udisco r d systems af960 spp1  (R&D Systems)

Journal: Frontiers in Immunology

Article Title: The CXCL9/SPP1 polarity axis in tumor-associated macrophages: immunoregulatory and prognostic significance in non-small cell lung cancer

doi: 10.3389/fimmu.2026.1763652

Figure Lengend Snippet: Identification of key cells and annotation of macrophage subsets. (A, B) t-SNE plots showed that CXCL9 and SPP1 were distributed mainly in macrophages among the differential cells. (C) The localization of CXCL9 and SPP1 genes in different cell types was determined, and macrophages accounted for a high proportion. (D) The expression of CXCL9 and SPP1 in differential cells was analyzed, and both genes were differentially expressed in macrophages. (E, F) The identification of 30 PCs was performed. (G) Macrophages were clustered into 15 subclusters after dimensionality reduction. (H) A bubble plot was used to show the expression of CXCL9 and SPP1 in different cell types; both genes were highly expressed in subcluster 3. (I) Macrophage subsets were annotated into 4 subpopulations. (J) A bubble plot was generated to display the high expression of CXCL9 and SPP1 in the annotated cells.

Article Snippet: After antibody stripping, the second round was performed using rabbit anti-human SPP1 monoclonal antibody (1:5000, Servicebio, Cat# GB11500) with HRP-conjugated goat anti-rabbit secondary antibody, followed by iF647-Tyramide (1:500, Servicebio, Cat# G1232).

Techniques: Expressing, Generated

Journal: Frontiers in Immunology

Article Title: The CXCL9/SPP1 polarity axis in tumor-associated macrophages: immunoregulatory and prognostic significance in non-small cell lung cancer

doi: 10.3389/fimmu.2026.1763652

Figure Lengend Snippet: RCTD cell type deconvolution. (A) Visualization of spatial distribution of cell types in spatial transcriptomics between disease and healthy groups; (B) Cell type composition of spatial transcriptomics between disease and healthy groups; (C) Comparative analysis of M1/M2 module scores in four macrophage subtypes (Significance levels were indicated as follows: *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, and ns for not statistically significant); Spatial distribution characteristics of macrophage subtypes in adjacent normal tissue (P24_B2, (D) ] and tumor tissue [P24_T2, (E) ] of non-small cell lung cancer (NSCLC) patient P24; (F) UMAP distribution of macrophage subtypes in NSCLC tumor and paratumor tissues; (G) Average proportion of the 8 nearest neighbor cells around the two macrophage subtypes; (H) Differences in neighborhood composition between the two macrophage subtypes; (I) Spatial lag Moran scatter plot analysis of CXCL9 gene expression; (J) Spatial lag Moran scatter plot analysis of SPP1 gene expression; (K) CXCL9/SPP1 spatial hotspot distribution map; (L) Spatial lag distribution map of SPP1; (M) Spatial lag distribution map of CXCL9.

Article Snippet: After antibody stripping, the second round was performed using rabbit anti-human SPP1 monoclonal antibody (1:5000, Servicebio, Cat# GB11500) with HRP-conjugated goat anti-rabbit secondary antibody, followed by iF647-Tyramide (1:500, Servicebio, Cat# G1232).

Techniques: Spatial Transcriptomics, Gene Expression

Journal: Frontiers in Immunology

Article Title: The CXCL9/SPP1 polarity axis in tumor-associated macrophages: immunoregulatory and prognostic significance in non-small cell lung cancer

doi: 10.3389/fimmu.2026.1763652

Figure Lengend Snippet: Validation of CXCL9 + and SPP1 + macrophage expression and polarization characteristics in lung cancer tissues. (A) Representative mIF staining for CXCL9+ TAMs and SPP1+ TAMs in lung tissue sections from the normal and LUAD tumor groups. DAPI (blue), CD68 (red), CXCL9 (yellow), SPP1 (green) are shown, along with individual and merged channels. (n = 3 per group). Scale bar, 20 μm. (B) Representative mIF staining of CXCL9+ TAMs (yellow) and SPP1+ TAMs (green) in lung tissue sections from the normal and LUAD tumor groups (n = 3 per group). Scale bar, 20 μm. (C) qRT-PCR validation of macrophage polarization. The M1 group showed high expression of Cxcl9 and iNOS, while the M2 group showed high expression of Spp1 and Arg1. Asterisks indicate statistically significant differences (*p < 0.05).

Article Snippet: After antibody stripping, the second round was performed using rabbit anti-human SPP1 monoclonal antibody (1:5000, Servicebio, Cat# GB11500) with HRP-conjugated goat anti-rabbit secondary antibody, followed by iF647-Tyramide (1:500, Servicebio, Cat# G1232).

Techniques: Biomarker Discovery, Expressing, Staining, Quantitative RT-PCR

Journal: Frontiers in Immunology

Article Title: The CXCL9/SPP1 polarity axis in tumor-associated macrophages: immunoregulatory and prognostic significance in non-small cell lung cancer

doi: 10.3389/fimmu.2026.1763652

Figure Lengend Snippet: Acquisition of DEGs. (A) The distribution of 485 DEGs1 between CXCL9 + SPP1 − and CXCL9 − SPP1 + macrophages was analyzed. (B, C) A total of 10,589 DEGs2 in the TCGA training cohort were identified. (D) Differential expression of CXCL9 between the high and low expression groups was detected. (E) Significant differences were observed in the K-M survival curves between the CXCL9 high and low expression groups. (F, G) A total of 1,838 DEGs between the CXCL9 high and low expression groups were identified. (H) Differential expression of SPP1 between the high and low expression groups was detected. (I) Significant differences were found in the KM survival curves between the SPP1 high and low expression groups. (J, K) A total of 2,100 DEGs between the SPP1 high and low expression groups were identified. (L) A Venn diagram was constructed to show the overlap of 91 intersecting genes.

Article Snippet: After antibody stripping, the second round was performed using rabbit anti-human SPP1 monoclonal antibody (1:5000, Servicebio, Cat# GB11500) with HRP-conjugated goat anti-rabbit secondary antibody, followed by iF647-Tyramide (1:500, Servicebio, Cat# G1232).

Techniques: Quantitative Proteomics, Expressing, Construct

Journal: Frontiers in Immunology

Article Title: The CXCL9/SPP1 polarity axis in tumor-associated macrophages: immunoregulatory and prognostic significance in non-small cell lung cancer

doi: 10.3389/fimmu.2026.1763652

Figure Lengend Snippet: Gene set enrichment analysis (GSEA) of CXCL9 and SPP1 high/low expression groups. (A) GSEA was performed between the CXCL9 high- and low-expression groups, and 41 KEGG pathways were significantly enriched. (B) GSEA was conducted between the SPP1 high- and low-expression groups, and 14 KEGG pathways were significantly enriched. (C, D) Single−gene GSEA was performed for CXCL9 and SPP1, and 54 and 51 KEGG pathways were significantly enriched, respectively.

Article Snippet: After antibody stripping, the second round was performed using rabbit anti-human SPP1 monoclonal antibody (1:5000, Servicebio, Cat# GB11500) with HRP-conjugated goat anti-rabbit secondary antibody, followed by iF647-Tyramide (1:500, Servicebio, Cat# G1232).

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: The CXCL9/SPP1 polarity axis in tumor-associated macrophages: immunoregulatory and prognostic significance in non-small cell lung cancer

doi: 10.3389/fimmu.2026.1763652

Figure Lengend Snippet: Pseudotime analysis and cell-cell communication of macrophages. (A) Pseudotime analysis of macrophages. (B) Analysis of pseudotime differentiation trajectories of macrophage subsets with different CXCL9-SPP1 phenotypes. (C) Spatial distribution characteristics of core functional gene expression in macrophage pseudotime trajectories. (D) Heatmap of expression profiles of key macrophage marker genes across different cell clusters. (E) Network diagram of the number of connections between different cell types in the control group. (F) Network diagram of the number of connections between different cell types in the tumor group. (G) Network diagram of the number of connections between macrophages and other cell types in the control group. (H) Network diagram of the number of connections between macrophages and other cell types in the tumor group. (I) Network diagram of connection weights between different cell types in the control group. (J) Network diagram of connection weights between different cell types in the tumor group. (K) Network diagram of connection weights between macrophages and other cell types in the control group. (L) Network diagram of connection weights between macrophages and other cell types in the tumor group.

Article Snippet: After antibody stripping, the second round was performed using rabbit anti-human SPP1 monoclonal antibody (1:5000, Servicebio, Cat# GB11500) with HRP-conjugated goat anti-rabbit secondary antibody, followed by iF647-Tyramide (1:500, Servicebio, Cat# G1232).

Techniques: Functional Assay, Gene Expression, Expressing, Marker, Control

Journal: Frontiers in Immunology

Article Title: The CXCL9/SPP1 polarity axis in tumor-associated macrophages: immunoregulatory and prognostic significance in non-small cell lung cancer

doi: 10.3389/fimmu.2026.1763652

Figure Lengend Snippet: Ligand-receptor interactions and metabolic activity in cell-cell communication. (A) Bubble plot of ligand-receptor interactions in cell-cell communication across different cell types in the control group. (B) Bubble plot of ligand-receptor interactions in cell-cell communication across different cell types in the tumor group. (C) Normalized activity of the top five transcription factors (TFs) in macrophage subsets. (D) Bubble plot of metabolic activity in macrophage subsets. (E, F) Correlation between prognostic genes and CXCL9/SPP1.

Article Snippet: After antibody stripping, the second round was performed using rabbit anti-human SPP1 monoclonal antibody (1:5000, Servicebio, Cat# GB11500) with HRP-conjugated goat anti-rabbit secondary antibody, followed by iF647-Tyramide (1:500, Servicebio, Cat# G1232).

Techniques: Activity Assay, Control

Journal: Cell Biology and Toxicology

Article Title: Revealing an enhanced cytotoxic immune microenvironment at the human maternal–fetal interface in preeclampsia

doi: 10.1007/s10565-026-10170-7

Figure Lengend Snippet: Alteration of decidual NK and macrophage expression levels of SPP1 and CCL pathway associated genes in PE patients. ( A ) Bar plot ranking significant signaling pathways based on differences in overall information flow within inferred cell-cell communication networks between PE and control groups. ( B ) Volcano plot displaying DEGs in dMacro from PE patients compared to Controls. ( C ) UMAP visualization and bar plot of decidua macrophage(dMacro), identifying three major subclusters. ( D ) Violin plots comparing signature scores of dMacro between PE and Control groups, specifically for phagocytosis (GO:0006909). **** p < 0.0001. ( E ) Confocal laser scanning microscopy (CLSM) images showing the secretion of PTN and OGN in peripheral natural killer (pNK) cells, decidual natural killer (dNK) cells, and induced NK (iNK) cells. Scale bars, 10 µm. ( F ) mRNA expression levels of CCL3, CCL4, CCR1, SPP1, and CD44 in decidual tissue from PE (n = 6) and Control (n = 6) samples. Data are presented as mean ± SD, unpaired t test was used for comparisons among two groups. Statistical significance: ** p < 0.01, **** p < 0.0001. ( G ) Circular diagram illustrating cell-cell interactions mediated by the CCL and SPP1 pathways in PE and Control groups. ( H ) Flow cytometry analysis of the percentage of PRF1, GZMA, GZMB, and GNLY expression in iNK cells cultured with or without SPP1(n=5). Data are presented as mean ± SD, unpaired t test was used for comparisons among two groups. Statistical significance: * p < 0.05

Article Snippet: To investigate SPP1's effects on cytotoxic function, differentiated decidual-like NK cells were replated in 24-well plates at 1–1.5 × 106 cells/mL and divided into two groups: 1 Experimental group: Treated with 2 μg/mL recombinant human SPP1 (MedChemExpress; Cat# HY- P70499 ); 2 Control group: Received equivalent volume of sterile double-distilled water.

Techniques: Expressing, Protein-Protein interactions, Control, Confocal Laser Scanning Microscopy, Flow Cytometry, Cell Culture

Journal: bioRxiv

Article Title: NicheSphere reveals Spp1⁺ macrophages as central hubs coordinating fibrotic remodeling in myeloproliferative neoplasms

doi: 10.64898/2026.03.16.711605

Figure Lengend Snippet: (A) Sankey plot showing top 30 differential LR interactions from OsteoCAR cells through ECM glycoproteins towards other co-localized cells. Colors represent LR expression per interaction: orange indicates stronger signaling in disease, blue indicates weaker signaling in disease. (B) Volcano plot for differential gene expression of OsteoCAR-Fibroblast multiplets cell. Genes with logFC > 1 and p-value < 0.05 shown in red; genes with p-value < 0.05 but low logFC in blue and others in green. (C) Representative images of Spp1 staining and quantification of mean DAB intensity in 4um femoral sections of TPO– or EV-induced murine BM fibrosis. Scale bar: 50um. P-value is based on an unpaired t-test.(*) = p<0.05. (D) Schematic representation of experimental timeline, where WT ckit+ donor cells are transduced with TPO-GFP or EV-GFP control lentiviral vectors, and transplanted into lethally-irradiated Spp1KO recipient mice (n=4-5/group). (E) Hemoglobin and red blood cell counts in TPO-driven fibrosis in WT and Spp1KO recipient mice at the time of sacrifice (82 days post-transplant). P-value is based on a one-way ANOVA.(*) = p<0.05,(**)= p<0.01, (***) = p<0.001, (****) = p<0.0001. (F) Blood monocytes quantified by flow cytometry in TPO-driven fibrosis in WT and Spp1KO recipient mice at the time of sacrifice (82 days post-transplant). (G) Spleen weight in TPO-driven fibrosis in WT and Spp1KO recipient mice at the time of sacrifice (82 days post-transplant). P-value is based on a one-way ANOVA.(*) = p<0.05,(**)= p<0.01, (***) = p<0.001, (****) = p<0.0001. (H) Image of spleens isolated from TPO-driven fibrosis in WT and Spp1KO recipient mice at sacrifice. (I) Bone marrow cellularity in TPO-driven fibrosis in WT and Spp1KO recipient mice at the time of sacrifice (82 days post-transplant). (J) Quantification of reticulin staining in 4um femoral sections in TPO-driven fibrosis in WT and Spp1KO recipient mice at the time of sacrifice (82 days post-transplant). (K) Representative imaging of reticulin staining in TPO-driven fibrosis in WT and Spp1KO recipient mice. Scale bar: 50um.

Article Snippet: Samples were diluted 1:50 and SPP1 concentration was quantified using the Human SPP1 Quantikine ELISA (R&D Systems, Human Osteopontin DOST00) according to the manufacturer’s instructions.

Techniques: Expressing, Gene Expression, Staining, Transduction, Control, Irradiation, Flow Cytometry, Isolation, Imaging

Journal: bioRxiv

Article Title: NicheSphere reveals Spp1⁺ macrophages as central hubs coordinating fibrotic remodeling in myeloproliferative neoplasms

doi: 10.64898/2026.03.16.711605

Figure Lengend Snippet: (A) Sankey plot showing top 25 LR pairs associated with Spp1+ macrophage-driven signaling via Spp1 as a ligand. (B) Heatmap of pathway score based on SPP1-associated intracellular signaling, highlighting differential transcriptional response between TPO vs. EV. Higher scores indicate stronger evidence of differential pathway regulation due to up-stream SPP1 signaling. (C) Sankey plot showing SPP1-associated intracellular signaling in Ly6C low infiltrating macrophages and resident-like macrophages, and pathways with enriched differential pathway score from panel B. Genes on the right represent genes downstream of SPP1. Here, only target genes that are ligands are shown. The color map reflects the log-transformed fold change (logFC) between TPO vs. EV. (D) Cytokines localized differential communication network showing cell types (nodes) colored by Niche with node size based on betweenness score. Edge colors represent normalized differential communication scores for cell type pairs. Red color in edges indicates increased signaling in disease, blue indicates decreased signaling in disease. (E) mRNA expression of Col1a1, αSMA and Spp1 in Gli1tdtomato MSCs stimulated with DMSO control, Tgf-β, IL-6, IL-1a, IL-1b and S100A8 for 48 hours, n=3/group, P-value is based on a one-way ANOVA.(*) = p<0.05,(**)= p<0.01, (***) = p<0.001, (****) = p<0.0001. (F) mRNA expression of Col1a1, Spp1 and IL-6 in Gli1tdtomato MSCs stimulated with DMSO control, Tgf-β and recombinant Spp1 for 48 hours, n=3/group, P-value is based on a one-way ANOVA.(*) = p<0.05,(**)= p<0.01, (***) = p<0.001, (****) = p<0.0001.

Article Snippet: Samples were diluted 1:50 and SPP1 concentration was quantified using the Human SPP1 Quantikine ELISA (R&D Systems, Human Osteopontin DOST00) according to the manufacturer’s instructions.

Techniques: Transformation Assay, Expressing, Control, Recombinant

Journal: bioRxiv

Article Title: NicheSphere reveals Spp1⁺ macrophages as central hubs coordinating fibrotic remodeling in myeloproliferative neoplasms

doi: 10.64898/2026.03.16.711605

Figure Lengend Snippet: A. Schematic representation of experimental timeline, where WT or Spp1KO ckit+ donor cells are transduced with TPO-GFP or EV-GFP control lentiviral vectors, and transplanted into lethally-irradiated WT recipient mice (n=5/group, experiment duration: 55 days after transplant). B. Hemoglobin and red blood cell counts in TPO-driven fibrosis in WT recipient mice with WT or Spp1KO BM at the time of sacrifice (55 days post-transplant). P-value is based on a one-way ANOVA.(*) = p<0.05,(**)= p<0.01, (***) = p<0.001, (****) = p<0.0001. C. Blood CD11b+ F4/80+ monocyte-derived macrophages quantified by flow cytometry in TPO-driven WT or Spp1KO fibrosis in WT recipient mice at the time of sacrifice (55 days post-transplant). D. Quantification of reticulin staining in 4um femoral sections in EV– or TPO-driven fibrosis in WT or Spp1KO-driven fibrosis at the time of sacrifice (55 days post-transplant). E. Representative imaging of reticulin staining in 4um femoral sections in TPO-driven WT or Spp1KO-driven fibrosis in WT recipient mice. Scale bar: 50um. F. ELISA analysis of IL-1b protein in plasma of WT or Spp1KO-driven TPO fibrosis. n=3-5/group. G. Representative imaging of IL-1b staining and DAB quantification in 4um femoral sections TPO-driven WT or Spp1KO-driven fibrosis in WT recipient mice. Scale bar: 50um. H. mRNA expression of Spp1, Tnfa and Il-6 in WT or Spp1KO FL-HoxB8 cells (undifferentiated) carrying either the EV or MPLW515L mutation. n=3/group, P-value is based on a one-way ANOVA.(*) = p<0.05,(**)= p<0.01, (***) = p<0.001, (****) = p<0.0001. I. Flow cytometry analysis of macrophage differentiation (CD11b+ F4/80+ cells) from FL-HoxB8 cells (WT vs. Spp1KO, MPLW515L vs. EV) over 5 days, followed by PE+ latex bead phagocytosis assay performed for 1 hour. n=3/group, P-value is based on a one-way ANOVA.(*) = p<0.05, (****) = p<0.0001.

Article Snippet: Samples were diluted 1:50 and SPP1 concentration was quantified using the Human SPP1 Quantikine ELISA (R&D Systems, Human Osteopontin DOST00) according to the manufacturer’s instructions.

Techniques: Transduction, Control, Irradiation, Derivative Assay, Flow Cytometry, Staining, Imaging, Enzyme-linked Immunosorbent Assay, Clinical Proteomics, Expressing, Mutagenesis, Phagocytosis Assay

Journal: bioRxiv

Article Title: NicheSphere reveals Spp1⁺ macrophages as central hubs coordinating fibrotic remodeling in myeloproliferative neoplasms

doi: 10.64898/2026.03.16.711605

Figure Lengend Snippet: (A) ELISA analysis of SPP1 protein in human blood plasma of MPN patients and healthy controls. n=6 healthy, 15 MPN. Statistical test performed is an unpaired t-test, p=0.2796. (B) Normalized SPP1 plasma protein abundance in MPN patients for distinct outcomes. Differences between groups were evaluated using a student t-test. (C) Kaplan–Meier survival curves of primary myelofibrosis patients stratified by quartiles of SPP1 concentration at baseline assessment. Differences between groups were evaluated using the log-rank test.

Article Snippet: Samples were diluted 1:50 and SPP1 concentration was quantified using the Human SPP1 Quantikine ELISA (R&D Systems, Human Osteopontin DOST00) according to the manufacturer’s instructions.

Techniques: Enzyme-linked Immunosorbent Assay, Clinical Proteomics, Quantitative Proteomics, Concentration Assay

Journal: bioRxiv

Article Title: NicheSphere reveals Spp1⁺ macrophages as central hubs coordinating fibrotic remodeling in myeloproliferative neoplasms

doi: 10.64898/2026.03.16.711605

Figure Lengend Snippet:

Article Snippet: Samples were diluted 1:50 and SPP1 concentration was quantified using the Human SPP1 Quantikine ELISA (R&D Systems, Human Osteopontin DOST00) according to the manufacturer’s instructions.

Techniques: