Journal: Molecular Cancer

Article Title: Integrated multi-omics identifies a CD54 + iCAF-ITGAL + macrophage niche driving immunosuppression via CXCL8-PDL1 axis in cervical cancer

doi: 10.1186/s12943-025-02471-y

Figure Lengend Snippet: CD54⁺ iCAFs promote monocyte migration and M2-like polarization through CCL2 secretion. A Left: Transwell migration assay of THP-1 cells co-cultured with CD54⁺ iCAFs or normal fibroblasts (NFs) for 48 h. Right: Quantification of migrated cells (n = 3 independent experiments). Scale bar: 100 μm. B-C THP-1 cells co-cultured with CD54⁺ iCAFs show upregulated expression of M2-like macrophage biomarkers and cytokines. D CD54⁺ iCAFs were transfected with CD54-targeting siRNA (si-CD54) or negative control siRNA (si-NC). Volcano plot of differentially expressed genes (DEGs) identified by RNA-seq analysis between si-CD54 and si-NC groups. E CD54 + iCAFs were transfected with si-CD54 or si-NC. Then, CD54 and CCL2 protein levels were assessed by Western blot. Left: Representative images. Right: Quantification from n = 3 independent experiments. F ELISA quantification of CCL2 secretion from CD54⁺ iCAFs and NFs. G Left: THP-1 cell migration in response to recombinant CCL2 treatment (50 ng/ml, 48 h) or PBS (Control group). Right: Quantification of migrated cells. All quantitative data are presented as mean ± SD. Statistical significance was determined using Student’s t-test for panels A (right), B, C, E (right), F, and G (right). *: P < 0.05, **: P < 0.01, ***: P < 0.001; ns, not significant

Article Snippet: A neutralizing anti-human CCL2 antibody (MedChemExpress) or an IgG isotype control was introduced to the culture medium.

Techniques: Migration, Transwell Migration Assay, Cell Culture, Expressing, Transfection, Negative Control, RNA Sequencing, Western Blot, Enzyme-linked Immunosorbent Assay, Recombinant, Control

Journal: Molecular Cancer

Article Title: Integrated multi-omics identifies a CD54 + iCAF-ITGAL + macrophage niche driving immunosuppression via CXCL8-PDL1 axis in cervical cancer

doi: 10.1186/s12943-025-02471-y

Figure Lengend Snippet: CD54⁺ iCAF-driven macrophage reprogramming promotes CXCL8 expression and tumor progression in vivo. A THP-1-derived macrophages, transfected with control siRNA (Control group) or ITGAL-targeting siRNA (Treatment group), were co-cultured with CD54⁺ iCAFs for 48 h and subsequently subjected to RNA-seq analysis. Volcano plot shows differentially expressed genes (DEGs) between control and treatment groups. B Left: Representative flow cytometry plots of CD54⁺ iCAF abundance. Middle: Quantitative analysis of CXCL8⁺ cell frequencies in CD54⁺ iCAF-high tumors (n = 3). Right: quantitative analysis of CXCL8 expression in CD54 + iCAF-high versus CD54 + iCAF-low (Displaying in Supplementary Fig. 5F) tumors. C Flow cytometric quantification of CXCL8-producing immune cell subsets in cervical cancer tissues. D Multiplex immunohistochemistry images showing macrophage-specific CXCL8 expression in cervical cancer tissue (Scale bar: 100 μm). E NIH/3T3 fibroblasts transfected with CD54 overexpression plasmid (OE-CD54) or empty vector control (OE-NC). CD54 and CCL2 secretion levels were quantified by ELISA. F Representative tumor images from C57BL/6 mice co-injected with TC-1 cells and NIH/3T3 fibroblasts expressing CD54 or empty vector (Control). G Tumor growth curves measured every 3 days (n = 3 mice/group). H-I Flow cytometry analysis of CD206⁺ macrophage infiltration in tumors from (F). I (Right): Quantification of CD206⁺ macrophage proportions. J Serum MIP-2 levels measured by ELISA in experimental groups from (F). All quantitative data are presented as mean ± SD. Statistical significance was determined using Student’s t-test for panels B (right), E, G(right), I (right), and J. *: P < 0.05, **: P < 0.001, ***: P <0.001

Article Snippet: A neutralizing anti-human CCL2 antibody (MedChemExpress) or an IgG isotype control was introduced to the culture medium.

Techniques: Expressing, In Vivo, Derivative Assay, Transfection, Control, Cell Culture, RNA Sequencing, Flow Cytometry, Multiplex Assay, Immunohistochemistry, Over Expression, Plasmid Preparation, Enzyme-linked Immunosorbent Assay, Injection

Journal: Molecular Cancer

Article Title: Integrated multi-omics identifies a CD54 + iCAF-ITGAL + macrophage niche driving immunosuppression via CXCL8-PDL1 axis in cervical cancer

doi: 10.1186/s12943-025-02471-y

Figure Lengend Snippet: CXCL8 correlates with CD8⁺ T cell exclusion and promotes PD-L1 expression on macrophages through cell-contact and soluble-factor dependent mechanisms. A CIBERSORT analysis showing an inverse correlation between CXCL8 mRNA levels and CD8⁺ T cell infiltration. B Representative immunohistochemistry (IHC) images showing CD8⁺ T cell density in high- versus low-CXCL8 expressing tumors (Scale bar: 100 μm). C Negative correlation between protein levels of CXCL8 and PD-L1 based on IHC scoring (Pearson correlation). D Left: Flow cytometry plots of PD-L1⁺ cells. Right: Quantification of PD-L1 expression in high- versus low-CXCL8 tumors (n = 3 per group). E Representative IHC staining confirming macrophage-specific PD-L1 expression (Scale bar: 100 μm). F Left: Gating strategy for identifying PD-L1⁺ cells. Right: Quantification of PD-L1 expression across immune cell subtypes, showing macrophage dominance (n = 6). G Left: Flow cytometry profiles of PD-L1 expression. Right: PD-L1 levels in high- versus low-CXCL8 tumors. H Flow cytometry analysis of CXCL8 and PD-L1 expression on CD68+ macrophages co-cultured with CD54⁺ iCAFs under direct contact or Transwell conditions, with normal fibroblasts (NFs) and macrophage-only cultures as controls. The bar graph shows geometric mean fluorescence intensity from three independent experiments. Corresponding representative flow cytometry plots are shown in Supplementary Fig. 6D. I CXCL8 and PD-L1 expression on macrophages after co-culture with CD54⁺ iCAFs and treatment with IgG control, anti-CD54, or anti-ITGAL blocking antibodies. See Supplementary Fig. 6E for flow plots. J CXCL8 and PD-L1 expression on macrophages transfected with control siRNA (si-NC) or ITGAL-targeting siRNA (si-ITGAL), with or without CD54 + iCAF co-culture. Corresponding representative flow cytometry plots are shown in Supplementary Fig. 6F. K CXCL8 and PD-L1 expression on macrophages co-cultured with CD54⁺ iCAFs and treated with IgG control or anti-CCL2 neutralizing antibody. Representative flow plots are provided in Supplementary Fig. 6G. Data are presented as mean ± SD. Statistical tests used: two-tailed Student’s t-test (D, right; G, right; K); one-way ANOVA with Tukey's multiple comparisons test (F, right; H, I, J). **: P < 0.01, ***: P < 0.001; ns, not significant

Article Snippet: A neutralizing anti-human CCL2 antibody (MedChemExpress) or an IgG isotype control was introduced to the culture medium.

Techniques: Expressing, Immunohistochemistry, Flow Cytometry, Cell Culture, Fluorescence, Co-Culture Assay, Control, Blocking Assay, Transfection, Two Tailed Test