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mouse anti human il6  (R&D Systems)


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    Structured Review

    R&D Systems mouse anti human il6
    Metastatic osteosarcoma cells promote osteosarcoma chemotaxis via the <t>IL6/CXCL8</t> mechanism. A, Schematic for testing migration to IL6 and CXCL8 in a Transwell setup and quantification of migrated cells relative to media control (down arrow). B, Schematic for testing the role of IL6 and CXCL8 signaling in osteosarcoma-induced migration and quantification relative to vehicle-treated osteosarcoma (OS)–conditioned media (CM; arrow). ANOVA with Dunnett multiple comparisons test. n = 4. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.
    Mouse Anti Human Il6, supplied by R&D Systems, used in various techniques. Bioz Stars score: 93/100, based on 39 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/mouse+anti+human+il6/pmc12616249-175-55-58?v=R%26D+Systems
    Average 93 stars, based on 39 article reviews
    mouse anti human il6 - by Bioz Stars, 2026-07
    93/100 stars

    Images

    1) Product Images from "Metastasis-Initiating Osteosarcoma Subpopulations Establish Paracrine Interactions with Lung and Tumor Cells to Create a Metastatic Niche"

    Article Title: Metastasis-Initiating Osteosarcoma Subpopulations Establish Paracrine Interactions with Lung and Tumor Cells to Create a Metastatic Niche

    Journal: Cancer Research

    doi: 10.1158/0008-5472.CAN-24-3360

    Metastatic osteosarcoma cells promote osteosarcoma chemotaxis via the IL6/CXCL8 mechanism. A, Schematic for testing migration to IL6 and CXCL8 in a Transwell setup and quantification of migrated cells relative to media control (down arrow). B, Schematic for testing the role of IL6 and CXCL8 signaling in osteosarcoma-induced migration and quantification relative to vehicle-treated osteosarcoma (OS)–conditioned media (CM; arrow). ANOVA with Dunnett multiple comparisons test. n = 4. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.
    Figure Legend Snippet: Metastatic osteosarcoma cells promote osteosarcoma chemotaxis via the IL6/CXCL8 mechanism. A, Schematic for testing migration to IL6 and CXCL8 in a Transwell setup and quantification of migrated cells relative to media control (down arrow). B, Schematic for testing the role of IL6 and CXCL8 signaling in osteosarcoma-induced migration and quantification relative to vehicle-treated osteosarcoma (OS)–conditioned media (CM; arrow). ANOVA with Dunnett multiple comparisons test. n = 4. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

    Techniques Used: Chemotaxis Assay, Migration, Control

    Epithelial-derived IL1α drives tumor IL6/CXCL8 production. A, In vitro lung epithelial–osteosarcoma organotypic coculture. Red, mCherry-expressing osteosarcoma cells; green, phalloidin (actin); blue, DAPI. Scale bar, 50 µm. B and C, scRNA-seq analysis of coculture with Seurat-based clustering on Uniform Manifold Approximation and Projection (UMAP) with epithelial cells annotated by expression of KRT19 and osteosarcoma cells annotated by expression of COL1A1 . D, NicheNet heatmap and dot plot analysis demonstrating candidate epithelial-derived ligands (rows) and osteosarcoma-derived receptors (columns). The strength of evidence for ligand–receptor interaction is indicated by the shade of blue in the heatmap, whereas the level of expression and percentage of cells expressing ligand or receptor are noted in dot plots. E, Heatmap demonstrating the regulatory potential of epithelial-derived ligands for the top 50 osteosarcoma differentially expressed genes (coculture vs. monoculture). Red arrows, IL6 , CXCL3 , and CXCL8 , which are strongly regulated IL1 ligands. F, Dot plot of osteosarcoma and epithelial (HBEC3-KT) ligands. Note that IL6 upregulation in coculture occurs in a minority of cells. G, Stimulation with IL1α or IL1β significantly increases secretion of IL6 and CXCL8 in human osteosarcoma cells. H, Epithelial-induced [HBEC3-KT conditioned media (CM)] osteosarcoma IL6 and CXCL8 production requires IL1 signaling (IL1Ra, IL1 receptor antagonist anakinra). ANOVA with Dunnett multiple comparisons test. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.
    Figure Legend Snippet: Epithelial-derived IL1α drives tumor IL6/CXCL8 production. A, In vitro lung epithelial–osteosarcoma organotypic coculture. Red, mCherry-expressing osteosarcoma cells; green, phalloidin (actin); blue, DAPI. Scale bar, 50 µm. B and C, scRNA-seq analysis of coculture with Seurat-based clustering on Uniform Manifold Approximation and Projection (UMAP) with epithelial cells annotated by expression of KRT19 and osteosarcoma cells annotated by expression of COL1A1 . D, NicheNet heatmap and dot plot analysis demonstrating candidate epithelial-derived ligands (rows) and osteosarcoma-derived receptors (columns). The strength of evidence for ligand–receptor interaction is indicated by the shade of blue in the heatmap, whereas the level of expression and percentage of cells expressing ligand or receptor are noted in dot plots. E, Heatmap demonstrating the regulatory potential of epithelial-derived ligands for the top 50 osteosarcoma differentially expressed genes (coculture vs. monoculture). Red arrows, IL6 , CXCL3 , and CXCL8 , which are strongly regulated IL1 ligands. F, Dot plot of osteosarcoma and epithelial (HBEC3-KT) ligands. Note that IL6 upregulation in coculture occurs in a minority of cells. G, Stimulation with IL1α or IL1β significantly increases secretion of IL6 and CXCL8 in human osteosarcoma cells. H, Epithelial-induced [HBEC3-KT conditioned media (CM)] osteosarcoma IL6 and CXCL8 production requires IL1 signaling (IL1Ra, IL1 receptor antagonist anakinra). ANOVA with Dunnett multiple comparisons test. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

    Techniques Used: Derivative Assay, In Vitro, Expressing

    Persistent IL6 and CXCL8 production is limited to a small subpopulation of cells. A, Uniform Manifold Approximation and Projection (UMAP) of tumor cells subsetted from coculture demonstrating that IL6 and CXCL8 expression is limited to a single cluster of cells. B and C, IF of osteosarcoma cells stimulated with IL1α (72 hours) and stained for IL6 (green) and DAPI (blue). Scale bar, 50 µm. The percentage of IL6+ cells was quantified in n = 3 experiments, 100 cells/experiment. D, scRNA-seq analysis of OS-17 tibial xenograft tumors, OS-17 experimental lung metastasis, human primary tumors, and human lung metastasis samples demonstrating the heterogeneity of IL1R1 expression. E, Flow cytometry analysis quantifying surface IL1R1 expression in human osteosarcoma cell lines.
    Figure Legend Snippet: Persistent IL6 and CXCL8 production is limited to a small subpopulation of cells. A, Uniform Manifold Approximation and Projection (UMAP) of tumor cells subsetted from coculture demonstrating that IL6 and CXCL8 expression is limited to a single cluster of cells. B and C, IF of osteosarcoma cells stimulated with IL1α (72 hours) and stained for IL6 (green) and DAPI (blue). Scale bar, 50 µm. The percentage of IL6+ cells was quantified in n = 3 experiments, 100 cells/experiment. D, scRNA-seq analysis of OS-17 tibial xenograft tumors, OS-17 experimental lung metastasis, human primary tumors, and human lung metastasis samples demonstrating the heterogeneity of IL1R1 expression. E, Flow cytometry analysis quantifying surface IL1R1 expression in human osteosarcoma cell lines.

    Techniques Used: Expressing, Staining, Flow Cytometry

    IL1α-induced IL6 and CXCL8 production requires NF-κB signaling in a subpopulation of cells in the G 1 cell-cycle phase. A, OS-17 cells were stimulated with IL1α and then fixed and stained for NF-κB (green) or IL1R1 (red) at the indicated time points. Note the nuclear translocation of NF-κB. Scale bar, 50 µm. B, IKK inhibitors IKK-16 and TPCA-1 abrogate IL1α-induced IL6 and CXCL8 secretion as measured by ELISA. n = 2 biological replicates done in technical triplicates. ANOVA with Dunnett multiple comparisons test. **, P < 0.01; ****, P < 0.0001. C, scRNA-seq analysis of OS-17 tibial xenograft tumors, OS-17 experimental lung metastasis, human primary tumors, and human lung metastasis samples demonstrating restriction of NF-κB activity (see Materials and Methods) in a small subpopulation of G 1 cells.
    Figure Legend Snippet: IL1α-induced IL6 and CXCL8 production requires NF-κB signaling in a subpopulation of cells in the G 1 cell-cycle phase. A, OS-17 cells were stimulated with IL1α and then fixed and stained for NF-κB (green) or IL1R1 (red) at the indicated time points. Note the nuclear translocation of NF-κB. Scale bar, 50 µm. B, IKK inhibitors IKK-16 and TPCA-1 abrogate IL1α-induced IL6 and CXCL8 secretion as measured by ELISA. n = 2 biological replicates done in technical triplicates. ANOVA with Dunnett multiple comparisons test. **, P < 0.01; ****, P < 0.0001. C, scRNA-seq analysis of OS-17 tibial xenograft tumors, OS-17 experimental lung metastasis, human primary tumors, and human lung metastasis samples demonstrating restriction of NF-κB activity (see Materials and Methods) in a small subpopulation of G 1 cells.

    Techniques Used: Staining, Translocation Assay, Enzyme-linked Immunosorbent Assay, Activity Assay

    Sustained IL6 production is maintained by a small subset of hypoproliferative cells that anchor osteosarcoma cells to the lung niche. A, IF expression of IL6 (red) and CXCL8 (green) over time (8–72 hours) following stimulation with IL1α. White, proliferating cells (Ki-67); blue, DAPI. Scale bar, 50 µm. B and C, ELISA and IF quantitation demonstrating that IL6 and CXCL8 production remains constant over time, whereas IL6 and CXCL8+ expression by IF becomes progressively restricted to Ki-67− cells. ELISA, two biological replicates done in technical triplicate. Percentage of IL6, CXCL8, and Ki-67 quantified in n = 3 experiments, 100 cells/experiment. ANOVA with Dunnett multiple comparisons test. ***, P < 0.001; ****, P < 0.0001. D, IL6, CXCL8, and Ki-67 expression in NCH-OS-4 cells following stimulation with IL1α (72 hours). Scale bar, 50 µm. E, F420 murine cells stimulated with murine IL1α (72 hours) and stained with Ki-67 (red), IL6 (green), and DAPI (blue). Scale bar, 50 µm. F, Quantitation of IL6 and Ki-67 staining in NCH-OS-4 and F420 cells.
    Figure Legend Snippet: Sustained IL6 production is maintained by a small subset of hypoproliferative cells that anchor osteosarcoma cells to the lung niche. A, IF expression of IL6 (red) and CXCL8 (green) over time (8–72 hours) following stimulation with IL1α. White, proliferating cells (Ki-67); blue, DAPI. Scale bar, 50 µm. B and C, ELISA and IF quantitation demonstrating that IL6 and CXCL8 production remains constant over time, whereas IL6 and CXCL8+ expression by IF becomes progressively restricted to Ki-67− cells. ELISA, two biological replicates done in technical triplicate. Percentage of IL6, CXCL8, and Ki-67 quantified in n = 3 experiments, 100 cells/experiment. ANOVA with Dunnett multiple comparisons test. ***, P < 0.001; ****, P < 0.0001. D, IL6, CXCL8, and Ki-67 expression in NCH-OS-4 cells following stimulation with IL1α (72 hours). Scale bar, 50 µm. E, F420 murine cells stimulated with murine IL1α (72 hours) and stained with Ki-67 (red), IL6 (green), and DAPI (blue). Scale bar, 50 µm. F, Quantitation of IL6 and Ki-67 staining in NCH-OS-4 and F420 cells.

    Techniques Used: Expressing, Enzyme-linked Immunosorbent Assay, Quantitation Assay, Staining

    IL1 signaling is required for osteosarcoma metastasis progression. A, Functional validation of IL1R1 CRISPR knockout by loss of epithelial-induced IL6 secretion measured by ELISA. n = 4 biological replicates done in triplicate. B, Representative hematoxylin and eosin–stained lungs from mice inoculated with OS-17 electroporated control (NG) or OS-17 IL1R1 CRISPR knockout cell lines ( B and C ). Scale bar, 2 mm. C, Number of metastatic lesions/slice quantified by a blinded examiner. n = 15 mice/condition. ANOVA with Dunnett multiple comparisons test. D and E, Representative images and quantification of the number of metastatic lesions and percentage of metastasis burden (relative to whole lung area) in mice inoculated via tail vein with OS-17 cells, then treated with vehicle (PBS) or anakinra 1 day after tumor injection. Welch t test. Scale bar, 2 mm. F and G, Representative images and quantification of the number of metastatic lesions and percentage of metastasis burden (relative to whole lung area) in mice inoculated via tail vein with F420 cells then treated with vehicle (PBS) or anakinra 1 day after tumor injection. Scale bar, 2 mm. Welch t test. **, P < 0.01; ****, P < 0.0001.
    Figure Legend Snippet: IL1 signaling is required for osteosarcoma metastasis progression. A, Functional validation of IL1R1 CRISPR knockout by loss of epithelial-induced IL6 secretion measured by ELISA. n = 4 biological replicates done in triplicate. B, Representative hematoxylin and eosin–stained lungs from mice inoculated with OS-17 electroporated control (NG) or OS-17 IL1R1 CRISPR knockout cell lines ( B and C ). Scale bar, 2 mm. C, Number of metastatic lesions/slice quantified by a blinded examiner. n = 15 mice/condition. ANOVA with Dunnett multiple comparisons test. D and E, Representative images and quantification of the number of metastatic lesions and percentage of metastasis burden (relative to whole lung area) in mice inoculated via tail vein with OS-17 cells, then treated with vehicle (PBS) or anakinra 1 day after tumor injection. Welch t test. Scale bar, 2 mm. F and G, Representative images and quantification of the number of metastatic lesions and percentage of metastasis burden (relative to whole lung area) in mice inoculated via tail vein with F420 cells then treated with vehicle (PBS) or anakinra 1 day after tumor injection. Scale bar, 2 mm. Welch t test. **, P < 0.01; ****, P < 0.0001.

    Techniques Used: Functional Assay, Biomarker Discovery, CRISPR, Knock-Out, Enzyme-linked Immunosorbent Assay, Staining, Control, Injection



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    Image Search Results


    Metastatic osteosarcoma cells promote osteosarcoma chemotaxis via the IL6/CXCL8 mechanism. A, Schematic for testing migration to IL6 and CXCL8 in a Transwell setup and quantification of migrated cells relative to media control (down arrow). B, Schematic for testing the role of IL6 and CXCL8 signaling in osteosarcoma-induced migration and quantification relative to vehicle-treated osteosarcoma (OS)–conditioned media (CM; arrow). ANOVA with Dunnett multiple comparisons test. n = 4. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

    Journal: Cancer Research

    Article Title: Metastasis-Initiating Osteosarcoma Subpopulations Establish Paracrine Interactions with Lung and Tumor Cells to Create a Metastatic Niche

    doi: 10.1158/0008-5472.CAN-24-3360

    Figure Lengend Snippet: Metastatic osteosarcoma cells promote osteosarcoma chemotaxis via the IL6/CXCL8 mechanism. A, Schematic for testing migration to IL6 and CXCL8 in a Transwell setup and quantification of migrated cells relative to media control (down arrow). B, Schematic for testing the role of IL6 and CXCL8 signaling in osteosarcoma-induced migration and quantification relative to vehicle-treated osteosarcoma (OS)–conditioned media (CM; arrow). ANOVA with Dunnett multiple comparisons test. n = 4. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

    Article Snippet: Primary antibodies used are as follows: rabbit anti-CXCL8 (Abcam, ab289967, IF), mouse anti-human vimentin (Abnova, SRL33, IF, AR = TE), rabbit anti-p21 [Cell Signaling Technology, 2947, IF, AR = CB], rabbit anti–NF-κB (Cell Signaling Technology, 8242, IF), hamster anti-podoplanin (Developmental Studies Hybridoma Bank, 8.1.1, AR = TE), rat anti–Ki-67 (Invitrogen, 14-5698-82, IF, AR = CB), mouse anti-human IL6 (R&D Systems, MAB2061, IF), mouse anti-human IL1α membrane form 488 (R&D Systems, FAB2001G, IF), goat anti-mouse IL1α (R&D Systems, AF-400-NA, IF, AR = TE).

    Techniques: Chemotaxis Assay, Migration, Control

    Epithelial-derived IL1α drives tumor IL6/CXCL8 production. A, In vitro lung epithelial–osteosarcoma organotypic coculture. Red, mCherry-expressing osteosarcoma cells; green, phalloidin (actin); blue, DAPI. Scale bar, 50 µm. B and C, scRNA-seq analysis of coculture with Seurat-based clustering on Uniform Manifold Approximation and Projection (UMAP) with epithelial cells annotated by expression of KRT19 and osteosarcoma cells annotated by expression of COL1A1 . D, NicheNet heatmap and dot plot analysis demonstrating candidate epithelial-derived ligands (rows) and osteosarcoma-derived receptors (columns). The strength of evidence for ligand–receptor interaction is indicated by the shade of blue in the heatmap, whereas the level of expression and percentage of cells expressing ligand or receptor are noted in dot plots. E, Heatmap demonstrating the regulatory potential of epithelial-derived ligands for the top 50 osteosarcoma differentially expressed genes (coculture vs. monoculture). Red arrows, IL6 , CXCL3 , and CXCL8 , which are strongly regulated IL1 ligands. F, Dot plot of osteosarcoma and epithelial (HBEC3-KT) ligands. Note that IL6 upregulation in coculture occurs in a minority of cells. G, Stimulation with IL1α or IL1β significantly increases secretion of IL6 and CXCL8 in human osteosarcoma cells. H, Epithelial-induced [HBEC3-KT conditioned media (CM)] osteosarcoma IL6 and CXCL8 production requires IL1 signaling (IL1Ra, IL1 receptor antagonist anakinra). ANOVA with Dunnett multiple comparisons test. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

    Journal: Cancer Research

    Article Title: Metastasis-Initiating Osteosarcoma Subpopulations Establish Paracrine Interactions with Lung and Tumor Cells to Create a Metastatic Niche

    doi: 10.1158/0008-5472.CAN-24-3360

    Figure Lengend Snippet: Epithelial-derived IL1α drives tumor IL6/CXCL8 production. A, In vitro lung epithelial–osteosarcoma organotypic coculture. Red, mCherry-expressing osteosarcoma cells; green, phalloidin (actin); blue, DAPI. Scale bar, 50 µm. B and C, scRNA-seq analysis of coculture with Seurat-based clustering on Uniform Manifold Approximation and Projection (UMAP) with epithelial cells annotated by expression of KRT19 and osteosarcoma cells annotated by expression of COL1A1 . D, NicheNet heatmap and dot plot analysis demonstrating candidate epithelial-derived ligands (rows) and osteosarcoma-derived receptors (columns). The strength of evidence for ligand–receptor interaction is indicated by the shade of blue in the heatmap, whereas the level of expression and percentage of cells expressing ligand or receptor are noted in dot plots. E, Heatmap demonstrating the regulatory potential of epithelial-derived ligands for the top 50 osteosarcoma differentially expressed genes (coculture vs. monoculture). Red arrows, IL6 , CXCL3 , and CXCL8 , which are strongly regulated IL1 ligands. F, Dot plot of osteosarcoma and epithelial (HBEC3-KT) ligands. Note that IL6 upregulation in coculture occurs in a minority of cells. G, Stimulation with IL1α or IL1β significantly increases secretion of IL6 and CXCL8 in human osteosarcoma cells. H, Epithelial-induced [HBEC3-KT conditioned media (CM)] osteosarcoma IL6 and CXCL8 production requires IL1 signaling (IL1Ra, IL1 receptor antagonist anakinra). ANOVA with Dunnett multiple comparisons test. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

    Article Snippet: Primary antibodies used are as follows: rabbit anti-CXCL8 (Abcam, ab289967, IF), mouse anti-human vimentin (Abnova, SRL33, IF, AR = TE), rabbit anti-p21 [Cell Signaling Technology, 2947, IF, AR = CB], rabbit anti–NF-κB (Cell Signaling Technology, 8242, IF), hamster anti-podoplanin (Developmental Studies Hybridoma Bank, 8.1.1, AR = TE), rat anti–Ki-67 (Invitrogen, 14-5698-82, IF, AR = CB), mouse anti-human IL6 (R&D Systems, MAB2061, IF), mouse anti-human IL1α membrane form 488 (R&D Systems, FAB2001G, IF), goat anti-mouse IL1α (R&D Systems, AF-400-NA, IF, AR = TE).

    Techniques: Derivative Assay, In Vitro, Expressing

    Persistent IL6 and CXCL8 production is limited to a small subpopulation of cells. A, Uniform Manifold Approximation and Projection (UMAP) of tumor cells subsetted from coculture demonstrating that IL6 and CXCL8 expression is limited to a single cluster of cells. B and C, IF of osteosarcoma cells stimulated with IL1α (72 hours) and stained for IL6 (green) and DAPI (blue). Scale bar, 50 µm. The percentage of IL6+ cells was quantified in n = 3 experiments, 100 cells/experiment. D, scRNA-seq analysis of OS-17 tibial xenograft tumors, OS-17 experimental lung metastasis, human primary tumors, and human lung metastasis samples demonstrating the heterogeneity of IL1R1 expression. E, Flow cytometry analysis quantifying surface IL1R1 expression in human osteosarcoma cell lines.

    Journal: Cancer Research

    Article Title: Metastasis-Initiating Osteosarcoma Subpopulations Establish Paracrine Interactions with Lung and Tumor Cells to Create a Metastatic Niche

    doi: 10.1158/0008-5472.CAN-24-3360

    Figure Lengend Snippet: Persistent IL6 and CXCL8 production is limited to a small subpopulation of cells. A, Uniform Manifold Approximation and Projection (UMAP) of tumor cells subsetted from coculture demonstrating that IL6 and CXCL8 expression is limited to a single cluster of cells. B and C, IF of osteosarcoma cells stimulated with IL1α (72 hours) and stained for IL6 (green) and DAPI (blue). Scale bar, 50 µm. The percentage of IL6+ cells was quantified in n = 3 experiments, 100 cells/experiment. D, scRNA-seq analysis of OS-17 tibial xenograft tumors, OS-17 experimental lung metastasis, human primary tumors, and human lung metastasis samples demonstrating the heterogeneity of IL1R1 expression. E, Flow cytometry analysis quantifying surface IL1R1 expression in human osteosarcoma cell lines.

    Article Snippet: Primary antibodies used are as follows: rabbit anti-CXCL8 (Abcam, ab289967, IF), mouse anti-human vimentin (Abnova, SRL33, IF, AR = TE), rabbit anti-p21 [Cell Signaling Technology, 2947, IF, AR = CB], rabbit anti–NF-κB (Cell Signaling Technology, 8242, IF), hamster anti-podoplanin (Developmental Studies Hybridoma Bank, 8.1.1, AR = TE), rat anti–Ki-67 (Invitrogen, 14-5698-82, IF, AR = CB), mouse anti-human IL6 (R&D Systems, MAB2061, IF), mouse anti-human IL1α membrane form 488 (R&D Systems, FAB2001G, IF), goat anti-mouse IL1α (R&D Systems, AF-400-NA, IF, AR = TE).

    Techniques: Expressing, Staining, Flow Cytometry

    IL1α-induced IL6 and CXCL8 production requires NF-κB signaling in a subpopulation of cells in the G 1 cell-cycle phase. A, OS-17 cells were stimulated with IL1α and then fixed and stained for NF-κB (green) or IL1R1 (red) at the indicated time points. Note the nuclear translocation of NF-κB. Scale bar, 50 µm. B, IKK inhibitors IKK-16 and TPCA-1 abrogate IL1α-induced IL6 and CXCL8 secretion as measured by ELISA. n = 2 biological replicates done in technical triplicates. ANOVA with Dunnett multiple comparisons test. **, P < 0.01; ****, P < 0.0001. C, scRNA-seq analysis of OS-17 tibial xenograft tumors, OS-17 experimental lung metastasis, human primary tumors, and human lung metastasis samples demonstrating restriction of NF-κB activity (see Materials and Methods) in a small subpopulation of G 1 cells.

    Journal: Cancer Research

    Article Title: Metastasis-Initiating Osteosarcoma Subpopulations Establish Paracrine Interactions with Lung and Tumor Cells to Create a Metastatic Niche

    doi: 10.1158/0008-5472.CAN-24-3360

    Figure Lengend Snippet: IL1α-induced IL6 and CXCL8 production requires NF-κB signaling in a subpopulation of cells in the G 1 cell-cycle phase. A, OS-17 cells were stimulated with IL1α and then fixed and stained for NF-κB (green) or IL1R1 (red) at the indicated time points. Note the nuclear translocation of NF-κB. Scale bar, 50 µm. B, IKK inhibitors IKK-16 and TPCA-1 abrogate IL1α-induced IL6 and CXCL8 secretion as measured by ELISA. n = 2 biological replicates done in technical triplicates. ANOVA with Dunnett multiple comparisons test. **, P < 0.01; ****, P < 0.0001. C, scRNA-seq analysis of OS-17 tibial xenograft tumors, OS-17 experimental lung metastasis, human primary tumors, and human lung metastasis samples demonstrating restriction of NF-κB activity (see Materials and Methods) in a small subpopulation of G 1 cells.

    Article Snippet: Primary antibodies used are as follows: rabbit anti-CXCL8 (Abcam, ab289967, IF), mouse anti-human vimentin (Abnova, SRL33, IF, AR = TE), rabbit anti-p21 [Cell Signaling Technology, 2947, IF, AR = CB], rabbit anti–NF-κB (Cell Signaling Technology, 8242, IF), hamster anti-podoplanin (Developmental Studies Hybridoma Bank, 8.1.1, AR = TE), rat anti–Ki-67 (Invitrogen, 14-5698-82, IF, AR = CB), mouse anti-human IL6 (R&D Systems, MAB2061, IF), mouse anti-human IL1α membrane form 488 (R&D Systems, FAB2001G, IF), goat anti-mouse IL1α (R&D Systems, AF-400-NA, IF, AR = TE).

    Techniques: Staining, Translocation Assay, Enzyme-linked Immunosorbent Assay, Activity Assay

    Sustained IL6 production is maintained by a small subset of hypoproliferative cells that anchor osteosarcoma cells to the lung niche. A, IF expression of IL6 (red) and CXCL8 (green) over time (8–72 hours) following stimulation with IL1α. White, proliferating cells (Ki-67); blue, DAPI. Scale bar, 50 µm. B and C, ELISA and IF quantitation demonstrating that IL6 and CXCL8 production remains constant over time, whereas IL6 and CXCL8+ expression by IF becomes progressively restricted to Ki-67− cells. ELISA, two biological replicates done in technical triplicate. Percentage of IL6, CXCL8, and Ki-67 quantified in n = 3 experiments, 100 cells/experiment. ANOVA with Dunnett multiple comparisons test. ***, P < 0.001; ****, P < 0.0001. D, IL6, CXCL8, and Ki-67 expression in NCH-OS-4 cells following stimulation with IL1α (72 hours). Scale bar, 50 µm. E, F420 murine cells stimulated with murine IL1α (72 hours) and stained with Ki-67 (red), IL6 (green), and DAPI (blue). Scale bar, 50 µm. F, Quantitation of IL6 and Ki-67 staining in NCH-OS-4 and F420 cells.

    Journal: Cancer Research

    Article Title: Metastasis-Initiating Osteosarcoma Subpopulations Establish Paracrine Interactions with Lung and Tumor Cells to Create a Metastatic Niche

    doi: 10.1158/0008-5472.CAN-24-3360

    Figure Lengend Snippet: Sustained IL6 production is maintained by a small subset of hypoproliferative cells that anchor osteosarcoma cells to the lung niche. A, IF expression of IL6 (red) and CXCL8 (green) over time (8–72 hours) following stimulation with IL1α. White, proliferating cells (Ki-67); blue, DAPI. Scale bar, 50 µm. B and C, ELISA and IF quantitation demonstrating that IL6 and CXCL8 production remains constant over time, whereas IL6 and CXCL8+ expression by IF becomes progressively restricted to Ki-67− cells. ELISA, two biological replicates done in technical triplicate. Percentage of IL6, CXCL8, and Ki-67 quantified in n = 3 experiments, 100 cells/experiment. ANOVA with Dunnett multiple comparisons test. ***, P < 0.001; ****, P < 0.0001. D, IL6, CXCL8, and Ki-67 expression in NCH-OS-4 cells following stimulation with IL1α (72 hours). Scale bar, 50 µm. E, F420 murine cells stimulated with murine IL1α (72 hours) and stained with Ki-67 (red), IL6 (green), and DAPI (blue). Scale bar, 50 µm. F, Quantitation of IL6 and Ki-67 staining in NCH-OS-4 and F420 cells.

    Article Snippet: Primary antibodies used are as follows: rabbit anti-CXCL8 (Abcam, ab289967, IF), mouse anti-human vimentin (Abnova, SRL33, IF, AR = TE), rabbit anti-p21 [Cell Signaling Technology, 2947, IF, AR = CB], rabbit anti–NF-κB (Cell Signaling Technology, 8242, IF), hamster anti-podoplanin (Developmental Studies Hybridoma Bank, 8.1.1, AR = TE), rat anti–Ki-67 (Invitrogen, 14-5698-82, IF, AR = CB), mouse anti-human IL6 (R&D Systems, MAB2061, IF), mouse anti-human IL1α membrane form 488 (R&D Systems, FAB2001G, IF), goat anti-mouse IL1α (R&D Systems, AF-400-NA, IF, AR = TE).

    Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Quantitation Assay, Staining

    IL1 signaling is required for osteosarcoma metastasis progression. A, Functional validation of IL1R1 CRISPR knockout by loss of epithelial-induced IL6 secretion measured by ELISA. n = 4 biological replicates done in triplicate. B, Representative hematoxylin and eosin–stained lungs from mice inoculated with OS-17 electroporated control (NG) or OS-17 IL1R1 CRISPR knockout cell lines ( B and C ). Scale bar, 2 mm. C, Number of metastatic lesions/slice quantified by a blinded examiner. n = 15 mice/condition. ANOVA with Dunnett multiple comparisons test. D and E, Representative images and quantification of the number of metastatic lesions and percentage of metastasis burden (relative to whole lung area) in mice inoculated via tail vein with OS-17 cells, then treated with vehicle (PBS) or anakinra 1 day after tumor injection. Welch t test. Scale bar, 2 mm. F and G, Representative images and quantification of the number of metastatic lesions and percentage of metastasis burden (relative to whole lung area) in mice inoculated via tail vein with F420 cells then treated with vehicle (PBS) or anakinra 1 day after tumor injection. Scale bar, 2 mm. Welch t test. **, P < 0.01; ****, P < 0.0001.

    Journal: Cancer Research

    Article Title: Metastasis-Initiating Osteosarcoma Subpopulations Establish Paracrine Interactions with Lung and Tumor Cells to Create a Metastatic Niche

    doi: 10.1158/0008-5472.CAN-24-3360

    Figure Lengend Snippet: IL1 signaling is required for osteosarcoma metastasis progression. A, Functional validation of IL1R1 CRISPR knockout by loss of epithelial-induced IL6 secretion measured by ELISA. n = 4 biological replicates done in triplicate. B, Representative hematoxylin and eosin–stained lungs from mice inoculated with OS-17 electroporated control (NG) or OS-17 IL1R1 CRISPR knockout cell lines ( B and C ). Scale bar, 2 mm. C, Number of metastatic lesions/slice quantified by a blinded examiner. n = 15 mice/condition. ANOVA with Dunnett multiple comparisons test. D and E, Representative images and quantification of the number of metastatic lesions and percentage of metastasis burden (relative to whole lung area) in mice inoculated via tail vein with OS-17 cells, then treated with vehicle (PBS) or anakinra 1 day after tumor injection. Welch t test. Scale bar, 2 mm. F and G, Representative images and quantification of the number of metastatic lesions and percentage of metastasis burden (relative to whole lung area) in mice inoculated via tail vein with F420 cells then treated with vehicle (PBS) or anakinra 1 day after tumor injection. Scale bar, 2 mm. Welch t test. **, P < 0.01; ****, P < 0.0001.

    Article Snippet: Primary antibodies used are as follows: rabbit anti-CXCL8 (Abcam, ab289967, IF), mouse anti-human vimentin (Abnova, SRL33, IF, AR = TE), rabbit anti-p21 [Cell Signaling Technology, 2947, IF, AR = CB], rabbit anti–NF-κB (Cell Signaling Technology, 8242, IF), hamster anti-podoplanin (Developmental Studies Hybridoma Bank, 8.1.1, AR = TE), rat anti–Ki-67 (Invitrogen, 14-5698-82, IF, AR = CB), mouse anti-human IL6 (R&D Systems, MAB2061, IF), mouse anti-human IL1α membrane form 488 (R&D Systems, FAB2001G, IF), goat anti-mouse IL1α (R&D Systems, AF-400-NA, IF, AR = TE).

    Techniques: Functional Assay, Biomarker Discovery, CRISPR, Knock-Out, Enzyme-linked Immunosorbent Assay, Staining, Control, Injection

    Immunohistochemical Staining

    Journal: Cellular and Molecular Gastroenterology and Hepatology

    Article Title: Concomitant NAFLD Facilitates Liver Metastases and PD-1-Refractory by Recruiting MDSCs via CXCL5/CXCR2 in Colorectal Cancer

    doi: 10.1016/j.jcmgh.2024.04.008

    Figure Lengend Snippet: Immunohistochemical Staining

    Article Snippet: Rabbit anti-mouse/human IL6 , N/A , Proteintech; 21865-1-AP.

    Techniques: Immunohistochemical staining