t-test)

Article Snippet: Chemical reagents included recombinant human TGF-β (MCE, HY-P7118), the proteasome inhibitor bortezomib (MCE, HY-10227) and recombinant human EGF (MCE, HY-P7109).

Techniques: Staining, Quantitative RT-PCR, Expressing, Migration, Transfection, Plasmid Preparation, Western Blot, Clone Assay, Control, Two Tailed Test

Journal: Cell Communication and Signaling : CCS

Article Title: LDOC1 connects histone H2B monoubiquitination to tumor cell plasticity in non-small cell lung cancer

doi: 10.1186/s12964-025-02607-z

Figure Lengend Snippet: LDOC1 loss promotes a hybrid epithelial–mesenchymal state and elevated H2Bub1 associated with STAS and clinical outcome in NSCLC. a Immunoblot analysis of LDOC1 and EMT markers in A549-shCtrl-1/-2 and A549-shLDOC1-1/-2 cells; GAPDH and β-actin served as loading controls. b Confocal immunofluorescent images of E-cadherin (green) and nuclei (DAPI, blue) in A549 sublines. Scale bars, 10 μm. c–e Flow-cytometric analysis of A549-shCtrl-1 and A549-shLDOC1-1 cells stained for E-cadherin–APC and vimentin–PE: (c) representative single-parameter histograms, (d) bivariate plots showing E-cad⁺/Vim⁻, E-cad⁻/Vim⁺ and E-cad⁺/Vim⁺ populations, and (e) quantification of hybrid E/M (E-cad⁺/Vim⁺) cells (mean ± SEM; *P < 0.05, two-tailed t-test). f Adhesion of A549-shCtrl-1/-2 and A549-shLDOC1-1/-2 cells with or without TGF-β (mean ± SEM, n = 3; *P < 0.05, **P < 0.01, ***P < 0.001, two-tailed t-test). g–h Representative IHC for H&E, E-cadherin and vimentin (g) and H2Bub1 (h) in paired primary tumors and matched STAS lesions from NSCLC; insets show higher magnification. Scale bars, 100 μm. i Kaplan–Meier curves of progression-free survival in chemotherapy-treated NSCLC patients stratified by H2Bub1 expression. j LDOC1 mRNA levels in KRAS-mutant versus KRAS^WT tumors in the TCGA LUAD cohort (n = 520; Welch’s t-test)

Article Snippet: Chemical reagents included recombinant human TGF-β (MCE, HY-P7118), the proteasome inhibitor bortezomib (MCE, HY-10227) and recombinant human EGF (MCE, HY-P7109).

Techniques: Western Blot, Staining, Two Tailed Test, Expressing, Mutagenesis