Journal: OncoImmunology

Article Title: MICA/B-targeted antibody promotes NK cell–driven tumor immunity in patients with intrahepatic cholangiocarcinoma

doi: 10.1080/2162402x.2022.2035919

Figure Lengend Snippet: Figure 6. Anti-MICA/B 7C6 mAb boosts anti-tumor cytotoxic function of peripheral NK cells from iCCA patients against the HuCCT-1 cell line. A): peripheral NK cell degranulation, evaluated as frequency of CD107a+ NK cells, in iCCA patients (n = 13) and HC (n = 16) in the presence of 7C6 mAb or IgG1-Fc. Parametric paired and unpaired t tests were used to compare data. B): dot plots showing the frequency of CD3-CD56+ CD107a+ NK cells in a HC and a patient (iCCA) in the presence of 7C6 mAb or IgG1-Fc. C): the proportion of circulating IFNγ+ NK cells in patients (n = 10) and HC (n = 11) in the presence of 7C6 mAb compared with IgG1-Fc. To compare paired data, the parametric t test and the non-parametric Wilcoxon t test were used. To compare unpaired data, the parametric t test and the non-parametric Mann-Whitney U test were used. D): representative dot plots showing the frequency of CD3-CD56+ IFNγ+ NK cells in a HC and in a patient (iCCA) in the presence of 7C6 mAb or IgG1-Fc.

Article Snippet: The recombinant human IgG1 Fc (BioXcell, Lebanon, NH, USA) and the humanized anti-MICA/B 7C6-IgG1 mAb were added at a final concentration of 10 μg/ml.

Techniques: MANN-WHITNEY

Journal: OncoImmunology

Article Title: MICA/B-targeted antibody promotes NK cell–driven tumor immunity in patients with intrahepatic cholangiocarcinoma

doi: 10.1080/2162402x.2022.2035919

Figure Lengend Snippet: Figure 7. Anti-MICA/B 7C6 mAb boosts anti-tumor cytotoxic function of peripheral NK cells from iCCA patients against patient-derived iCCA cell lines. A): peripheral NK cell degranulation, evaluated as CD107a+NK frequency, in iCCA patients (n = 12) and HC (n = 8) in the presence of 7C6 mAb or IgG1-Fc using patient- derived primary tumor cell lines as targets. Parametric paired and unpaired t tests were used to compare data. B): dot plots showing the frequency of CD3-CD56 + CD107a+ NK cells in a HC and in a patient (iCCA) in the presence of 7C6 mAb or IgG1-Fc. C): proportion of circulating IFNγ+NK cells in patients (n = 10) and in HC (n = 8) in the presence of 7C6 mAb compared with IgG1-Fc. To compare paired data, we used the parametric t test and the non-parametric Wilcoxon t test. To compare unpaired data, the parametric t test and the non-parametric Mann-Whitney U test were used. D): representative dot plots showing the frequency of CD3-CD56+ IFNγ +NK cells in a HC and in a patient (iCCA) in the presence of 7C6 mAb or IgG1-Fc.

Article Snippet: The recombinant human IgG1 Fc (BioXcell, Lebanon, NH, USA) and the humanized anti-MICA/B 7C6-IgG1 mAb were added at a final concentration of 10 μg/ml.

Techniques: Derivative Assay, MANN-WHITNEY

Journal: OncoImmunology

Article Title: MICA/B-targeted antibody promotes NK cell–driven tumor immunity in patients with intrahepatic cholangiocarcinoma

doi: 10.1080/2162402x.2022.2035919

Figure Lengend Snippet: Figure 8. 7C6 mAb enhances the anti-tumor effect of liver- and tumor-infiltrating NK cells in iCCA patients. A): Frequency of degranulating CD107a+NK cells in LIL (n = 13) and TIL (n = 10) of iCCA patients in the presence of anti-MICA/B 7C6 mAb or IgG1-Fc using autologous tumor-derived cell lines as targets. Parametric paired and unpaired t tests were used to compare data. B): representative dot plots showing the frequency of CD3-CD56+ CD107a+ LIL- and TIL-NK cells in the presence of 7C6 mAb or IgG1-Fc. C): proportion of IFNγ+ NK cells in LIL (n = 10) and TIL (n = 8) of iCCA patients in the presence of 7C6 mAb compared with IgG1-Fc using autologous tumor-derived cell lines as targets. The parametric t test and non-parametric Wilcoxon t test were used to compare paired data. The parametric t test was used to compare unpaired data.

Article Snippet: The recombinant human IgG1 Fc (BioXcell, Lebanon, NH, USA) and the humanized anti-MICA/B 7C6-IgG1 mAb were added at a final concentration of 10 μg/ml.

Techniques: Derivative Assay

Journal: OncoImmunology

Article Title: MICA/B-targeted antibody promotes NK cell–driven tumor immunity in patients with intrahepatic cholangiocarcinoma

doi: 10.1080/2162402x.2022.2035919

Figure Lengend Snippet: Figure 9. Cytotoxicity assay of HC PBMC, patient PBMC, LIL and TIL cells. A, B): Frequency of CFSE+LIVE/DEAD (LD)+ HuCCT-1 cell line targets when HC PBMC (n = 5), patient PBMC (n = 10), LIL (n = 8) and TIL (n = 5) were used as effector cells in the presence of 7C6 mAb and isotype control (IgG1). The parametric paired t tests were used to compare data. C, D): Frequency of CFSE+LD+ patient-derived cell line targets when HC PBMC (n = 5), patient PBMC (n = 8), LIL (n = 8) and TIL (n = 4) were used as effectors in the presence of 7C6 and isotype control. The parametric paired t test was used to compare data in panel C. The non-parametric Wilcoxon t test was used to compare paired data in panel D. Target cell death was determined as frequency of CFSE+LD+ cells.

Article Snippet: The recombinant human IgG1 Fc (BioXcell, Lebanon, NH, USA) and the humanized anti-MICA/B 7C6-IgG1 mAb were added at a final concentration of 10 μg/ml.

Techniques: Cytotoxicity Assay, Control, Derivative Assay

Journal: Journal of Translational Medicine

Article Title: N-acetyltransferase 10 affects the proliferation of intrahepatic cholangiocarcinoma and M2-type polarization of macrophages by regulating C-C motif chemokine ligand 2

doi: 10.1186/s12967-024-05664-z

Figure Lengend Snippet: NAT10 polarizes macrophages toward the M2 type through CCL2 . (A) Macrophages were polarized by treating them with the supernatant of ICC cells for 24 h. (B) After co-culturing ICC cells and macrophages for 24 h, the macrophages underwent polarization. (C) Co-culturing ICC cells with macrophages for 24 h resulted in the polarization of macrophages towards the M2 phenotype. (D) Immunofluorescence showed that CD86 expression increased and CD163 expression decreased in NAT10-knockdown tumors ( n = 6). Scale bars: 50 μm. (E and F) Western blot and ELISA showed that NAT10 knockdown decreased CCL2 expression levels in ICC cells and cell supernatant. (G) CCL2-knockdown cell lines were constructed and verified at the protein level. (H) Flow cytometry confirmed that CCL2 knockdown reduced the polarization of macrophages toward M2. (I) Immunofluorescence showed that CD86 expression increased and CD163 expression decreased in CCL2-knockdown tumors ( n = 6). Scale bars: 50 μm. Data are representative of three or more independent experimental replicates. Data are displayed as the mean ± SD. P -values were determined by Student’s t-test and one-way ANOVA in panels. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. ICC, intrahepatic cholangiocarcinoma; ELISA, enzyme-linked immunosorbent assay; SD, standard deviation; ANOVA, analysis of variance

Article Snippet: The levels of CCL2 in the cell supernatants were determined using an enzyme-linked immunosorbent assay (ELISA) kit (E-EL-H6005, Elabscience).

Techniques: Immunofluorescence, Expressing, Knockdown, Western Blot, Enzyme-linked Immunosorbent Assay, Construct, Flow Cytometry, Standard Deviation

Journal: Journal of Nanobiotechnology

Article Title: Folate-modified biomimetic nanovesicles loaded with a PU.1 inhibitor alleviate atherosclerosis by suppressing inflammation

doi: 10.1186/s12951-025-03825-w

Figure Lengend Snippet: PU.1 is upregulated in macrophages of advanced atherosclerotic lesions and promotes inflammation through IL-1β/NF-κB signaling. ( A ) Boxplot showing expression levels of PU.1 in early versus advanced atherosclerotic plaques based on the GSE43292 dataset. ( B ) UMAP plot of single-cell RNA sequencing data depicting major immune and stromal cell populations in atherosclerotic lesions. ( C ) UMAP feature plot showing SPI1 (encoding PU.1) expression predominantly enriched in macrophages. ( D ) Representative immunofluorescence staining of PU.1 (green) and CD68 (red) in aortic root sections from chow diet– and high fat diet–fed mice. Nuclei were counterstained with DAPI (blue). Scale bar: 200 μm. ( E ) CUT&Tag analysis showing genome-wide binding of PU.1 in macrophages. Heatmap indicates PU.1 enrichment near transcription start sites (TSS). ( F ) Genomic distribution of PU.1 binding peaks identified by CUT&Tag. ( G – H ) Representative CUT&Tag tracks showing PU.1 binding at the promoters of pro-inflammatory cytokines. ( I ) Dual-luciferase reporter assay confirming the transcriptional activation of the IL-1β promoter by PU.1 overexpression (OE). ( J ) Western blot showing that PU.1 knockdown suppressed ox-LDL–induced IL-1β expression and NF-κB pathway activation (p-IκB and p-p65). ( K ) Western blot demonstrating that IL-1β knockdown reversed PU.1-induced NF-κB activation. ( L ) Western blot analysis showing that the PU.1 inhibitor DB1976 attenuated ox-LDL–induced IL-1β expression and NF-κB activation. ( M ) qRT-PCR analysis of pro-inflammatory cytokines (IL-1β, IL-6, TNF-α, MCP-1) in macrophages treated with ox-LDL with or without DB1976. One-way ANOVA with Tukey’s multiple comparison post hoc test was used for statistical analysis. Data are presented as the mean ± standard deviation (SD). * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001

Article Snippet: ELISA kits for mouse IL-1β (E-EL-M0037), IL-6 (E-EL-M0044), TNF-α (E-EL-M3063), and MCP-1 (E-EL-M3001) were purchased from Elabscience (China).

Techniques: Expressing, RNA Sequencing, Immunofluorescence, Staining, Genome Wide, Binding Assay, Luciferase, Reporter Assay, Activation Assay, Over Expression, Western Blot, Knockdown, Quantitative RT-PCR, Comparison, Standard Deviation

Journal: Journal of Nanobiotechnology

Article Title: Folate-modified biomimetic nanovesicles loaded with a PU.1 inhibitor alleviate atherosclerosis by suppressing inflammation

doi: 10.1186/s12951-025-03825-w

Figure Lengend Snippet: The anti-atherosclerotic effects of the D-FNVs. ( A - D ) BMDMs were co-treated with ox-LDL (100 µg/mL) and various formulations (free DB1976, D-NVs, D-FNVs). The levels of IL-1β, IL-6, TNF-α, and MCP-1 in the supernatant were measured by ELISA. ( E , F ) Flow cytometry analysis and quantification of intracellular ROS levels in BMDM cells treated with ox-LDL (100 µg/mL) and various formulations (free DB1976, D-NVs, D-FNVs), respectively, at 2 mM DB1976 for 24 h. ( G , H ) Flow cytometry analysis and quantification of apoptosis rates in BMDMs treated with ox-LDL (100 µg/mL) and various formulations (free DB1976, D-NVs, D-FNVs) at 2 mM DB1976 for 24 h. One-way ANOVA with Tukey’s multiple comparison post hoc test was used for statistical analysis. Data are presented as the mean ± standard deviation (SD). * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001

Article Snippet: ELISA kits for mouse IL-1β (E-EL-M0037), IL-6 (E-EL-M0044), TNF-α (E-EL-M3063), and MCP-1 (E-EL-M3001) were purchased from Elabscience (China).

Techniques: Enzyme-linked Immunosorbent Assay, Flow Cytometry, Comparison, Standard Deviation

Journal: Journal of Nanobiotechnology

Article Title: Folate-modified biomimetic nanovesicles loaded with a PU.1 inhibitor alleviate atherosclerosis by suppressing inflammation

doi: 10.1186/s12951-025-03825-w

Figure Lengend Snippet: D-FNVs ameliorated inflammation in an atherosclerotic mouse model. ( A , B ) Representative images of plaques within the aortic root subjected to immunofluorescent staining for the macrophage marker CD68. Scale bar: 100 μm. ( C – F ) Levels of IL-1β, IL-6, TNF-α, and MCP-1 in aortic tissues collected from atherosclerotic mice treated with various formulations (saline, DB1976, D-NVs, D-FNVs). ( G – J ) Levels of IL-1β, IL-6, TNF-α, and MCP-1 in blood serum collected from the same groups of atherosclerotic mice. The n values are all biological replicates. One-way ANOVA with Tukey’s multiple comparison post hoc test was used for statistical analysis. Data are presented as the mean ± standard deviation (SD). * p < 0.05, ** p < 0.01, and *** p < 0.001. **** p < 0.0001

Article Snippet: ELISA kits for mouse IL-1β (E-EL-M0037), IL-6 (E-EL-M0044), TNF-α (E-EL-M3063), and MCP-1 (E-EL-M3001) were purchased from Elabscience (China).

Techniques: Staining, Marker, Saline, Comparison, Standard Deviation