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CCR1 + -G-MDSCs accumulate to SMAD4-deficient CRC tissues via the CCL15/CCR1 axis and CCL9/CCR1 axis in clinical specimens and mouse models. (A) Clinical specimens of human CRC were examined using immunohistochemistry staining for SMAD4, CCR1 and CD33, and (B) representative images of indicated specimens. Scale bar = 100 μm. (C) Immunohistochemistry analysis of human CRC specimens showing the correlations of SMAD4 and CCR1 expression. Pearson correlation analysis was used to indicate correlation. (D) Fluorescence Activated Cell Sorter (FACS) analysis of the percent of MDSCs in PBMC from healthy subjects and clinical patients with CRC. (E) Representative FACS plots of CD33 + CCR1 + in CD11b + HLA-DR – cells. (F) Quantification of CCL15 concentration in plasma from healthy subjects and clinical patients with CRC. (G) Representative FACS plots and (H) quantification of the percent of CD14 – CD15 + (G-MDSCs) in MDSCs, respectively. G: G-MDSCs, M: MDSCs. (I) MC38 or CT26 cells were injected into spleens of C57BL/6 or BALB/c mice respectively. Mice were reared for 14 days and sacrificed for analysis of liver metastasis, n = 4. (J) Representative FACS plots of MDSCs in CD11b + , CCR1 + in MDSCs from MC38 and CT26 metastasis tissues, respectively. (K) Liver metastasis from mice was stained for CD11b (green) and <t>Ly6G/Ly6C</t> (red). Inset shows invasion front of liver metastasis. Scale bar = 100 μm, scale bar for zoom out = 30 μm. (L) Quantification of the percent of G-MDSCs in CD11b + , n = 4, from MC38 and CT26 metastasis tissues, respectively. (M) Immunohistochemistry staining for CCL9 and CCR1 from MC38 and CT26 metastasis tissues, Scale bar = 100 μm, scale bar for zoom out = 20 μm. (N) Quantification of CCL9 concentration in tumors, n = 4. (O) Schematic summary showing SMAD4-deficient CRC recruited CCR1 + -G-MDSC via the CCL9/CCR1 axis. Data are presented as mean ± SD of at least two independent experiments. Two-tailed unpaired Student's t -test (H, L, N), and Brown-Forsythe and Welch ANOVA tests were used (D, F) for statistical analysis with calculated P values shown.
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CCR1 + -G-MDSCs accumulate to SMAD4-deficient CRC tissues via the CCL15/CCR1 axis and CCL9/CCR1 axis in clinical specimens and mouse models. (A) Clinical specimens of human CRC were examined using immunohistochemistry staining for SMAD4, CCR1 and CD33, and (B) representative images of indicated specimens. Scale bar = 100 μm. (C) Immunohistochemistry analysis of human CRC specimens showing the correlations of SMAD4 and CCR1 expression. Pearson correlation analysis was used to indicate correlation. (D) Fluorescence Activated Cell Sorter (FACS) analysis of the percent of MDSCs in PBMC from healthy subjects and clinical patients with CRC. (E) Representative FACS plots of CD33 + CCR1 + in CD11b + HLA-DR – cells. (F) Quantification of CCL15 concentration in plasma from healthy subjects and clinical patients with CRC. (G) Representative FACS plots and (H) quantification of the percent of CD14 – CD15 + (G-MDSCs) in MDSCs, respectively. G: G-MDSCs, M: MDSCs. (I) MC38 or CT26 cells were injected into spleens of C57BL/6 or BALB/c mice respectively. Mice were reared for 14 days and sacrificed for analysis of liver metastasis, n = 4. (J) Representative FACS plots of MDSCs in CD11b + , CCR1 + in MDSCs from MC38 and CT26 metastasis tissues, respectively. (K) Liver metastasis from mice was stained for CD11b (green) and Ly6G/Ly6C (red). Inset shows invasion front of liver metastasis. Scale bar = 100 μm, scale bar for zoom out = 30 μm. (L) Quantification of the percent of G-MDSCs in CD11b + , n = 4, from MC38 and CT26 metastasis tissues, respectively. (M) Immunohistochemistry staining for CCL9 and CCR1 from MC38 and CT26 metastasis tissues, Scale bar = 100 μm, scale bar for zoom out = 20 μm. (N) Quantification of CCL9 concentration in tumors, n = 4. (O) Schematic summary showing SMAD4-deficient CRC recruited CCR1 + -G-MDSC via the CCL9/CCR1 axis. Data are presented as mean ± SD of at least two independent experiments. Two-tailed unpaired Student's t -test (H, L, N), and Brown-Forsythe and Welch ANOVA tests were used (D, F) for statistical analysis with calculated P values shown.

Journal: Acta Pharmaceutica Sinica. B

Article Title: CCL9/CCR1 axis-driven chemotactic nanovesicles for attenuating metastasis of SMAD4-deficient colorectal cancer by trapping TGF- β

doi: 10.1016/j.apsb.2024.05.009

Figure Lengend Snippet: CCR1 + -G-MDSCs accumulate to SMAD4-deficient CRC tissues via the CCL15/CCR1 axis and CCL9/CCR1 axis in clinical specimens and mouse models. (A) Clinical specimens of human CRC were examined using immunohistochemistry staining for SMAD4, CCR1 and CD33, and (B) representative images of indicated specimens. Scale bar = 100 μm. (C) Immunohistochemistry analysis of human CRC specimens showing the correlations of SMAD4 and CCR1 expression. Pearson correlation analysis was used to indicate correlation. (D) Fluorescence Activated Cell Sorter (FACS) analysis of the percent of MDSCs in PBMC from healthy subjects and clinical patients with CRC. (E) Representative FACS plots of CD33 + CCR1 + in CD11b + HLA-DR – cells. (F) Quantification of CCL15 concentration in plasma from healthy subjects and clinical patients with CRC. (G) Representative FACS plots and (H) quantification of the percent of CD14 – CD15 + (G-MDSCs) in MDSCs, respectively. G: G-MDSCs, M: MDSCs. (I) MC38 or CT26 cells were injected into spleens of C57BL/6 or BALB/c mice respectively. Mice were reared for 14 days and sacrificed for analysis of liver metastasis, n = 4. (J) Representative FACS plots of MDSCs in CD11b + , CCR1 + in MDSCs from MC38 and CT26 metastasis tissues, respectively. (K) Liver metastasis from mice was stained for CD11b (green) and Ly6G/Ly6C (red). Inset shows invasion front of liver metastasis. Scale bar = 100 μm, scale bar for zoom out = 30 μm. (L) Quantification of the percent of G-MDSCs in CD11b + , n = 4, from MC38 and CT26 metastasis tissues, respectively. (M) Immunohistochemistry staining for CCL9 and CCR1 from MC38 and CT26 metastasis tissues, Scale bar = 100 μm, scale bar for zoom out = 20 μm. (N) Quantification of CCL9 concentration in tumors, n = 4. (O) Schematic summary showing SMAD4-deficient CRC recruited CCR1 + -G-MDSC via the CCL9/CCR1 axis. Data are presented as mean ± SD of at least two independent experiments. Two-tailed unpaired Student's t -test (H, L, N), and Brown-Forsythe and Welch ANOVA tests were used (D, F) for statistical analysis with calculated P values shown.

Article Snippet: CD11b, Ly6C, and Ly6G staining assays were served by Servicebio (Hubei Wuhan, China).

Techniques: Immunohistochemistry, Staining, Expressing, Fluorescence, Concentration Assay, Clinical Proteomics, Injection, Two Tailed Test

List of antibodies used to characterize the TIME (membrane staining)

Journal: STAR Protocols

Article Title: Protocol to characterize the melanoma tumor immune microenvironment in mice from single cell to flow cytometry analysis

doi: 10.1016/j.xpro.2023.102690

Figure Lengend Snippet: List of antibodies used to characterize the TIME (membrane staining)

Article Snippet: In Myeloid Derived Suppressor Cells (MDSCs) we could identify monocytic MDSCs (M-MDSC (CD11b+/Ly6C+/Ly6G-/Galectin+)) polymorphonucleate MDSC (PM-MDSC (CD11b+/Ly6C+/mild, Ly6G+/Galectin low)) ( ).

Techniques: Membrane, Staining, Concentration Assay, Derivative Assay

Flow cytometry gating strategy to identify Myeloid Derived Suppressor Cells (MDSCs)

Journal: STAR Protocols

Article Title: Protocol to characterize the melanoma tumor immune microenvironment in mice from single cell to flow cytometry analysis

doi: 10.1016/j.xpro.2023.102690

Figure Lengend Snippet: Flow cytometry gating strategy to identify Myeloid Derived Suppressor Cells (MDSCs)

Article Snippet: In Myeloid Derived Suppressor Cells (MDSCs) we could identify monocytic MDSCs (M-MDSC (CD11b+/Ly6C+/Ly6G-/Galectin+)) polymorphonucleate MDSC (PM-MDSC (CD11b+/Ly6C+/mild, Ly6G+/Galectin low)) ( ).

Techniques: Flow Cytometry, Derivative Assay

List of antibodies used to characterize the TIME (membrane staining)

Journal: STAR Protocols

Article Title: Protocol to characterize the melanoma tumor immune microenvironment in mice from single cell to flow cytometry analysis

doi: 10.1016/j.xpro.2023.102690

Figure Lengend Snippet: List of antibodies used to characterize the TIME (membrane staining)

Article Snippet: In Myeloid Derived Suppressor Cells (MDSCs) we could identify monocytic MDSCs (M-MDSC (CD11b+/Ly6C+/Ly6G-/Galectin+)) polymorphonucleate MDSC (PM-MDSC (CD11b+/Ly6C+/mild, Ly6G+/Galectin low)) ( ).

Techniques: Membrane, Staining, Concentration Assay, Derivative Assay

Flow cytometry gating strategy to identify Myeloid Derived Suppressor Cells (MDSCs)

Journal: STAR Protocols

Article Title: Protocol to characterize the melanoma tumor immune microenvironment in mice from single cell to flow cytometry analysis

doi: 10.1016/j.xpro.2023.102690

Figure Lengend Snippet: Flow cytometry gating strategy to identify Myeloid Derived Suppressor Cells (MDSCs)

Article Snippet: In Myeloid Derived Suppressor Cells (MDSCs) we could identify monocytic MDSCs (M-MDSC (CD11b+/Ly6C+/Ly6G-/Galectin+)) polymorphonucleate MDSC (PM-MDSC (CD11b+/Ly6C+/mild, Ly6G+/Galectin low)) ( ).

Techniques: Flow Cytometry, Derivative Assay