phospho mertk  (Novus Biologicals)

Journal: Frontiers in Immunology

Article Title: Hydroxychloroquine enhances efferocytosis and modulates inflammation via MerTK/Gas6 signaling in a pristane-induced lupus mouse model

doi: 10.3389/fimmu.2025.1524315

Figure Lengend Snippet: HCQ increases MerTK expression in vitro . (A, B) Gene expression of TAM receptors Mertk (A) and Axl (B) in RAW264.7 cells treated with indicated concentration of HCQ (5, 7.5, 10 μM) for 24 hours with or without co-incubation with apoptotic EL4 cells (n=3). (C, D) Gene expression of TAM ligands Gas6 (C) and Pros (D) in RAW264.7 cells treated with indicated concentration of HCQ (5, 7.5, 10 μM) for 24 hours with or without co-incubation with apoptotic EL4 cells (n=3). (E) Representative flow cytometry images (right) and data plot (left) show surface expression of MerTK on RAW264.7 cells treated with indicated concentration of HCQ (5, 7.5, 10 μM) for 24 hours with or without co-incubation with apoptotic EL4 cells (n=3). (F) Representative WB images (left) and data plot (right) show the protein expression of MerTK, p-MerTK, Gas6, p62, and LC3B in RAW264.7 cells treated with HCQ (5 and 10 μM) for 24 hours (n=3). MFI refers to the median fluorescence intensity. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.

Article Snippet: The membranes were incubated overnight at 4°C with primary antibodies against MerTK (AF591, R&D), p-MerTK (p186-749, PhosphoSolutions), Gas6 (PA5-72882, Thermo Fisher Scientific), p62 (A11250, ABclonal), LC3B (E-AB-70053, Elabscience), PPARγ (E-AB-60059, Elabscience), LXR (A04523-2, Boster), and actin (3700, Cell Signaling).

Techniques: Expressing, In Vitro, Gene Expression, Concentration Assay, Incubation, Flow Cytometry, Fluorescence

Journal: Frontiers in Immunology

Article Title: Hydroxychloroquine enhances efferocytosis and modulates inflammation via MerTK/Gas6 signaling in a pristane-induced lupus mouse model

doi: 10.3389/fimmu.2025.1524315

Figure Lengend Snippet: HCQ enhances efferocytosis, MerTK expression and reduces inflammation in vivo . (A) Representative flow cytometry images (left) and quantification (right) show efferocytosis of the peritoneal macrophages isolated from control, HCQ (30 mg/kg), PIL and PIL+HCQ mice after 1 week of treatment exposed to apoptotic thymocytes (Deep Red-labeled) for 30 minutes (n=5). (B) Representative immunofluorescence images (left) and data plot (right) display peritoneal macrophages attached to culture dishes from control, HCQ, PIL, and PIL+HCQ mice after one week of treatment, exposed to apoptotic thymocytes (Deep Red-labeled) for 30 minutes (n=5). (C) Representative flow cytometry images (left) and data plot (right) show the surface expression of MerTK on the peritoneal macrophages isolated from control, HCQ, PIL, and PIL+HCQ mice after 3 day of treatment (n=5). (D) Representative western blot images (left) and data plot (right) show the protein expression of MerTK and Gas6 in the spleen isolated from control, HCQ, PIL, PIL+HCQ mice after 1 week of treatment (n=3). (E) Spleen weight index from control, HCQ, PIL, and PIL+HCQ mice after 1 week of treatment (n=5). (F) The plasma levels of IL-17A in control, HCQ, PIL, and PIL+HCQ mice after 1 week of treatment (n=5). Scale bar: 170 μm. MFI refers to the median fluorescence intensity. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.

Article Snippet: The membranes were incubated overnight at 4°C with primary antibodies against MerTK (AF591, R&D), p-MerTK (p186-749, PhosphoSolutions), Gas6 (PA5-72882, Thermo Fisher Scientific), p62 (A11250, ABclonal), LC3B (E-AB-70053, Elabscience), PPARγ (E-AB-60059, Elabscience), LXR (A04523-2, Boster), and actin (3700, Cell Signaling).

Techniques: Expressing, In Vivo, Flow Cytometry, Isolation, Control, Labeling, Immunofluorescence, Western Blot, Clinical Proteomics, Fluorescence

Journal: Frontiers in Immunology

Article Title: Hydroxychloroquine enhances efferocytosis and modulates inflammation via MerTK/Gas6 signaling in a pristane-induced lupus mouse model

doi: 10.3389/fimmu.2025.1524315

Figure Lengend Snippet: HCQ promotes efferocytosis and anti-inflammatory signals via MerTK. (A) Representative western blot images (left) and quantitative analysis (right) show the levels of phosphorylated MerTK and total MerTK protein in RAW264.7 cells treated with HCQ (5 μM), UNC2025 (1 μM), or HCQ+UNC2025 after co-incubation with apoptotic EL4 cells for 15 minutes. Macrophages were pretreated with UNC2025 for 1 hour prior to exposure to apoptotic cells (n=3). (B) Representative flow cytometry plots (left) and quantification (right) of efferocytosis in RAW264.7 cells treated with HCQ, UNC2025, or HCQ+UNC2025 following co-incubation with apoptotic EL4 cells for 1 hour (n=3). (C) Representative flow cytometry plots (left) and quantification (right) of efferocytosis in peritoneal macrophages isolated from normal or PIL mice treated with HCQ, UNC2025, or HCQ+UNC2025, following co-incubation with apoptotic thymocytes for 30 minutes (n=3). (D-F) Relative gene expression levels of Ifnα (D) , Il6 (E) , Il10 (F) in RAW264.7 cells treated with HCQ, UNC2025, or HCQ+UNC2025 after co-incubation with apoptotic EL4 cells for 2 hours, compared to untreated controls (n=5). (G) Gene expression of Il6 in peritoneal macrophages isolated from normal or PIL mice treated as indicated, following co-incubation with apoptotic thymocytes for 1 hour (n=3). (H) Representative western blot images (left) and quantification (right) show the expression of anti-inflammatory transcription factors PPARγ and LXR in RAW264.7 cells treated with HCQ, UNC2025, or HCQ+UNC2025 after co-incubation with apoptotic EL4 cells for 2 hours (n=3). *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.

Article Snippet: The membranes were incubated overnight at 4°C with primary antibodies against MerTK (AF591, R&D), p-MerTK (p186-749, PhosphoSolutions), Gas6 (PA5-72882, Thermo Fisher Scientific), p62 (A11250, ABclonal), LC3B (E-AB-70053, Elabscience), PPARγ (E-AB-60059, Elabscience), LXR (A04523-2, Boster), and actin (3700, Cell Signaling).

Techniques: Western Blot, Incubation, Flow Cytometry, Isolation, Gene Expression, Expressing

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet: MerTK mediates HCC resistant to anti-PD-L1 treatment (A) Schematic illustrating the establishment of anti-PD-L1-resistant strains in vivo . (B) Tumor growth curves of subcutaneous implantation models in Hepa1-6 and Res1-6 strains treated with anti-PD-L1 (aPD-L1) or IgG. (C) Survival of orthotopic implantation models of Hepa1-6 and Res1-6 strains treated with aPD-L1 or IgG. (D) Proteomic sequencing analysis was used to analyze the differently expressed proteins in Hepa1-6 and Res1-6. The top 20 differently expressed proteins are presented in a heatmap, including 10 upregulated and 10 downregulated proteins. (E) Western blot of p-MerTK, MerTK, and β-actin in Hepa1-6, Res1-6, HCA-1, and Res-CA1 strains. (F) IHC staining of p-MerTK and MerTK in Hepa1-6 and Res1-6 subcutaneous tumor tissues. (G) Locations of sensitive and resistant tumors were exhibited by MRI, and AFP values before and after anti-PD-L1/PD-1 therapy between patient in sensitive group and patient in resistant group. IHC staining of MerTK expression in HCC tissues from sensitive and resistant patients received anti-PD-1/PD-L1 therapy. (H and I) Subcutaneous xenograft mouse model of Res1-6, Res1-6-sh-MerTK, Hepa1-6, and Hepa1-6-OE-MerTK strains treated with anti-PD-L1 or IgG. When the tumor volume approximately reached 100 mm 3 in size, tumor volume was measured every 3 days. After 25 days of treatment, mice were sacrificed. Shown are tumor appearance and tumor growth curves. All results are shown as mean ± SEM (n = 5). One- or two-way ANOVA was used to analyze the data; ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001.

Article Snippet: Anti-mouse p -MerTK antibody , SAB Signalway Antibody , Cat#13320.

Techniques: In Vivo, Sequencing, Western Blot, Immunohistochemistry, Expressing

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet: MerTK promotes anti-PD-L1 resistance by suppressing ferroptosis in HCC (A) All the differential genes between Hepa1-6 and Res1-6 cells were analyzed using KEGG pathway analysis using the functional gene sets in MSigDB (literature vs. databases containing signaling pathways). (B) Fluorescence detection of lipid ROS by C11-BODIPY (left) and statistical analysis of relative lipid ROS fluorescence signal (right). (C) Fluorescence detection of dead cells by SYTOX Green (left) and statistical analysis of percentage dead cells (right). (D–F) Cell viability of Hepa1-6, Hepa1-6-OE-MerTK, Res1-6, and Res1-6-sh-MerTK strains treated with erastin (5.0 μM) in cocultured condition (left) and statistical analysis of cell survival rate in each time point (right). (G–J) In subcutaneous xenograft mouse model, the statistical analysis of relative lipid ROS and MDA content in Hepa1-6, Hepa1-6-OE-MerTK, Res1-6, and Res1-6-sh-MerTK strains treated with anti-PD-L1 or IgG. All results are shown as mean ± SEM (n = 5). One- or two-way ANOVA was used to analyze the data; ∗∗p < 0.01 and ∗∗∗p < 0.001.

Article Snippet: Anti-mouse p -MerTK antibody , SAB Signalway Antibody , Cat#13320.

Techniques: Functional Assay, Protein-Protein interactions, Fluorescence

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet: Suppression of SLC7A11 sensitizes anti-PD-L1 treatment caused by MerTK (A) Volcano plot of differentially expressed proteins in protein mass spectrometry. (B) Western blot of SLC7A11 expression in Hepa1-6, Res1-6, HCA-1, and Res-CA1. (C) IHC staining of SLC7A11 in Hepa1-6, Res1-6, HCA-1, and Res-CA1 subcutaneous tumor tissues. (D) IHC staining of SLC7A11 expression in HCC tissues from sensitive and resistant patients with anti-PD-1/PD-L1 therapy, and statistical analysis. (E) The correlation between MerTK and SLC7A11 expression in tumor tissues from HCC patients received anti-PD-1/PD-L1 therapy, Pearson product-moment correlation coefficients and p values are shown. (F and G) (F) Schematic illustrating the procedure of anti-PD-L1 or IgG treatment in Res1-6 and Res1-6-shSLC7A11 subcutaneous tumor model, and (G) the representative images of subcutaneous tumor in different groups. (H) Statistical analysis of tumor growth curves. (I and J) Western blot analysis of p-MerTK, MerTK, SLC7A11, and β-actin expression in Hepa1-6, Hepa1-6-OE-MerTK, Res1-6, and Res1-6-shMerTK subcutaneous tumor treated with anti-PD-L1 or IgG. All results are shown as mean ± SEM (n = 5). One- or two-way ANOVA was used to analyze the data; ∗∗p < 0.01 and ∗∗∗p < 0.001.

Article Snippet: Anti-mouse p -MerTK antibody , SAB Signalway Antibody , Cat#13320.

Techniques: Mass Spectrometry, Western Blot, Expressing, Immunohistochemistry

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet: MerTK regulates HCC tumor cell ferroptosis via the ERK/SP1 pathway (A) Western blot analysis of p-MerTK, MerTK, p-ERK, ERK, p-SP1, SP1, SLC7A11, and β-actin expression in Res1-6, Res1-6-shMerTK, and Res1-6 treated with ERK1/2 inhibitor (HY-126288). (B) Western blot analysis of p-MerTK, MerTK, p-ERK, ERK, p-SP1, SP1, SLC7A11, and β-actin expression in Hepa1-6, Hepa1-6-OE-MerTK, and Hepa1-6-OE-MerTK treated with ERK1/2 inhibitor. (C–E) (C) Fluorescence detection of lipid ROS by C11-BODIPY and (D) statistical analysis of relative lipid ROS fluorescence signal in Hepa1-6 tumor cells and (E) in Res1-6 tumor cells. (F–H) (F) Fluorescence detection of dead cells by SYTOX Green, (G) statistical analysis of percentage dead cells in Hepa1-6 tumor cells and (H) in Res1-6 tumor cells. All results are shown as mean ± SEM (n = 5). One- or two-way ANOVA was used to analyze the data; ∗∗p < 0.01 and ∗∗∗p < 0.001.

Article Snippet: Anti-mouse p -MerTK antibody , SAB Signalway Antibody , Cat#13320.

Techniques: Western Blot, Expressing, Fluorescence

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet: MerTK induces anti-PD-L1 resistance by favoring a protumor microenvironment (A) Relationship between overall survival and CTL levels in HCC patients with low and high MerTK gene copy numbers. (B) The correlations between the mRNA expression levels of MerTK and cytotoxic CD8 + T cells. (C) The representative image of HCC tissue stained with MerTK (red), CD8 (gold), CD11b (purple), CD15 (green), and CD14 (pink). (D–G) (D) The percentage statistical analysis of CD8 + T cells (E) MDSCs, (F) gMDSCs, and (G) mMDSCs in tumor tissues. (H) The correlation analysis between the expression of MerTK and the enrichment of MDSCs. (I and J) T-distributed stochastic neighbor embedding (t-SNE) plot of tumor-infiltrating leukocytes overlaid with color-coded clusters and the frequency of clusters of the indicated immune cell subsets, including CD3 + T cells, CD8 + T cells, IFNγ + CD8 + T cells, CD4 + T cells, CD11b + cells, and MDSCs in Hepa1-6, Hepa1-6-OE-MerTK, Res1-6, and Res1-6-shMerTK subcutaneous tumor model treated with anti-PD-L1 or IgG (left) and the statistical analysis (right). All results are shown as mean ± SEM (n = 5). One- or two-way ANOVA was used to analyze the data; ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001.

Article Snippet: Anti-mouse p -MerTK antibody , SAB Signalway Antibody , Cat#13320.

Techniques: Expressing, Staining

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet: Inhibition of MerTK promotes ferroptosis and increases the efficacy of PD-L1 antibody in resistant HCC (A) Cell viability (percentage) analysis of Res1-6 cells following MerTK inhibitors (sitravatinib, UNC5293, UNC2541, and UNC1267) at different concentrations (0, 0.1, 0.5, 1.0, 2.0, 4.0, 8.0, 16.0, and 32.0 μM). (B) Cell death detection by propidium iodide (PI) staining (left) and statistical analysis (right). (C) Fluorescence detection of lipid ROS by C11-BODIPY (left) and statistical analysis of relative lipid ROS fluorescence signal (right). (D) Fluorescence detection of dead cells by SYTOX Green (left) and statistical analysis of percentage dead cells (right). (E) The representative images of subcutaneous tumor in Res1-6 strains were treated with IgG, sitravatinib, anti-PD-L1 or their combination (left) and the statistical analysis of tumor growth curves (right). (F) The representative images of orthotopic tumor in Res1-6 strains were treated with IgG, sitravatinib, anti-PD-L1 or their combination (left) and the statistical analysis of survival curves (right). All results are shown as mean ± SEM (n = 5). One- or two-way ANOVA was used to analyze the data; ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001.

Article Snippet: Anti-mouse p -MerTK antibody , SAB Signalway Antibody , Cat#13320.

Techniques: Inhibition, Staining, Fluorescence

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet: Sitravatinib combined with PD-L1 antibody increases ferroptosis and reduces MDSC infiltration in HCC microenvironment (A and B) (A) T-SNE plot of tumor-infiltrating leukocytes overlaid with color-coded clusters and the frequency of clusters of the indicated immune cell subsets, including CD3 + T cells, CD8 + T cells, IFNγ + CD8 + T cells, CD4 + T cells, CD11b + cells, and MDSCs in Res1-6 strains treated with IgG, sitravatinib, anti-PD-L1, or their combination and (B) the statistical analysis. (C) Western blot analysis of p-MerTK, MerTK, SLC7A11, and β-actin expression in different groups. (D) The representative imagines of IHC staining of p-MerTK, MerTK, and Ki-67 from subcutaneous tumors treated with IgG, sitravatinib, anti-PD-L1 or their combination. Scale bar: 100 μm. (E–H) In subcutaneous xenograft mouse model, the statistical analysis of relative lipid ROS and MDA content in Res1-6 and Res-CA1 strains treated with IgG, sitravatinib, anti-PD-L1, or their combination. All results are shown as mean ± SEM (n = 5). One- or two-way ANOVA was used to analyze the data; ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001.

Article Snippet: Anti-mouse p -MerTK antibody , SAB Signalway Antibody , Cat#13320.

Techniques: Western Blot, Expressing, Immunohistochemistry

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet:

Article Snippet: Anti-mouse p -MerTK antibody , SAB Signalway Antibody , Cat#13320.

Techniques: Virus, Recombinant, Red Blood Cell Lysis, Enzyme-linked Immunosorbent Assay, CCK-8 Assay, Isolation, Multiple Displacement Amplification, Expressing, shRNA, Software

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet: Suppression of SLC7A11 sensitizes anti-PD-L1 treatment caused by MerTK (A) Volcano plot of differentially expressed proteins in protein mass spectrometry. (B) Western blot of SLC7A11 expression in Hepa1-6, Res1-6, HCA-1, and Res-CA1. (C) IHC staining of SLC7A11 in Hepa1-6, Res1-6, HCA-1, and Res-CA1 subcutaneous tumor tissues. (D) IHC staining of SLC7A11 expression in HCC tissues from sensitive and resistant patients with anti-PD-1/PD-L1 therapy, and statistical analysis. (E) The correlation between MerTK and SLC7A11 expression in tumor tissues from HCC patients received anti-PD-1/PD-L1 therapy, Pearson product-moment correlation coefficients and p values are shown. (F and G) (F) Schematic illustrating the procedure of anti-PD-L1 or IgG treatment in Res1-6 and Res1-6-shSLC7A11 subcutaneous tumor model, and (G) the representative images of subcutaneous tumor in different groups. (H) Statistical analysis of tumor growth curves. (I and J) Western blot analysis of p-MerTK, MerTK, SLC7A11, and β-actin expression in Hepa1-6, Hepa1-6-OE-MerTK, Res1-6, and Res1-6-shMerTK subcutaneous tumor treated with anti-PD-L1 or IgG. All results are shown as mean ± SEM (n = 5). One- or two-way ANOVA was used to analyze the data; ∗∗p < 0.01 and ∗∗∗p < 0.001.

Article Snippet: Briefly, following deparaffinization, rehydrating and antigen retrieval, primary antibodies were applied to slides, incubated at 4°C overnight, followed by incubation with secondary antibody (Dako Denmark A/S, Glostrup, Denmark) at 37°C for 30 min. SLC7A11 antibody (Proteintech, #26864-1-AP), MerTK antibody (SAB Signalway Antibody, #32851), p -MerTK antibody (SAB Signalway Antibody, #13320), Ki67 antibody (Abcam, #ab92742), CD3 antibody (Abcam, #ab16669), CD8 antibody (Abcam, #ab237709), CD11b antibody (Abcam, #ab52478), Gr1 antibody (Abcam, #ab25377) were used as primary antibodies.

Techniques: Mass Spectrometry, Western Blot, Expressing, Immunohistochemistry

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet: MerTK regulates HCC tumor cell ferroptosis via the ERK/SP1 pathway (A) Western blot analysis of p-MerTK, MerTK, p-ERK, ERK, p-SP1, SP1, SLC7A11, and β-actin expression in Res1-6, Res1-6-shMerTK, and Res1-6 treated with ERK1/2 inhibitor (HY-126288). (B) Western blot analysis of p-MerTK, MerTK, p-ERK, ERK, p-SP1, SP1, SLC7A11, and β-actin expression in Hepa1-6, Hepa1-6-OE-MerTK, and Hepa1-6-OE-MerTK treated with ERK1/2 inhibitor. (C–E) (C) Fluorescence detection of lipid ROS by C11-BODIPY and (D) statistical analysis of relative lipid ROS fluorescence signal in Hepa1-6 tumor cells and (E) in Res1-6 tumor cells. (F–H) (F) Fluorescence detection of dead cells by SYTOX Green, (G) statistical analysis of percentage dead cells in Hepa1-6 tumor cells and (H) in Res1-6 tumor cells. All results are shown as mean ± SEM (n = 5). One- or two-way ANOVA was used to analyze the data; ∗∗p < 0.01 and ∗∗∗p < 0.001.

Article Snippet: Briefly, following deparaffinization, rehydrating and antigen retrieval, primary antibodies were applied to slides, incubated at 4°C overnight, followed by incubation with secondary antibody (Dako Denmark A/S, Glostrup, Denmark) at 37°C for 30 min. SLC7A11 antibody (Proteintech, #26864-1-AP), MerTK antibody (SAB Signalway Antibody, #32851), p -MerTK antibody (SAB Signalway Antibody, #13320), Ki67 antibody (Abcam, #ab92742), CD3 antibody (Abcam, #ab16669), CD8 antibody (Abcam, #ab237709), CD11b antibody (Abcam, #ab52478), Gr1 antibody (Abcam, #ab25377) were used as primary antibodies.

Techniques: Western Blot, Expressing, Fluorescence

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet: MerTK induces anti-PD-L1 resistance by favoring a protumor microenvironment (A) Relationship between overall survival and CTL levels in HCC patients with low and high MerTK gene copy numbers. (B) The correlations between the mRNA expression levels of MerTK and cytotoxic CD8 + T cells. (C) The representative image of HCC tissue stained with MerTK (red), CD8 (gold), CD11b (purple), CD15 (green), and CD14 (pink). (D–G) (D) The percentage statistical analysis of CD8 + T cells (E) MDSCs, (F) gMDSCs, and (G) mMDSCs in tumor tissues. (H) The correlation analysis between the expression of MerTK and the enrichment of MDSCs. (I and J) T-distributed stochastic neighbor embedding (t-SNE) plot of tumor-infiltrating leukocytes overlaid with color-coded clusters and the frequency of clusters of the indicated immune cell subsets, including CD3 + T cells, CD8 + T cells, IFNγ + CD8 + T cells, CD4 + T cells, CD11b + cells, and MDSCs in Hepa1-6, Hepa1-6-OE-MerTK, Res1-6, and Res1-6-shMerTK subcutaneous tumor model treated with anti-PD-L1 or IgG (left) and the statistical analysis (right). All results are shown as mean ± SEM (n = 5). One- or two-way ANOVA was used to analyze the data; ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001.

Article Snippet: Briefly, following deparaffinization, rehydrating and antigen retrieval, primary antibodies were applied to slides, incubated at 4°C overnight, followed by incubation with secondary antibody (Dako Denmark A/S, Glostrup, Denmark) at 37°C for 30 min. SLC7A11 antibody (Proteintech, #26864-1-AP), MerTK antibody (SAB Signalway Antibody, #32851), p -MerTK antibody (SAB Signalway Antibody, #13320), Ki67 antibody (Abcam, #ab92742), CD3 antibody (Abcam, #ab16669), CD8 antibody (Abcam, #ab237709), CD11b antibody (Abcam, #ab52478), Gr1 antibody (Abcam, #ab25377) were used as primary antibodies.

Techniques: Expressing, Staining

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet: Sitravatinib combined with PD-L1 antibody increases ferroptosis and reduces MDSC infiltration in HCC microenvironment (A and B) (A) T-SNE plot of tumor-infiltrating leukocytes overlaid with color-coded clusters and the frequency of clusters of the indicated immune cell subsets, including CD3 + T cells, CD8 + T cells, IFNγ + CD8 + T cells, CD4 + T cells, CD11b + cells, and MDSCs in Res1-6 strains treated with IgG, sitravatinib, anti-PD-L1, or their combination and (B) the statistical analysis. (C) Western blot analysis of p-MerTK, MerTK, SLC7A11, and β-actin expression in different groups. (D) The representative imagines of IHC staining of p-MerTK, MerTK, and Ki-67 from subcutaneous tumors treated with IgG, sitravatinib, anti-PD-L1 or their combination. Scale bar: 100 μm. (E–H) In subcutaneous xenograft mouse model, the statistical analysis of relative lipid ROS and MDA content in Res1-6 and Res-CA1 strains treated with IgG, sitravatinib, anti-PD-L1, or their combination. All results are shown as mean ± SEM (n = 5). One- or two-way ANOVA was used to analyze the data; ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001.

Article Snippet: Briefly, following deparaffinization, rehydrating and antigen retrieval, primary antibodies were applied to slides, incubated at 4°C overnight, followed by incubation with secondary antibody (Dako Denmark A/S, Glostrup, Denmark) at 37°C for 30 min. SLC7A11 antibody (Proteintech, #26864-1-AP), MerTK antibody (SAB Signalway Antibody, #32851), p -MerTK antibody (SAB Signalway Antibody, #13320), Ki67 antibody (Abcam, #ab92742), CD3 antibody (Abcam, #ab16669), CD8 antibody (Abcam, #ab237709), CD11b antibody (Abcam, #ab52478), Gr1 antibody (Abcam, #ab25377) were used as primary antibodies.

Techniques: Western Blot, Expressing, Immunohistochemistry

Journal: Cell Reports Medicine

Article Title: Disruption of MerTK increases the efficacy of checkpoint inhibitor by enhancing ferroptosis and immune response in hepatocellular carcinoma

doi: 10.1016/j.xcrm.2024.101415

Figure Lengend Snippet:

Article Snippet: Briefly, following deparaffinization, rehydrating and antigen retrieval, primary antibodies were applied to slides, incubated at 4°C overnight, followed by incubation with secondary antibody (Dako Denmark A/S, Glostrup, Denmark) at 37°C for 30 min. SLC7A11 antibody (Proteintech, #26864-1-AP), MerTK antibody (SAB Signalway Antibody, #32851), p -MerTK antibody (SAB Signalway Antibody, #13320), Ki67 antibody (Abcam, #ab92742), CD3 antibody (Abcam, #ab16669), CD8 antibody (Abcam, #ab237709), CD11b antibody (Abcam, #ab52478), Gr1 antibody (Abcam, #ab25377) were used as primary antibodies.

Techniques: Virus, Recombinant, Lysis, Enzyme-linked Immunosorbent Assay, CCK-8 Assay, Isolation, Multiple Displacement Amplification, Expressing, shRNA, Software

Journal: Aging (Albany NY)

Article Title: Myeloid-specific knockout of SHP2 regulates PI3K/PLCγ signaling pathway to protect against early myocardial infarction injury

doi: 10.18632/aging.205096

Figure Lengend Snippet: SHP2 MAC-KO can affect the expression of PI3K/PLCγ signaling pathway, the expression of inflammatory factors and proteins related to phagocytic activity. ( A ) Protein band diagram of p-SHP2, p-PI3K, p-MerTK, p-PLCγ2, p-MyD88, p-NF-κB, TNFα, IFNγ, IL-1β; ( B ) Relative protein expression of p-SHP2, p-PI3K, p-MerTK, p-PLCγ2, p-MyD88, p-NF-κB, TNFα, IFNγ, IL-1β. **P<0.01.

Article Snippet: Then the membrane was transferred, and put in the TBST buffer containing 5% skim milk powder, blocked for 2 h. Primary antibodies p-SHP2(Abcam, ab62322, 1:1000), p-PI3K (CellSignaling, 17366, 1:1000), p-MerTK (CellSignaling, 44463,1:1000), p-PLCγ2 (CellSignaling, 3871, 1:1000), p-Myd88 (CellSignaling, 17366, 1:1000), p-NF-κB (CellSignaling, 3303,1:1000), TNFα (CellSignaling, 11948, 1:1000), IFNγ (CellSignaling, 8455, 1:1000), IL-1β (CellSignaling, 12703, 1:1000), and GADPH (CellSignaling, 5174, 1:1000) were added, and were incubated at room temperature for 1 hour, then incubated at 4° C overnight.

Techniques: Expressing, Activity Assay

Journal: Aging (Albany NY)

Article Title: Myeloid-specific knockout of SHP2 regulates PI3K/PLCγ signaling pathway to protect against early myocardial infarction injury

doi: 10.18632/aging.205096

Figure Lengend Snippet: SHP2 MAC-KO can mediate the PI3K/PLCγ signaling pathway to influence the expression of inflammatory factors and proteins related to phagocytic activity. ( A ) Protein band diagram of p-PI3K, p-MerTK, p-PLC Γ2, p-MyD88, p-NF-κb, TNF α, IFN γ, il-1β; ( B ) Relative protein expression amounts of p-PI3K, p-MerTK, p-PLCγ2, p-MyD88, p-NF-κb, TNF α, IL-1β. **P<0.01, ns P>0.05.

Article Snippet: Then the membrane was transferred, and put in the TBST buffer containing 5% skim milk powder, blocked for 2 h. Primary antibodies p-SHP2(Abcam, ab62322, 1:1000), p-PI3K (CellSignaling, 17366, 1:1000), p-MerTK (CellSignaling, 44463,1:1000), p-PLCγ2 (CellSignaling, 3871, 1:1000), p-Myd88 (CellSignaling, 17366, 1:1000), p-NF-κB (CellSignaling, 3303,1:1000), TNFα (CellSignaling, 11948, 1:1000), IFNγ (CellSignaling, 8455, 1:1000), IL-1β (CellSignaling, 12703, 1:1000), and GADPH (CellSignaling, 5174, 1:1000) were added, and were incubated at room temperature for 1 hour, then incubated at 4° C overnight.

Techniques: Expressing, Activity Assay