Journal: Cell Death & Disease

Article Title: Targeting ENO1 reprograms macrophage polarization to trigger antitumor immunity and improves the therapeutic effect of radiotherapy

doi: 10.1038/s41419-026-08416-7

Figure Lengend Snippet: A A total of 2.5 × 10 5 CT26 cells were subcutaneously injected into the left legs of BALB/c mice for 4 days and then intraperitoneally administered anti-ENO1 antibodies (HuL001, 20 mg/kg) five times at three-day intervals. Local radiotherapy was given on Day 10. The tumor volume was recorded every three days. * p < 0.05 and ** p < 0.01. Two-Way ANOVA test ( n = 4). B The resected tumors were weighed on Day 30. * p < 0.05 and *** p < 0.001. One-Way ANOVA test ( n = 3). C The densities of M1 (CD80 + ) and M2 (CD206 + ) macrophages in resected tumors were analyzed by immunofluorescence staining. D The quantification of M1 (CD80 + ) and M2 (CD206 + ) macrophages in resected tumors. * p < 0.05 and ** p < 0.01. One-Way ANOVA test ( n = 3). E The mRNA levels of Cd86 and Cd163 in resected tumors were analyzed by qRT‒PCR ( n = 3). * p < 0.05, ** p < 0.01 and *** p < 0.001. One-Way ANOVA test ( n = 3). F The frequencies of M1 (CD11c + CD11b + F4/80 + CD45 + 7AAD - CD3 - CD19 - ) and M2 (CD206 + CD11b + F4/80 + CD45 + 7AAD - CD3 - CD19 - ) tumor-infiltrating macrophages were analyzed by flow cytometry ( n = 3-4). * p < 0.05 and *** p < 0.001. One-Way ANOVA test. G The quantification of the M1/M2 ratio is shown ( n = 3). * p < 0.05 and ** p < 0.01. One-Way ANOVA test ( n = 3). H The frequencies of CD4 (CD4 + CD3 + CD45 + 7AAD - ) and CD8 (CD8 + CD3 + CD45 + 7AAD - ) tumor-infiltrating T cells were analyzed by flow cytometry ( n = 3-4). * p < 0.05. One-Way ANOVA test. I The quantification of CD8 + TILs is shown. * p < 0.05. One-Way ANOVA test ( n = 3). J The frequency of regulatory CD4 T cells (CD25 + CD127 - CD4 + CD3 + CD45 + 7AAD - ) was analyzed by flow cytometry ( n = 3-4). K The quantification of the CD8/Treg ratio is shown. * p < 0.05 and ** p < 0.01. One-Way ANOVA test ( n = 3). L The densities of dendritic cells (CD11c + ) and cytotoxic T cells (CD8 + ) in resected tumors were analyzed by immunofluorescence staining. The density of CD11c + DCs is shown. * p < 0.05 and ** p < 0.01. One-Way ANOVA test ( n = 3). M The density of CD8 + TILs is shown. * p < 0.05 and ** p < 0.01. One-Way ANOVA test ( n = 3).

Article Snippet: The cells were stained with different antibodies against combinations of surface markers: (1) CD4 and CD8 T cells: ViaDyeRed, CD45-BV785 (E-AB-F1136UD, Elabscience, Texas, USA), CD3-PE-Fir700, CD4-PE/Cy7, CD8a-BV570 (E-AB-F1104UF, Elabscience, Texas, USA), CD127-BV711, CD25, IFNγ-PE and GzmB-AF647; (2) Tumor-associated macrophages: ViaDyeRed, CD3-PE-Fir700, CD19-PE-Fir700, CD45-BV785, CD11b-BV421, F4/80-PE, CD11c-AF488, CD206-APC.

Techniques: Injection, Immunofluorescence, Staining, Flow Cytometry

Journal: Cell Death & Disease

Article Title: Targeting ENO1 reprograms macrophage polarization to trigger antitumor immunity and improves the therapeutic effect of radiotherapy

doi: 10.1038/s41419-026-08416-7

Figure Lengend Snippet: A A total of 5 × 10 5 CT26 cells were subcutaneously injected into the left legs of BALB/c mice for 5 days and then intraperitoneally administered with anti-ENO1 antibodies (HuL001, 40 mg/kg) or clodronate liposomes (50 μL/mouse) on the indicated days ( n = 5). Local radiotherapy was given on Day 10. The tumor volume was recorded every three days. * p < 0.05. Two-Way ANOVA test ( n = 4). B The densities of M1 (CD80 + ) and M2 (CD206 + ) macrophages in resected tumors were analyzed by immunofluorescence staining. The quantification of M1 (CD80 + ) and M2 (CD206 + ) macrophages in resected tumors. * p < 0.05 and *** p < 0.001. One-Way ANOVA test ( n = 3). C 4T1 cells (5 ×10 4 ) were subcutaneously injected into the left legs of BALB/c mice for 4 days and then intraperitoneally administered anti-ENO1 antibodies (HuL001, 20 mg/kg) six times on the indicated days ( n = 5). Local radiotherapy was given on Days 10 and 12. The tumor volume was recorded every three days. * p < 0.05 and *** p < 0.001. Two-Way ANOVA test ( n = 4). D The resected tumors were weighed on Day 40. * p < 0.05. One-Way ANOVA test ( n = 4). E The densities of M1 (CD80 + ) and M2 (CD206 + ) macrophages in resected tumors were analyzed by immunofluorescence staining. F The quantification of M1 (CD80 + ) and M2 (CD206 + ) macrophages in resected tumors. * p < 0.05. One-Way ANOVA test ( n = 3). G The frequencies of M1 (CD11c + CD11b + F4/80 + CD45 + 7AAD - CD3 - CD19 - ) and M2 (CD1206 + CD11b + F4/80 + CD45 + 7AAD - CD3 - CD19 - ) tumor-infiltrating macrophages were analyzed by flow cytometry. * p < 0.05. One-Way ANOVA test ( n = 3-4). H The quantification of the M1/M2 ratio is shown. * p < 0.05. One-Way ANOVA test ( n = 3). I A representative image of flow cytometric analysis of GzmB + (GzmB hi CD8 + CD3 + CD45 + 7AAD - ) T cells. J The frequency of GzmB + (GzmB hi CD8 + CD3 + CD45 + 7AAD - ) T cells was analyzed by flow cytometry. * p < 0.05. One-Way ANOVA test ( n = 3-4). K The density of GzmB + (GzmB hi CD8 + CD3 + CD45 + 7AAD - ) T cells is shown. * p < 0.05. One-Way ANOVA test ( n = 3-4). L The density of IFNγ + (IFNγ hi CD8 + CD3 + CD45 + 7AAD - ) T cells is shown ( n = 3-4). * p < 0.05 and ** p < 0.01. One-Way ANOVA test ( n = 3-4). M The proposed mechanism of TGFβ1/TGFβR/Smad3/PRMT5-mediated ENO1 translocation for lactate release via MCT4.

Article Snippet: The cells were stained with different antibodies against combinations of surface markers: (1) CD4 and CD8 T cells: ViaDyeRed, CD45-BV785 (E-AB-F1136UD, Elabscience, Texas, USA), CD3-PE-Fir700, CD4-PE/Cy7, CD8a-BV570 (E-AB-F1104UF, Elabscience, Texas, USA), CD127-BV711, CD25, IFNγ-PE and GzmB-AF647; (2) Tumor-associated macrophages: ViaDyeRed, CD3-PE-Fir700, CD19-PE-Fir700, CD45-BV785, CD11b-BV421, F4/80-PE, CD11c-AF488, CD206-APC.

Techniques: Injection, Liposomes, Immunofluorescence, Staining, Flow Cytometry, Translocation Assay

Journal: Journal of nanobiotechnology

Article Title: Cerium oxide nanoparticles-carrying human umbilical cord mesenchymal stem cells counteract oxidative damage and facilitate tendon regeneration.

doi: 10.1186/s12951-023-02125-5

Figure Lengend Snippet: Fig. 1 Characterization of hUCMSCs. A Primary hUCMSCs were spindle-shaped under light microscopy. B Flow cytometry analysis showed that 98.5% of cells expressed CD29, 97.4% expressed CD44, 98.6% of cells expressed CD90, and 98.1% of cells expressed CD105. Meanwhile, only 0.48% and 0.28% expressed CD45 and CD34. C Lipid droplets were stained by oil red O after adipogenic differentiation induction of hUCMSCs. Mineralized nodules were stained by Alizarin Red after osteogenic differentiation induction of hUCMSCs. Acid mucopolysaccharide was stained by Alcian Blue after chondrogenic differentiation induction of hUCMSCs

Article Snippet: Briefly, 107 hUCMSCs were subjected to analysis for surface markers CD34 (E-AB-F1143D, Elabscience), CD44 (E-AB-F1100D, Elabscience), CD45 (E-AB-F1137D, Elabscience), CD29 (E-AB-F1049D, Elabscience), CD90 (E-AB-F1167D, Elabscience), and CD105 (E-AB-F1310D, Elabscience) by flow cytometry following the manufacturer’s protocol.

Techniques: Light Microscopy, Flow Cytometry, Staining

Journal: BMC Molecular and Cell Biology

Article Title: Transcriptome and proteome profiling reveal complementary scavenger and immune features of rat liver sinusoidal endothelial cells and liver macrophages

doi: 10.1186/s12860-020-00331-9

Figure Lengend Snippet: Antibodies used in the study

Article Snippet: CD45-PE (OX-1) , CD45, PTPRC , Novus Biologicals , NB100–64895PE , 0.85 μg/ million cells.

Techniques: Concentration Assay, Flow Cytometry, Control, Staining, FACS

Journal: NPJ Precision Oncology

Article Title: Clock pathway inhibitor overcomes tumor immune-exclusion via regulation of fibrocyte differentiation

doi: 10.1038/s41698-025-01066-6

Figure Lengend Snippet: A Double staining of CD8 and αSMA in mouse various subcutaneous tumors. Scale bar, 200 μm. The quantitative evaluation of sections from each tumor stained for B αSMA ( n = 12 fields from four mice per group) and C CD8 ( n = 12 fields from four mice per group). D The correlation between the number of CD8 + T cells and αSMA + area in each mouse tumor tissue. E Double staining of collagen 1 and CD45 to detect fibrocytes in mouse various subcutaneous tumors. Scale bar, 100 μm. F The quantitative evaluation of sections from each tumor stained for fibrocytes ( n = 12 fields from four mice per group). G The correlation between the number of fibrocytes and αSMA + area in each mouse tumor tissue. The correlation was estimated by Spearman’s correlation and a linear regression analysis (the best-fit line is indicated). Representative images of tumor sections resected from non-small cell lung carcinoma (NSCLC) patients stained to detect αSMA + cancer-associated fibroblasts ( H , αSMA + FAP + ) and fibrocytes ( I , CD45 + FSP-1 + ). Scale bar, 100 μm. J Quantitative evaluation of tumor-infiltrating fibrocytes (CD45 + FSP-1 + ) in NSCLC tumor tissue stained in Fig. 1H and I ( n = 50 patients). K Representative images of resected NSCLC tissue stained to detect CD8 + T cells and EpCAM + tumor cells. Scale bar, 100 μm. L Quantitative evaluation of tumor-infiltrating fibrocytes (CD45 + FSP-1 + ) in each NSCLC group divided by the immune phenotypes of tumor-infiltrating CD8 + T cells (inflamed, desert, and exclusion). * P < 0.05 by a one-way ANOVA. All data are shown as the mean ± s.e.m.

Article Snippet: The FAP and CD45 cell surface expression in tumor-infiltrating fibrocytes isolated from AB1-HA tumor tissue were detected by PE-Cy7 conjugated anti-mouse CD45 antibody (1:100 dilution, 30-F11, eBioscience) and PE conjugated anti-mouse FAP antibody (1:100 dilution, Novus Biologicals), respectively.

Techniques: Double Staining, Staining

Journal: NPJ Precision Oncology

Article Title: Clock pathway inhibitor overcomes tumor immune-exclusion via regulation of fibrocyte differentiation

doi: 10.1038/s41698-025-01066-6

Figure Lengend Snippet: A A clustermap of relative transcription factor-activities in tumor-infiltrating fibrocytes and macrophages was estimated by wPGSA from a single cell RNA-seq (scRNA-seq) dataset of mouse tumor-infiltrating CD45 + immune cells in subcutaneous AB1-HA tissue. B UMAP plots of clock genes in scRNA-seq data generated from CD45 + cells in AB1-HA tumor tissue. C The gene expression levels of clock genes in tumor-infiltrating fibrocytes (CD34 + CD45 + cells) and macrophages (CD34 - CD45 + F4/80 + cells) isolated from AB1-HA tumor tissue ( n = 3 per group). * P < 0.05 by Student’s t- test. Data are shown as the mean ± s.e.m. D Monocle-generated plots presenting pseudotime ordering and differentiation trajectory of tumor-infiltrating fibrocyte cluster from AB1-HA tumor tissue. E Monocle-generated plots showing the pseudotime-ordered expression of selected marker genes and clock genes. Lines show the relative expression of each marker in pseudotime.

Article Snippet: The FAP and CD45 cell surface expression in tumor-infiltrating fibrocytes isolated from AB1-HA tumor tissue were detected by PE-Cy7 conjugated anti-mouse CD45 antibody (1:100 dilution, 30-F11, eBioscience) and PE conjugated anti-mouse FAP antibody (1:100 dilution, Novus Biologicals), respectively.

Techniques: RNA Sequencing, Generated, Gene Expression, Isolation, Expressing, Marker

Journal: NPJ Precision Oncology

Article Title: Clock pathway inhibitor overcomes tumor immune-exclusion via regulation of fibrocyte differentiation

doi: 10.1038/s41698-025-01066-6

Figure Lengend Snippet: A A diagram showing the strategy used to isolate fibrocytes from AB1-HA tumor-bearing mice. B The isolation of fibrocytes (CD34 + CD45 + F4/8 mid cells) and macrophages (CD34 - CD45 + F4/80 high cells) from AB1-HA tumor tissue C The gene expression levels of COL1A1 and ACTA2 in AB1-HA tumor-infiltrating fibrocytes treated with KL001 and/or TGF-β ( n = 3 per group). * P < 0.05 by Student’s t- test. D A diagram showing the strategy used to isolate fibrocytes from LLC tumor-bearing mice. E The gene expression level of ACTA2 in LLC tumor-infiltrating fibrocytes treated with KL001 ( n = 3 per group). * P < 0.05 by Student’s t- test. F A diagram showing the strategy used to isolate fibrocytes and fibroblasts from LLC tumor-bearing COL1A2-GFP reporter mice. G The isolation of fibrocytes (CD45 + GFP + cells), fibroblasts (CD45 - GFP + cells), and other immune cells (CD45 + GFP - cells) from LLC tumor - bearing COL1A2-GFP reporter mice. H The gene expression levels of COL1A1, ACTA2, and CD45 in each population isolated in Fig. ( n = 3 per group). * P < 0.05 by Student’s t- test. I The gene expression levels of ACTA2 in LLC tumor-infiltrating fibrocytes and fibroblasts treated with KL001 and/or TGF-β ( n = 3 per group). J A diagram showing the strategy used to isolate fibrocytes from mouse lung. K The gene expression levels of COL1A1 and ACTA2 in lung fibrocytes treated with KL001 and/or TGF-β ( n = 3 per group). * P < 0.05 by Student’s t- test. * P < 0.05 by Student’s t- test. All data are shown as the mean ± s.e.m.

Article Snippet: The FAP and CD45 cell surface expression in tumor-infiltrating fibrocytes isolated from AB1-HA tumor tissue were detected by PE-Cy7 conjugated anti-mouse CD45 antibody (1:100 dilution, 30-F11, eBioscience) and PE conjugated anti-mouse FAP antibody (1:100 dilution, Novus Biologicals), respectively.

Techniques: Isolation, Gene Expression

Journal: NPJ Precision Oncology

Article Title: Clock pathway inhibitor overcomes tumor immune-exclusion via regulation of fibrocyte differentiation

doi: 10.1038/s41698-025-01066-6

Figure Lengend Snippet: A The evaluation of the tumor volume of AB1-HA-bearing mice treated with KL001 from 5 days after tumor cell injection ( n = 6 per group). * P < 0.05 by the Mann-Whitney U test. B The evaluation of the tumor volume of LLC-bearing mice treated with KL001 from 5 days after tumor cell injection ( n = 6 per group). * P < 0.05 by the Mann-Whitney U test. C Representative images and the D quantitative evaluation of sections from AB1-HA tumors stained for αSMA ( n = 15 fields from five mice per group). Scale bar, 200 μm. E Representative images and F the quantitative evaluation of sections from AB1-HA tumors stained for collagen 1a1 ( n = 15 fields from five mice per group). Scale bar, 200 μm. G Representative images and H the quantitative evaluation of sections from AB1-HA tumors stained for collagen 1a1 and CD45 to detect fibrocytes ( n = 16 fields from four mice per group). Scale bar, 100 μm. The tumors were harvested at day 16 from each group studied in Fig. 4A. * P < 0.05 by the Mann-Whitney U test. I The evaluation of the tumor volume of AB1-HA-bearing mice treated with CLK8 from 5 days after tumor cell injection ( n = 5 per group). * P < 0.05 by the Mann-Whitney U test. The quantitative evaluation of sections from AB1-HA tumors stained for αSMA ( J , n = 15 fields from five mice per group), collagen 1a1 ( K , n = 15 from five mice fields per group), and fibrocytes ( L , n = 15 fields from five mice per group). The tumors were harvested at day 16 from each group studied in Fig. 4G. * P < 0.05 by the Mann-Whitney U test. All data are shown as the mean ± s.e.m.

Article Snippet: The FAP and CD45 cell surface expression in tumor-infiltrating fibrocytes isolated from AB1-HA tumor tissue were detected by PE-Cy7 conjugated anti-mouse CD45 antibody (1:100 dilution, 30-F11, eBioscience) and PE conjugated anti-mouse FAP antibody (1:100 dilution, Novus Biologicals), respectively.

Techniques: Injection, MANN-WHITNEY, Staining

Journal: NPJ Precision Oncology

Article Title: Clock pathway inhibitor overcomes tumor immune-exclusion via regulation of fibrocyte differentiation

doi: 10.1038/s41698-025-01066-6

Figure Lengend Snippet: A The evaluation of the tumor volume and B the representative image of tumor tissue of LLC-bearing mice treated with KL001 and/or αPD-L1 Ab (n = 7 per group). * P < 0.05 by a one-way ANOVA. C Representative images and D the quantitative evaluation of sections from LLC tumors stained for αSMA ( n = 15 fields from five mice per group). The tumors were harvested at day 21 from each group studied in Fig. 6A. Scale bar, 200 μm. Representative images and the quantitative evaluation of sections from LLC tumors stained for collagen 1 + CD45 + fibrocytes ( E, F ), CD8 + T cells ( G, H ) and CD4 + T cells ( I, J ). Scale bar, 100 μm. * P < 0.05 by a one-way ANOVA. K The evaluation of the tumor volume of AB1-HA-bearing mice treated with KL001 and/or αCTLA-4 Ab ( n = 6 per group). * P < 0.05 by a one-way ANOVA. The quantitative evaluation of sections from AB1-HA tumors ( n = 15 fields from five mice per group) stained for CD8 ( L ), CD4 ( M ), and Foxp3 ( N ). The tumors were harvested at day 18 from each group studied in Fig. 6K. * P < 0.05 by a one-way ANOVA. All data are shown as the mean ± s.e.m.

Article Snippet: The FAP and CD45 cell surface expression in tumor-infiltrating fibrocytes isolated from AB1-HA tumor tissue were detected by PE-Cy7 conjugated anti-mouse CD45 antibody (1:100 dilution, 30-F11, eBioscience) and PE conjugated anti-mouse FAP antibody (1:100 dilution, Novus Biologicals), respectively.

Techniques: Staining