Journal: Chinese Journal of Cancer Research

Article Title: Tim-3 promotes cell aggressiveness and paclitaxel resistance through NF-κB/STAT3 signalling pathway in breast cancer cells

doi: 10.21147/j.issn.1000-9604.2020.05.02

Figure Lengend Snippet: Effect of Tim-3 OE on tube formation of endothelial cells. (A) Tube formation ability of HUVECs cultured in conditioned medium from MDA-MB-231 Tim-3-overexpressing cells (P=0.014 vs . Scr); (B) Tube formation ability of HUVECs cultured in conditioned medium from MCF7 Tim-3-overexpressing cells (P=0.016 vs . Scr); (C) Protein levels of VEGFA, VEGFB and VEGFD following Tim-3 OE (left) and quantitative densitometric analysis (right); (D) mRNA expression of VEGFA and VEGFD genes in breast cancer cells. OE, overexpression; Tim-3, T-cell immunoglobulin and mucin-domain containing molecule-3; HUVEC, human umbilical vein endothelial cell; VEGF, vascular endothelial growth factor. * , P<0.05; ** , P<0.01; *** , P<0.001.

Article Snippet: ELISA was performed using the human VEGFC (cat. no. E-EL-H1600; Elabscience) and vascular endothelial growth factor receptor 2 (VEGFR2) ELISA kit (cat. no. E-EL-H1603; Elabscience), according to the manufacturer’s protocol.

Techniques: Cell Culture, Expressing, Over Expression

Journal: Chinese Journal of Cancer Research

Article Title: Tim-3 promotes cell aggressiveness and paclitaxel resistance through NF-κB/STAT3 signalling pathway in breast cancer cells

doi: 10.21147/j.issn.1000-9604.2020.05.02

Figure Lengend Snippet: VEGFC and VEGFR2 protein levels in conditioned media of stable cell lines indicated by ELISA. (A) VEGFC; (B) VEGFR2. VEGF, vascular endothelial growth factor; ELISA, enzyme-linked immunosorbent assay.

Article Snippet: ELISA was performed using the human VEGFC (cat. no. E-EL-H1600; Elabscience) and vascular endothelial growth factor receptor 2 (VEGFR2) ELISA kit (cat. no. E-EL-H1603; Elabscience), according to the manufacturer’s protocol.

Techniques: Stable Transfection, Enzyme-linked Immunosorbent Assay

Journal: Chinese Journal of Cancer Research

Article Title: Tim-3 promotes cell aggressiveness and paclitaxel resistance through NF-κB/STAT3 signalling pathway in breast cancer cells

doi: 10.21147/j.issn.1000-9604.2020.05.02

Figure Lengend Snippet: Schematic illustration of role of Tim-3 in breast cancer. Upregulation of Tim-3 not only promotes cell proliferation, migration and invasion, but also disrupts cell-cell tight junction, increases angiogenesis of endothelial cells and paclitaxel-resistance. Tim-3 functions in breast cancer cells by activating NF-κB/STAT3 pathway and downstream target genes. Tim-3, T-cell immunoglobulin and mucin-domain containing molecule-3; IL-6, interleukin 6; ZO, zona occludens; VEGF, vascular endothelial growth factor; CCND1, cyclin D1; MMP-1, matrix metalloproteinase-1.

Article Snippet: ELISA was performed using the human VEGFC (cat. no. E-EL-H1600; Elabscience) and vascular endothelial growth factor receptor 2 (VEGFR2) ELISA kit (cat. no. E-EL-H1603; Elabscience), according to the manufacturer’s protocol.

Techniques: Migration

Journal: EMBO Molecular Medicine

Article Title: Selective tubulin-binding drugs induce pericyte phenotype switching and anti-cancer immunity

doi: 10.1038/s44321-025-00222-6

Figure Lengend Snippet: ( A ) 8-week drug treatment scheme in RIP1-Tag5 mice including a 2-week priming and 6-week maintenance phase. ( B ) Representative fluorescent micrographs show vessel perfusion in untreated mice (U), or mice treated with CA-4 (C), eribulin (E), or anti-VEGFR2 antibodies (DC101, D). FITC-lectin (green) overlay (yellow, marked by arrows) with CD31 + (red) blood vessels is shown. Scale bar, 50 µm. ( C ) Representative fluorescent micrographs show staining of the contractile marker CNN1 (red) in NG2 + pericytes (green) and overlay (yellow, marked by arrows). Scale bar, 50 µm. ( D ) Quantification of lectin-perfusion in treatment groups: n = 4 mice for untreated, CA-4 and DC101, n = 11 mice for erbulin treatment groups, ** P = 0.0024, *** P = 0.0002, **** P <0.0001. CD31 + tumor blood vessels: n = 4 mice for untreated, CA-4 and DC101, n = 11 mice for erbulin treatment groups, ** P = 0.0081, DC101 compared to U. CNN1: n = 10 mice for untreated and CA-4, n = 5 mice for eribulin and n = 6 mice for DC101 treatment groups, * P = 0.0139, ** P = 0.0048, *** P = 0.006. NG2 + intratumoral pericytes: n = 10 mice for untreated and CA-4, n = 5 mice for eribulin and n = 6 mice for DC101 treatment groups, * P = 0.0365, DC101 compared to untreated. ( E ) Representative H&E micrographs showing tumor cellularity and collagen-rich tumor capsule (black dotted line) in treatment groups. Quantification of percentage of RIP1-Tag5 tumors displaying an intact collagen capsule (dotted line), n = 8 mice in untreated, n = 5 mice in all treatment groups, **** P <0.0001, DC101 compared to untreated. Scale bar 200 µm (upper images), scale bars for DC101 (lower images), 50 µm. All data were analyzed using one-way ANOVA. Data are expressed as mean ± SD. .

Article Snippet: Combretastatin (CA-4, S7783, 1.5–2.5 mg/kg in 5% DMSO, 10% PEG300 and 85% PBS, Selleck Chemical), paclitaxel (AG-CN2-0045, 0.05, 0.25, 0.5 or 2 mg/kg in 5% DMSO, 95% PBS, AdipoGen Life Sciences), vinorelbine (A10976, Navelbine, 0.25 mg/kg in 5% DMSO, 95% PBS, AdooQ Bioscience), eribulin (0.05 and 0.25 mg/kg in 5% DMSO, 95% PBS, Eisai Inc.) were injected i.v.; for VEGF blocking studies, anti-VEGFR2 antibodies (DC101, 15 mg/kg in PBS, BioXCell) was injected intraperitoneally (i.p.).

Techniques: Staining, Marker

Journal: Cancer Research

Article Title: Apolipoprotein E Promotes Immune Suppression in Pancreatic Cancer through NF-κB–Mediated Production of CXCL1

doi: 10.1158/0008-5472.can-20-3929

Figure Lengend Snippet: Figure 4. ApoE/ mice have fewer monocytic-MDSCs and increased CD8þ T-cell infiltration. A, Experimental scheme for orthotopic transplantation of 7940b, KPC tumor cells. B, tSNE visualization of the 6 cell populations identified using CyTOF in WT and ApoE–/ tumors. Populations identified include macrophages (blue), immature myeloidcells (orange), CD8 T cells (green), CD4 T cells (red), B cells (purple), and nonimmune(brown). C, Manual gating quantitation of cell populations in WT (n ¼ 5– 6) and ApoE–/ (n ¼ 7) tumors. Populations include total immune (CD45þ), B cells (CD45þ CD19þ), total myeloid (CD45þ CD11bþ), macrophages (CD11bþ F4/80þ), TAMs (F4/80þ CD206þ; F4/80þ PD-L1þ), granulocytic-MDSCs (Ly-6Cþ Ly6Gþ), monocytic-MDSCs (Ly-6Cþ Ly-6G), total T cells (CD45þ CD3þ), CD4 T cells (CD3þ

Article Snippet: C57BL/6 and ApoE / mice were administered anti-CD8 antibody (Bio XCell; #BP0060) and anti-CD4 antibody (Bio XCell; # BE0003-1) via intraperitoneal injection at 200 mg/mouse three times weekly.

Techniques: Transplantation Assay, Quantitation Assay

Journal: Cancer Research

Article Title: Apolipoprotein E Promotes Immune Suppression in Pancreatic Cancer through NF-κB–Mediated Production of CXCL1

doi: 10.1158/0008-5472.can-20-3929

Figure Lengend Snippet: Figure 5. Antitumor phenotype in ApoE–/ mice is rescued upon T-cell depletion. A, Experimental design schematic for T-cell depletion in WT and ApoE–/ mice. B, Final tumor weight (g) from WT (n ¼ 6), WT anti-CD4/CD8 (n ¼ 3), ApoE–/ (n ¼ 6), and ApoE–/ anti-CD4/CD8 (n ¼ 6). Statistical significance was determined with a nonparametric Mann–Whitney test. C, Representative SPADE analysis of cellular infiltrate in WT tumor. Identified populations include nonimmune cells, CD8 T cells, CD4 T cells, B cells, immature myeloid cells, macrophages, and CD11cþ myeloid cells. The SPADE plot is colored to indicate CD45 expression. Red, high expression; blue, low expression. D, Manual gating quantitation of cell populations in WT (n ¼ 4), WT anti-CD4/CD8 (n ¼ 2), ApoE–/ (n ¼ 4), and ApoE–/ anti-CD4/CD8 (n ¼ 5) tumors. Populations include CD4 T cells (CD3þ CD4þ) and CD8 T cells (CD3þ CD8þ) E, total myeloid cells (CD45þ CD11bþ), macrophages (CD11bþ F4/80þ), CD11cþ

Article Snippet: C57BL/6 and ApoE / mice were administered anti-CD8 antibody (Bio XCell; #BP0060) and anti-CD4 antibody (Bio XCell; # BE0003-1) via intraperitoneal injection at 200 mg/mouse three times weekly.

Techniques: MANN-WHITNEY, Expressing, Quantitation Assay

Journal: Cancer Research

Article Title: Apolipoprotein E Promotes Immune Suppression in Pancreatic Cancer through NF-κB–Mediated Production of CXCL1

doi: 10.1158/0008-5472.can-20-3929

Figure Lengend Snippet: Figure 7. APOE regulates tumor cell Cxcl1 production via NF-kB signaling. A, Representative Western blot analysis of 7940b KPC tumor cells that were either untreated, treated with scrambled siRNA-negative control, or with LDLR siRNA for 24 hours. a-Tubulin was used as a loading control. Normalized protein expression is denoted under each lane. B, qRT-PCR analysis of Ldlr, Cxcl1, and Cxcl5 mRNA levels relative to Cyclophilin A in 7940b KPC cells that underwent LDLR knockdown for 48 hours and were treated with 0.3 mg/mL recombinant ApoE (n ¼ 3) for 1 hour. Statistical significance was determined using one-way ANOVA with Tukey test for multiple correction. C, Heat map of NF-kB/cytokine signatures in 7940b KPC cells treated with 0.3 mg/mL recombinant ApoE (n ¼ 3) compared with vehicle (n ¼ 3) for 48 hours. Red, high expression; blue, low expression. D, Representative coimmunofluorescence staining of p65 (green), CK19 (red), and DAPI (blue) in 7940b tumor cells in vitro treated with vehicle or 0.3 mg/mL recombinant ApoE for 48 hours. Scale bars, 25 mm. Quantitation of percent nuclear p65 in a 40 field in 7940b cells (n ¼ 4) and 7940b cells treated with 0.3 mg/mL recombinant ApoE (n ¼ 4) for 48 hours. White box represents higher magnification. Statistical significance was determined using two-tailed t tests. E, qRT-PCR analysis of Cxcl1, and Cxcl5 mRNA levels relative to Cyclophilin A in 7940b cells (n ¼ 3), 7940b cells treated with 0.3 mg/mL recombinant ApoE for 2 hours (n ¼ 3), 7940b cells pretreated with 5 mmol/L BAY11-7082 for 1 hour and treated with 0.3 mg/mL recombinant ApoE for 2 hours (n ¼ 3), and 7940b cells pretreated with 10 mmol/L BAY11-7082 for 1 hour and treated with 0.3 mg/mL recombinant ApoE for 2 hours (n ¼ 3). Statistical significance was determined using one-way ANOVA with Tukey test for multiple comparisons. F, Working model. PDAC tumors with active ApoE secretion regulate CXCL1 production from tumor cells and fibroblasts, which in turn recruits immature myeloid cells, resulting in suppression of CD8þ T-cell infiltration.

Article Snippet: C57BL/6 and ApoE / mice were administered anti-CD8 antibody (Bio XCell; #BP0060) and anti-CD4 antibody (Bio XCell; # BE0003-1) via intraperitoneal injection at 200 mg/mouse three times weekly.

Techniques: Western Blot, Negative Control, Control, Expressing, Quantitative RT-PCR, Knockdown, Recombinant, Staining, In Vitro, Quantitation Assay, Two Tailed Test