anti-mouse pe-trem-1 (174031)  (R&D Systems)

Journal: Frontiers in Oncology

Article Title: Tumor-Infiltrating Myeloid Cells Co-Express TREM1 and TREM2 and Elevated TREM-1 Associates With Disease Progression in Renal Cell Carcinoma

doi: 10.3389/fonc.2021.662723

Figure Lengend Snippet: TREM-1 is expressed on peripheral myeloid-derived suppressor cells (MDSC) subsets, tumor-infiltrating MDSC, and tumor-associated macrophages in tumor-bearing mice. (A) TREM-1 expression on splenic granulocytic MDSC (PMN-MDSC) and monocytic MDSC (Mo-MDSC) from naïve or tumor-bearing mice 21 days after 4T1 inoculation. (B) TREM-1 levels on tumor-derived Mo-MDSC 0, 7, 15, and 21 days after the 4T1 tumor challenge. * p < 0.05 relative to naïve mice, ** p < 0.0. (C) TREM-1 expression on PMN-MDSC, Mo-MDSC, and tumor-associated macrophages isolated from tumors of tumor-bearing mice 21 days after 4T1 injection. One representative histogram is shown for each population out of more than three independent experiments.

Article Snippet: The anti-mouse PE-TREM-1 (174031) and PE-Rat IgG2A (54447) isotype control as well as recombinant mouse TREM-1 Fc chimera, TREM-2 Fc chimera, and human IgG1 Fc were purchased from R&D Systems.

Techniques: Derivative Assay, Expressing, Isolation, Injection

Journal: Frontiers in Oncology

Article Title: Tumor-Infiltrating Myeloid Cells Co-Express TREM1 and TREM2 and Elevated TREM-1 Associates With Disease Progression in Renal Cell Carcinoma

doi: 10.3389/fonc.2021.662723

Figure Lengend Snippet: TREM-1 engagement enhances pro-inflammatory cytokine release by myeloid-derived suppressor cells (MDSC) and Trem-1 + MDSC from tumor-bearing mice suppress T cell proliferation. (A) Peripheral myeloid-derived suppressor cells were sorted from spleens of 4T1 tumor-bearing mice, plated onto wells coated with either anti-TREM-1 or rat IgG2A isotype control Ab, and stimulated with lipopolysaccharide. The supernatants were harvested 20 h later, and the TNF levels in the supernatants were measured by ELISA. * p < 0.05. (B) Ly6C + CD11b + (Mo) and (C) Ly6G + CD11b + (PMN) MDSC were sorted from spleens of 4T1 tumor-bearing mice. The cells were cultured at the indicated ratios with splenocytes from naïve BALB/c mice. Con A was added to each well at a final concentration of 2 μg/ml. Tritiated thymidine incorporation was measured during the last 18 h of the 85-h incubation period. * p -value <0.05 relative to no suppressors. Data represents mean ± SEM from triplicates of a representative experiment.

Article Snippet: The anti-mouse PE-TREM-1 (174031) and PE-Rat IgG2A (54447) isotype control as well as recombinant mouse TREM-1 Fc chimera, TREM-2 Fc chimera, and human IgG1 Fc were purchased from R&D Systems.

Techniques: Derivative Assay, Enzyme-linked Immunosorbent Assay, Cell Culture, Concentration Assay, Incubation

Journal: Frontiers in Oncology

Article Title: Tumor-Infiltrating Myeloid Cells Co-Express TREM1 and TREM2 and Elevated TREM-1 Associates With Disease Progression in Renal Cell Carcinoma

doi: 10.3389/fonc.2021.662723

Figure Lengend Snippet: TREM-2 is expressed in monocytic MDSC (Mo-MDSC) and tumor-associated macrophages (TAM), and a possible ligand is expressed in TAM. (A) sTREM-2 levels in the plasma of naïve, 4TI-bearing mice or in the wash of 4T1 excised tumors. (B, C) Trem2 expression by qPCR in B6 macrophages, 4T1 cells, splenic MDSC, or TAM as indicated. (B) Relative Trem2 mRNA expression in B6 macrophages compared to 4TI cells demonstrating a neglectable Trem1 expression in tumor cells. (C, D) Relative Trem2 and Trem1 mRNA expression in sorted splenic PMN-MDSC, Mo-MDSC, or TAM sorted from 4T1 tumors removed from mice (see Supplementary Figures S2 – S4 for the gating strategy). Data are normalized to a Trem1 - and Trem2 -expressing macrophage cell line (B6 Line). (E) Single-cell suspensions from 4T1 tumors from tumor-bearing mice were stained with MDSC markers and human (hu) IgG-Fc (gray shaded), TREM-1/hu IgG-Fc (dashed line), or TREM-2/hu IgG-Fc (solid line, no shading), followed by PE-labeled anti-human IgG-Fc. The TREM/hu IgG-Fc expression is shown on gated Gr-1 - CD11b + TAM.

Article Snippet: The anti-mouse PE-TREM-1 (174031) and PE-Rat IgG2A (54447) isotype control as well as recombinant mouse TREM-1 Fc chimera, TREM-2 Fc chimera, and human IgG1 Fc were purchased from R&D Systems.

Techniques: Expressing, Staining, Labeling

Journal: Frontiers in Oncology

Article Title: Tumor-Infiltrating Myeloid Cells Co-Express TREM1 and TREM2 and Elevated TREM-1 Associates With Disease Progression in Renal Cell Carcinoma

doi: 10.3389/fonc.2021.662723

Figure Lengend Snippet: Expression of TREM in RENCA tumors. (A) Representative FACS plot (left) showing the staining of RENCA-associated CD45 + leukocytes with TREM-1 and F4/80. Data from a quadrant analysis of three individual mice are grafted (right). Staining of the quadrants is noted below the graph. (B) Soluble TREM-1 as determined by ELISA. The mean +/- SEM for five individual mice in each group is plotted. (C) qPCR analysis of Trem2 on RNA isolated from whole RENCA tumors (tumor) at day 22 after implantation. Relative values compared to splenocytes isolated from healthy mice (spleen) and splenocytes isolated from tumor-bearing mice (tumor spleen) are shown. * p < 0.05, **p < 0.01, and ****p < 0.0001.

Article Snippet: The anti-mouse PE-TREM-1 (174031) and PE-Rat IgG2A (54447) isotype control as well as recombinant mouse TREM-1 Fc chimera, TREM-2 Fc chimera, and human IgG1 Fc were purchased from R&D Systems.

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

Journal: Frontiers in Oncology

Article Title: Tumor-Infiltrating Myeloid Cells Co-Express TREM1 and TREM2 and Elevated TREM-1 Associates With Disease Progression in Renal Cell Carcinoma

doi: 10.3389/fonc.2021.662723

Figure Lengend Snippet: TREM-1+ myeloid cells infiltrate human renal tumors. (A) Average TREM-1 mean fluorescence intensity on peripheral blood polymorphonuclear neutrophils (PMN) and monocytes (B) from healthy (n = 3) vs. RCC donors (n = 5). (C) CD16 expression on peripheral blood PMN from healthy vs. RCC donors; *pvalue < 0.05.

Article Snippet: The anti-mouse PE-TREM-1 (174031) and PE-Rat IgG2A (54447) isotype control as well as recombinant mouse TREM-1 Fc chimera, TREM-2 Fc chimera, and human IgG1 Fc were purchased from R&D Systems.

Techniques: Fluorescence, Expressing

Journal: Frontiers in Oncology

Article Title: Tumor-Infiltrating Myeloid Cells Co-Express TREM1 and TREM2 and Elevated TREM-1 Associates With Disease Progression in Renal Cell Carcinoma

doi: 10.3389/fonc.2021.662723

Figure Lengend Snippet: sTREM-1 levels are increased in the plasma of patients with renal cell carcinoma. (A) Serum from healthy donors ( n = 10) or patients with either localized ( n = 5) or metastatic ( n = 9) renal cell carcinoma (RCC) was tested for soluble TREM-1 (sTREM-1) levels by ELISA. (B) sTREM-1 levels between stage I (localized) and stage IV (metastatic) RCC patients demonstrating a trend towards higher levels with worsening stage. (C) sTREM-1 levels in RCC patients from the Fox Chase Cancer Center ( n = 63) and healthy controls ( n = 20). (D) Percent of CD45+ blood leukocytes expressing TREM-1 in RCC patients and healthy controls. (E) Soluble TREM-1 levels are plotted vs . the percentages of blood leukocytes expressing TREM-1 in RCC (left) and healthy controls (right). The r values and p -values of the correlation analysis are shown. (F) Fluorescence intensity (geometric mean) of TREM-1 staining of RCC patients is plotted against the sTREM-1 values for those same patients. The r values and p -values of the correlation analysis are shown.

Article Snippet: The anti-mouse PE-TREM-1 (174031) and PE-Rat IgG2A (54447) isotype control as well as recombinant mouse TREM-1 Fc chimera, TREM-2 Fc chimera, and human IgG1 Fc were purchased from R&D Systems.

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

Journal: Frontiers in Oncology

Article Title: Tumor-Infiltrating Myeloid Cells Co-Express TREM1 and TREM2 and Elevated TREM-1 Associates With Disease Progression in Renal Cell Carcinoma

doi: 10.3389/fonc.2021.662723

Figure Lengend Snippet: The Cancer Genome Atlas analysis shows that TREM-1 expression is increased in renal cell carcinoma (RCC) patients and correlates with a poor outcome. (A) Box plots showing Trem1 mRNA expression for normal (72) and stages I–IV ( n = 267, 57, 123, and 84, respectively) RCC patients, with the medians represented as blue bars in a box, and each case is shown as a pink dot. (B) Trem1 expression levels in 531 RCC tumor samples from TCGA were stratified into tertiles of Trem1 expression levels (low = 3rd quartile, high = 3rd tertile, and NC = intertertile range. (C) Kaplan–Meier overall survival curves according to this strata and (D) for higher-stage diseases (stage III and above).

Article Snippet: The anti-mouse PE-TREM-1 (174031) and PE-Rat IgG2A (54447) isotype control as well as recombinant mouse TREM-1 Fc chimera, TREM-2 Fc chimera, and human IgG1 Fc were purchased from R&D Systems.

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: Identification of Extracellular Actin As a Ligand for Triggering Receptor Expressed on Myeloid Cells-1 Signaling

doi: 10.3389/fimmu.2017.00917

Figure Lengend Snippet: FACS analysis of surface triggering receptor expressed on myeloid cells-1 (TREM-1) and its ligand on cells. Mice ( n = 5) were inoculated with LPS for 6 h or mock treated with PBS as a control. The blood cells were collected, and red blood cells were removed for analysis of TREM-1 expression or analysis of the distribution of TREM-1-interacting proteins. All experiments were done in triplicate. (A) FACS analysis of TREM-1 expression on the cell surface with phycoerythrin (PE)-conjugated rat anti-mouse TREM-1 or PE Rat IgG2a, κ Isotype ctrl Antibody, allophycocyanin-conjugated anti-mouse F4/80, and Percp/cy5.5-conjugated anti-mouse Ly-6G. The signal for TREM-1 was specific (Figure in Supplementary Material), and the cells expressing TREM-1 was further analyzed (Figure in Supplementary Material). (B) FACS analysis of the distribution of TREM-1-interacting proteins on cells with Cy5.5-NHS-Ester-labeled recombinant extracellular domain of mouse TREM-1 (rTREM-1), PE/Cy7-conjugated anti-mouse CD41, and PE-conjugated anti-mouse Ly-6G. The cells expressing the ligand for TREM-1 were shown in details (Figure in Supplementary Material).

Article Snippet: The cells were then stained with allophycocyanin-conjugated anti-mouse F4/80 (BioLegend), Percp/cy5.5-conjugated anti-mouse Ly-6G (BioLegend), and phycoerythrin (PE)-conjugated rat IgG2A anti-mouse TREM-1 (R&D systems) or PE-conjugated Rat IgG2a, κIsotype Ctrl Antibody (BioLegend) to analyze which cells expressed surface TREM-1.

Techniques: Control, Expressing, Labeling, Recombinant

Journal: Frontiers in Immunology

Article Title: Identification of Extracellular Actin As a Ligand for Triggering Receptor Expressed on Myeloid Cells-1 Signaling

doi: 10.3389/fimmu.2017.00917

Figure Lengend Snippet: Analysis of the co-localization of triggering receptor expressed on myeloid cells-1 (TREM-1) and actin by laser scanning confocal microscopy. RAW264.7 cells were mock treated with PBS or treated with LPS, platelets, or rACTIN. All cells were fixed and then incubated with mouse TREM-1 antibody antigen affinity-purified polyclonal goat IgG and rabbit anti-beta actin polyclonal antibody, followed by FITC-conjugated affinipure donkey anti-rabbit IgG (H + L) and CY3-conjugated affinipure donkey anti-goat IgG (H + L). After staining the nuclei with Hoechest 33258, the slides were analyzed with a LSM880 with Airyscan laser scanning confocal microscope and ZEN2.3 LITE software. The scale bars in the figure represent 10 µm.

Article Snippet: The cells were then stained with allophycocyanin-conjugated anti-mouse F4/80 (BioLegend), Percp/cy5.5-conjugated anti-mouse Ly-6G (BioLegend), and phycoerythrin (PE)-conjugated rat IgG2A anti-mouse TREM-1 (R&D systems) or PE-conjugated Rat IgG2a, κIsotype Ctrl Antibody (BioLegend) to analyze which cells expressed surface TREM-1.

Techniques: Confocal Microscopy, Incubation, Affinity Purification, Staining, Microscopy, Software

Journal: Frontiers in Immunology

Article Title: Identification of Extracellular Actin As a Ligand for Triggering Receptor Expressed on Myeloid Cells-1 Signaling

doi: 10.3389/fimmu.2017.00917

Figure Lengend Snippet: Analysis of the co-localization of triggering receptor expressed on myeloid cells-1 (TREM-1) and actin in lung sections of cecal ligation and puncture (CLP) mice or healthy control mice. The lung sections from CLP mice ( n = 3) or mock-treated mice ( n = 3) were stained with mouse TREM-1 antibody antigen affinity-purified polyclonal goat IgG and rabbit anti-beta actin polyclonal antibody, followed by FITC-conjugated affinipure donkey anti-rabbit IgG (H + L) and CY3-conjugated affinipure donkey anti-goat IgG (H + L). The cells were analyzed with Fluoview™ Fv1000 laser scanning confocal microscope and FV10-ASW3.1 viewer software. The scale bars in the figure represent 20 µm.

Article Snippet: The cells were then stained with allophycocyanin-conjugated anti-mouse F4/80 (BioLegend), Percp/cy5.5-conjugated anti-mouse Ly-6G (BioLegend), and phycoerythrin (PE)-conjugated rat IgG2A anti-mouse TREM-1 (R&D systems) or PE-conjugated Rat IgG2a, κIsotype Ctrl Antibody (BioLegend) to analyze which cells expressed surface TREM-1.

Techniques: Ligation, Control, Staining, Affinity Purification, Microscopy, Software