Journal: Cell death & disease

Article Title: FGFR4 promotes CAF activation through the CXCL10-CXCR3 axis in colon cancer.

doi: 10.1038/s41419-025-07588-y

Figure Lengend Snippet: Fig. 3 Transcriptome analysis of FGFR4-overexpressing colon cancer cells. A Principal Component Analysis (PCA) plot showing distinct separation of EV and FGFR4 overexpression CT-26 groups (n = 3/group). B Volcano plot of DEGs in FGFR4-overexpressing CT-26 cells compared with those in EV controls. DEGs were defined as genes with a P < 0.001 and an absolute value of >2 for the log2 of fold-change in FGFR4-overexpressing cells compared with that in controls. Green and red dots show significantly downregulated and upregulated DEGs, respectively. C Reactome analysis for the 96 significantly upregulated genes. D Enrichment plots for the top ten gene sets enriched in Gene Set Enrichment Analysis (GSEA) Hallmark analysis. These plots show the running enrichment score (ES) profile and the distribution of gene set members along the rank-ordered list of genes. In each graph, the probes on the far left (red) represent genes most correlated with upregulation, while those on the far right (blue) indicate genes most correlated with downregulation in the FGFR4-overexpressing group. The green line represents the running ES across the gene list. The normalized enrichment score (NES) and false discovery rate (FDR) are provided for each gene set. E Changes in expression of IFN genes based on RNA-seq data. F RT-qPCR analysis confirmed that FGFR4 overexpression led to increased expression of Ifnβ and Cxcl10 in CT-26 and HT-29 cells. Gapdh was used as an internal reference control gene. G ELISA analysis of secretory Ifnβ and Cxcl10 levels in conditioned media of CT-26/EV and CT-26/FGFR4 cells. Data are presented as mean ± SD from three independent experiments. Statistical significance: ***P < 0.001, ****P < 0.0001 using Student’s t-test. DEGs, differentially expressed genes.

Article Snippet: Mouse CXCL10 (R&D Systems, Minneapolis, MN, USA; 466-CR) and mouse TGFβ (R&D Systems; 7666-MB) were used as recombinant proteins.

Techniques: Over Expression, Expressing, RNA Sequencing, Quantitative RT-PCR, Control, Enzyme-linked Immunosorbent Assay

Journal: Cell death & disease

Article Title: FGFR4 promotes CAF activation through the CXCL10-CXCR3 axis in colon cancer.

doi: 10.1038/s41419-025-07588-y

Figure Lengend Snippet: Fig. 4 FGFR4 induces CXCL10 secretion via activation of the TLR3-TBK-IRF axis and autocrine action of IFNβ. A Left: FGFR4-mediated activation of the TLR3-TBK-IRF and IFN signaling pathways was determined using western blotting for the indicated protein markers in CT-26 and HT-29 cells. Phosphorylated proteins are denoted by “P” before the protein name. Right: Schematic representation of the TLR3-TBK-IRF and IFN-STAT-CXCL10 signaling pathway. B The TLR3-TBK-IRF and IFN-STAT-CXCL10 signaling pathways are suppressed following FGFR4 knockdown or pharmacological inhibition by BLU9931 in FGFR4-overexpressing CT-26 and HT-29 cells. Cells were transfected with siRNA targeting FGFR4 for 48 h or treated with 1 μM BLU-9931 for 24 h. Western blot analysis shows a reduction in TBK1, IRF, and STAT1 phosphorylation, and the downstream targets IFN-β and CXCL10 following FGFR4 knockdown or BLU-9931 treatment. β-actin was used as a loading control. C Effect of siRNA-mediated FGFR4 knockdown on the gene expression levels of Ifnα, Ifnβ, and Cxcl10 induced by FGFR4 overexpression in CT-26 colon cancer cells. D Effect of pharmacological inhibition of FGFR4 activity by BLU9931 on the expression levels of Ifnα, Ifnβ, and Cxcl10 induced by FGFR4 overexpression in CT-26 colon cancer cells. Data are presented as mean ± SD from three independent experiments. Statistical significance: ns, not significant; **P < 0.01; ***P < 0.001; ****P < 0.0001 using one-way ANOVA. Veh vehicle, BLU BLU9931, siNC siRNA negative control, siF4 siFgfr4.

Article Snippet: Mouse CXCL10 (R&D Systems, Minneapolis, MN, USA; 466-CR) and mouse TGFβ (R&D Systems; 7666-MB) were used as recombinant proteins.

Techniques: Activation Assay, Protein-Protein interactions, Western Blot, Knockdown, Inhibition, Transfection, Phospho-proteomics, Control, Gene Expression, Over Expression, Activity Assay, Expressing, Negative Control

Journal: Cell death & disease

Article Title: FGFR4 promotes CAF activation through the CXCL10-CXCR3 axis in colon cancer.

doi: 10.1038/s41419-025-07588-y

Figure Lengend Snippet: Fig. 5 FGFR4 induces CAF differentiation and activation via the CXCL10-CXCR3 axis. A Collagen gel contraction assay results show the effect of recombinant CXCL10 protein on the contractility of NIH/3T3 cells (n = 6/group). Representative gel images are shown together with the quantification of collagen gel contraction. B Migration and invasion assay results show the effect of recombinant CXCL10 protein on the migration and invasion properties of NIH/3T3 cells. The effect of CXCL10 was compared to that of TGFβ, which induces CAF activation. C The effect of recombinant CXCL10 protein (ng) on CAF marker gene levels in NIH/3T3 cells was determined using western blotting and RT-qPCR analyses. D Effect of Cxcl10 knockdown on CAF marker gene expression. NIH/3T3 cells were incubated for 24 h with conditioned media (CM) obtained from siNC- or siCXCL10-transfected CT-26/FGFR4 cells. Western blotting and RT-qPCR analyses of CAF marker expression in NIH/3T3 cells. E Effect of CXCL10 neutralizing antibody on CAF marker gene expression. NIH/3T3 cells were incubated in CM obtained from CT-26/ FGFR4 cells in the presence of either normal anti-IgG or anti-CXCL10 neutralizing antibodies (concentrations shown in μg/mL) for 24 h. F Effect of AMG487, a pharmacological CXCR3 inhibitor, on gene expression of CAF markers. The CM of CT-26/FGFR4 cells, alone or in combination with the CXCR3 inhibitor, was used to treat the NIH/3T3 cells for 24 h. G Invasion assay results depicting the effect of the CXCR3 inhibitor AMG487 (2 μM) on the invasion capabilities of CT-26 primary tumor-derived CAFs. H Invasion assay results show the effects of various CXCR3 inhibitors on the invasive properties of CT26/FGFR4 tumor-derived CAFs. The CXCR3 inhibitors used (at 2 μM) were: TAK779, NBI74330, and SCH546738. Data are presented as mean ± SD. Statistical significance: *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001 using Student’s t-test and one-way ANOVA). NT no treatment, AMG AMG487, TAK TAK779, NBI NBI74330, SCH SCH54673.

Article Snippet: Mouse CXCL10 (R&D Systems, Minneapolis, MN, USA; 466-CR) and mouse TGFβ (R&D Systems; 7666-MB) were used as recombinant proteins.

Techniques: Activation Assay, Collagen Gel Contraction Assay, Recombinant, Migration, Invasion Assay, Marker, Western Blot, Quantitative RT-PCR, Knockdown, Gene Expression, Incubation, Transfection, Expressing, Derivative Assay

Journal: Cell death & disease

Article Title: FGFR4 promotes CAF activation through the CXCL10-CXCR3 axis in colon cancer.

doi: 10.1038/s41419-025-07588-y

Figure Lengend Snippet: Fig. 6 Correlation analysis of CAF markers and FGFR4 or CXCL10 expression in tissue samples from colorectal cancer patients. A FGFR4, CXCL10, and CAF marker mRNA expression in tissue samples from colorectal cancer patients (n = 137) and adjacent normal tissues (n = 137) were analyzed using RT-qPCR. Data presented as the mean ± SD (n = 137 independent samples; statistical analysis via Student’s t-test; ***P < 0.001, ****P < 0.0001). B Correlation analysis between the expression of FGFR4 and that of CAF markers, such as CXCL10, αSMA, PDGFRβ, and FAP in tissue samples from colorectal cancer patients (n = 137) was performed using Pearson’s correlation test. C Correlation analysis between CXCL10 and CAF marker expression in tissue samples from colorectal cancer patients (n = 137), was examined using Pearson’s correlation test. Statistical significance was assessed using Pearson’s correlation coefficient (r) and Student’s t-test (P-value).

Article Snippet: Mouse CXCL10 (R&D Systems, Minneapolis, MN, USA; 466-CR) and mouse TGFβ (R&D Systems; 7666-MB) were used as recombinant proteins.

Techniques: Expressing, Marker, Quantitative RT-PCR

Journal: Frontiers in Immunology

Article Title: FMNL1 and mDia1 promote efficient T cell migration through complex environments via distinct mechanisms

doi: 10.3389/fimmu.2024.1467415

Figure Lengend Snippet: FMNL1 mediates T cell migration independently of Myosin-II. WT, FMNL1 KO, and mDia1 KO T cells were collected from donor mice, ex vivo activated, and differentially fluorescently labeled. (A) T cells were seeded onto 3μm transwell inserts with 50μM Blebbistatin or a vehicle control and 100 ng/mL CXCL10 in the bottom well. Percent migration is calculated as the number of cells counted in the bottom well normalized to a 20% loading control well. (B–F) T cells were embedded in 1.0 mg/mL collagen with increasing concentrations of para-nitro-Blebbistatin. (B) Representative maximum Z-projection images of WT (red) FMNL1 KO (green), and mDia1 KO (cyan) T cell migration through 1.0 mg/mL collagen matrices treated with either DMSO (top) or 10 μM para-nitro-Blebbistatin (bottom). Track lines show the path taken by the cells over 25 min. (C) Trajectory plots of the DMSO and 10 μM Blebbistatin-treated groups for all cells analyzed in (D–F) . (D–F) Quantification of mean track speed, average arrest coefficient, and mean squared displacement of cells tracked continuously for 10 min. Cells were treated with a DMSO control, 3 μM or 10 μM para-nitro-Blebbistatin. Data in (A) represents the mean +/- SEM of three independent experiments. Significance was determined by One-way ANOVA. Data in (D–F) represent the mean +/- SEM of at least 165 cells per condition pooled from three independent experiments. Significance was determined by Brown-Forsythe and Welch ANOVA tests with Games-Howell’s multiple comparisons test. ns, not significant, * = p<0.05, ** = p<0.01, *** = p<0.001, **** = p<0.0001.

Article Snippet: Recombinant Mouse IP-10 (CXCL10) , Peprotech , 250-16.

Techniques: Migration, Ex Vivo, Labeling, Control

Journal: Frontiers in Immunology

Article Title: FMNL1 and mDia1 promote efficient T cell migration through complex environments via distinct mechanisms

doi: 10.3389/fimmu.2024.1467415

Figure Lengend Snippet: Key resources.

Article Snippet: Recombinant Mouse IP-10 (CXCL10) , Peprotech , 250-16.

Techniques: Recombinant, Fractionation, Cell Culture