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Tissue specification of neutrophils in the CRLM microenvironment. A, Expression levels of CXCR4, CXCR2, and CXCR1 across each TAN and PMN subset. B, mIHC staining of MPO, CXCR1, CXCR2, and CXCR4 in colorectal cancer and CRLM tissues. Scale bar, 50µm. C, Quantitative analysis of the percentage of CXCR1 + , CXCR2 + , and CXCR4 + neutrophils in paired colorectal cancer and CRLM tissues ( n = 5). D, Heatmap showing the mean strength of chemokine–receptor axis for each TAN subset in CRLM tissues. E, Predicted ordering of each TAN subset by CytoTRACE analysis using scRNA-seq dataset. F, Monocle 2 trajectory analysis of TAN subsets differentiation using the scRNA-seq dataset. Cells are colored by pseudotime, ordering predicted by CytoTRACE, and cell types. G, The density of each TAN subset along pseudotime. H, Clustering analysis of gene expression along pseudotime, with pathway enrichment analysis of four gene sets based on pseudotime. I, TAN1 similarity scores of TAN0_ELL2, TAN2_HSPA1B, TAN3_IFIT2, and TAN4_ARG1 across colorectal cancer, adjacent liver tissues (LP), and CRLM tissues. The TAN1 similarity score was calculated based on the top 50 differentially expressed genes in TAN1. J, Relative mRNA expression levels of BHLHE40 , CDKN1A (p21), LGALS3 , VEGFA , and LDHA as determined by qPCR in control neutrophils or neutrophils cocultured with <t>HCT116</t> or DLD-1 cells. K and L, Flow cytometry analysis of Gal3 surface expression in neutrophils cultured alone (control) or cocultured with DLD-1 ( K ) or HCT116 cells ( L ). P values were determined by paired Student t test ( C ) or unpaired Student t test ( J , K , and L ). n.s., not significant; **, P < 0.01; ***, P < 0.001. CRC, colorectal cancer; Ctrl, control; MFI, mean fluorescence intensity.
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Tissue specification of neutrophils in the CRLM microenvironment. A, Expression levels of CXCR4, CXCR2, and CXCR1 across each TAN and PMN subset. B, mIHC staining of MPO, CXCR1, CXCR2, and CXCR4 in colorectal cancer and CRLM tissues. Scale bar, 50µm. C, Quantitative analysis of the percentage of CXCR1 + , CXCR2 + , and CXCR4 + neutrophils in paired colorectal cancer and CRLM tissues ( n = 5). D, Heatmap showing the mean strength of chemokine–receptor axis for each TAN subset in CRLM tissues. E, Predicted ordering of each TAN subset by CytoTRACE analysis using scRNA-seq dataset. F, Monocle 2 trajectory analysis of TAN subsets differentiation using the scRNA-seq dataset. Cells are colored by pseudotime, ordering predicted by CytoTRACE, and cell types. G, The density of each TAN subset along pseudotime. H, Clustering analysis of gene expression along pseudotime, with pathway enrichment analysis of four gene sets based on pseudotime. I, TAN1 similarity scores of TAN0_ELL2, TAN2_HSPA1B, TAN3_IFIT2, and TAN4_ARG1 across colorectal cancer, adjacent liver tissues (LP), and CRLM tissues. The TAN1 similarity score was calculated based on the top 50 differentially expressed genes in TAN1. J, Relative mRNA expression levels of BHLHE40 , CDKN1A (p21), LGALS3 , VEGFA , and LDHA as determined by qPCR in control neutrophils or neutrophils cocultured with <t>HCT116</t> or DLD-1 cells. K and L, Flow cytometry analysis of Gal3 surface expression in neutrophils cultured alone (control) or cocultured with DLD-1 ( K ) or HCT116 cells ( L ). P values were determined by paired Student t test ( C ) or unpaired Student t test ( J , K , and L ). n.s., not significant; **, P < 0.01; ***, P < 0.001. CRC, colorectal cancer; Ctrl, control; MFI, mean fluorescence intensity.
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Tissue specification of neutrophils in the CRLM microenvironment. A, Expression levels of CXCR4, CXCR2, and CXCR1 across each TAN and PMN subset. B, mIHC staining of MPO, CXCR1, CXCR2, and CXCR4 in colorectal cancer and CRLM tissues. Scale bar, 50µm. C, Quantitative analysis of the percentage of CXCR1 + , CXCR2 + , and CXCR4 + neutrophils in paired colorectal cancer and CRLM tissues ( n = 5). D, Heatmap showing the mean strength of chemokine–receptor axis for each TAN subset in CRLM tissues. E, Predicted ordering of each TAN subset by CytoTRACE analysis using scRNA-seq dataset. F, Monocle 2 trajectory analysis of TAN subsets differentiation using the scRNA-seq dataset. Cells are colored by pseudotime, ordering predicted by CytoTRACE, and cell types. G, The density of each TAN subset along pseudotime. H, Clustering analysis of gene expression along pseudotime, with pathway enrichment analysis of four gene sets based on pseudotime. I, TAN1 similarity scores of TAN0_ELL2, TAN2_HSPA1B, TAN3_IFIT2, and TAN4_ARG1 across colorectal cancer, adjacent liver tissues (LP), and CRLM tissues. The TAN1 similarity score was calculated based on the top 50 differentially expressed genes in TAN1. J, Relative mRNA expression levels of BHLHE40 , CDKN1A (p21), LGALS3 , VEGFA , and LDHA as determined by qPCR in control neutrophils or neutrophils cocultured with <t>HCT116</t> or DLD-1 cells. K and L, Flow cytometry analysis of Gal3 surface expression in neutrophils cultured alone (control) or cocultured with DLD-1 ( K ) or HCT116 cells ( L ). P values were determined by paired Student t test ( C ) or unpaired Student t test ( J , K , and L ). n.s., not significant; **, P < 0.01; ***, P < 0.001. CRC, colorectal cancer; Ctrl, control; MFI, mean fluorescence intensity.
Hct116 Human Colorectal Cancer Cells, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Tissue specification of neutrophils in the CRLM microenvironment. A, Expression levels of CXCR4, CXCR2, and CXCR1 across each TAN and PMN subset. B, mIHC staining of MPO, CXCR1, CXCR2, and CXCR4 in colorectal cancer and CRLM tissues. Scale bar, 50µm. C, Quantitative analysis of the percentage of CXCR1 + , CXCR2 + , and CXCR4 + neutrophils in paired colorectal cancer and CRLM tissues ( n = 5). D, Heatmap showing the mean strength of chemokine–receptor axis for each TAN subset in CRLM tissues. E, Predicted ordering of each TAN subset by CytoTRACE analysis using scRNA-seq dataset. F, Monocle 2 trajectory analysis of TAN subsets differentiation using the scRNA-seq dataset. Cells are colored by pseudotime, ordering predicted by CytoTRACE, and cell types. G, The density of each TAN subset along pseudotime. H, Clustering analysis of gene expression along pseudotime, with pathway enrichment analysis of four gene sets based on pseudotime. I, TAN1 similarity scores of TAN0_ELL2, TAN2_HSPA1B, TAN3_IFIT2, and TAN4_ARG1 across colorectal cancer, adjacent liver tissues (LP), and CRLM tissues. The TAN1 similarity score was calculated based on the top 50 differentially expressed genes in TAN1. J, Relative mRNA expression levels of BHLHE40 , CDKN1A (p21), LGALS3 , VEGFA , and LDHA as determined by qPCR in control neutrophils or neutrophils cocultured with <t>HCT116</t> or DLD-1 cells. K and L, Flow cytometry analysis of Gal3 surface expression in neutrophils cultured alone (control) or cocultured with DLD-1 ( K ) or HCT116 cells ( L ). P values were determined by paired Student t test ( C ) or unpaired Student t test ( J , K , and L ). n.s., not significant; **, P < 0.01; ***, P < 0.001. CRC, colorectal cancer; Ctrl, control; MFI, mean fluorescence intensity.
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Tissue specification of neutrophils in the CRLM microenvironment. A, Expression levels of CXCR4, CXCR2, and CXCR1 across each TAN and PMN subset. B, mIHC staining of MPO, CXCR1, CXCR2, and CXCR4 in colorectal cancer and CRLM tissues. Scale bar, 50µm. C, Quantitative analysis of the percentage of CXCR1 + , CXCR2 + , and CXCR4 + neutrophils in paired colorectal cancer and CRLM tissues ( n = 5). D, Heatmap showing the mean strength of chemokine–receptor axis for each TAN subset in CRLM tissues. E, Predicted ordering of each TAN subset by CytoTRACE analysis using scRNA-seq dataset. F, Monocle 2 trajectory analysis of TAN subsets differentiation using the scRNA-seq dataset. Cells are colored by pseudotime, ordering predicted by CytoTRACE, and cell types. G, The density of each TAN subset along pseudotime. H, Clustering analysis of gene expression along pseudotime, with pathway enrichment analysis of four gene sets based on pseudotime. I, TAN1 similarity scores of TAN0_ELL2, TAN2_HSPA1B, TAN3_IFIT2, and TAN4_ARG1 across colorectal cancer, adjacent liver tissues (LP), and CRLM tissues. The TAN1 similarity score was calculated based on the top 50 differentially expressed genes in TAN1. J, Relative mRNA expression levels of BHLHE40 , CDKN1A (p21), LGALS3 , VEGFA , and LDHA as determined by qPCR in control neutrophils or neutrophils cocultured with <t>HCT116</t> or DLD-1 cells. K and L, Flow cytometry analysis of Gal3 surface expression in neutrophils cultured alone (control) or cocultured with DLD-1 ( K ) or HCT116 cells ( L ). P values were determined by paired Student t test ( C ) or unpaired Student t test ( J , K , and L ). n.s., not significant; **, P < 0.01; ***, P < 0.001. CRC, colorectal cancer; Ctrl, control; MFI, mean fluorescence intensity.
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Tissue specification of neutrophils in the CRLM microenvironment. A, Expression levels of CXCR4, CXCR2, and CXCR1 across each TAN and PMN subset. B, mIHC staining of MPO, CXCR1, CXCR2, and CXCR4 in colorectal cancer and CRLM tissues. Scale bar, 50µm. C, Quantitative analysis of the percentage of CXCR1 + , CXCR2 + , and CXCR4 + neutrophils in paired colorectal cancer and CRLM tissues ( n = 5). D, Heatmap showing the mean strength of chemokine–receptor axis for each TAN subset in CRLM tissues. E, Predicted ordering of each TAN subset by CytoTRACE analysis using scRNA-seq dataset. F, Monocle 2 trajectory analysis of TAN subsets differentiation using the scRNA-seq dataset. Cells are colored by pseudotime, ordering predicted by CytoTRACE, and cell types. G, The density of each TAN subset along pseudotime. H, Clustering analysis of gene expression along pseudotime, with pathway enrichment analysis of four gene sets based on pseudotime. I, TAN1 similarity scores of TAN0_ELL2, TAN2_HSPA1B, TAN3_IFIT2, and TAN4_ARG1 across colorectal cancer, adjacent liver tissues (LP), and CRLM tissues. The TAN1 similarity score was calculated based on the top 50 differentially expressed genes in TAN1. J, Relative mRNA expression levels of BHLHE40 , CDKN1A (p21), LGALS3 , VEGFA , and LDHA as determined by qPCR in control neutrophils or neutrophils cocultured with HCT116 or DLD-1 cells. K and L, Flow cytometry analysis of Gal3 surface expression in neutrophils cultured alone (control) or cocultured with DLD-1 ( K ) or HCT116 cells ( L ). P values were determined by paired Student t test ( C ) or unpaired Student t test ( J , K , and L ). n.s., not significant; **, P < 0.01; ***, P < 0.001. CRC, colorectal cancer; Ctrl, control; MFI, mean fluorescence intensity.

Journal: Cancer Discovery

Article Title: Senescent-Like Neutrophils Shape Angiogenic Immunosuppressive Niches in Colorectal Cancer Liver Metastasis

doi: 10.1158/2159-8290.CD-25-0820

Figure Lengend Snippet: Tissue specification of neutrophils in the CRLM microenvironment. A, Expression levels of CXCR4, CXCR2, and CXCR1 across each TAN and PMN subset. B, mIHC staining of MPO, CXCR1, CXCR2, and CXCR4 in colorectal cancer and CRLM tissues. Scale bar, 50µm. C, Quantitative analysis of the percentage of CXCR1 + , CXCR2 + , and CXCR4 + neutrophils in paired colorectal cancer and CRLM tissues ( n = 5). D, Heatmap showing the mean strength of chemokine–receptor axis for each TAN subset in CRLM tissues. E, Predicted ordering of each TAN subset by CytoTRACE analysis using scRNA-seq dataset. F, Monocle 2 trajectory analysis of TAN subsets differentiation using the scRNA-seq dataset. Cells are colored by pseudotime, ordering predicted by CytoTRACE, and cell types. G, The density of each TAN subset along pseudotime. H, Clustering analysis of gene expression along pseudotime, with pathway enrichment analysis of four gene sets based on pseudotime. I, TAN1 similarity scores of TAN0_ELL2, TAN2_HSPA1B, TAN3_IFIT2, and TAN4_ARG1 across colorectal cancer, adjacent liver tissues (LP), and CRLM tissues. The TAN1 similarity score was calculated based on the top 50 differentially expressed genes in TAN1. J, Relative mRNA expression levels of BHLHE40 , CDKN1A (p21), LGALS3 , VEGFA , and LDHA as determined by qPCR in control neutrophils or neutrophils cocultured with HCT116 or DLD-1 cells. K and L, Flow cytometry analysis of Gal3 surface expression in neutrophils cultured alone (control) or cocultured with DLD-1 ( K ) or HCT116 cells ( L ). P values were determined by paired Student t test ( C ) or unpaired Student t test ( J , K , and L ). n.s., not significant; **, P < 0.01; ***, P < 0.001. CRC, colorectal cancer; Ctrl, control; MFI, mean fluorescence intensity.

Article Snippet: The human colon cancer cell lines HCT116 (RRID: CVCL_0291) and DLD-1 (RRID: CVCL_0248), mouse colon cancer cell line CT26 (RRID: CVCL_7254), as well as the human neutrophil cell line HL-60 (RRID: CVCL_0002) were obtained from the American Type Culture Collection.

Techniques: Tissue, Expressing, Staining, Analysis, Gene Expression, Control, Flow Cytometry, Cell Culture, Fluorescence