Journal: OncoTargets and therapy

Article Title: Dynamic Changes in Serum Markers and Their Utility in the Early Diagnosis of All Stages of Hepatitis B-Associated Hepatocellular Carcinoma

doi: 10.2147/OTT.S229835

Figure Lengend Snippet: The Clinicopathological Characteristics of the Study Population

Article Snippet: The following antibodies were spotted on aldehyde-coated microscope slides (Shanghai Baiao, China) by a GeSiM Nano-Plotter TM Micropipetting System (Radeberg, Germany): mouse monoclonal AFP antibody (1.25 mg/mL, Frpon Botech Inc, Shenzhen, China), mouse monoclonal GPC3 antibody (1 mg/mL, R&D Bio-techne, Minneapolis, USA), mouse monoclonal Golgi membrane protein 1 (GOLM1) antibody (0.5 mg/mL, Novus, Centennial, USA) and mouse monoclonal prothrombin factor II antibody (2.5mg/mL, Fujirebio Inc, Tokyo, Japan).

Techniques:

Journal: OncoTargets and therapy

Article Title: Dynamic Changes in Serum Markers and Their Utility in the Early Diagnosis of All Stages of Hepatitis B-Associated Hepatocellular Carcinoma

doi: 10.2147/OTT.S229835

Figure Lengend Snippet: The levels of serum AFP, GPC3, GP73 and DCP in each subgroup. ( A ) AFP. ( B ) GPC3. ( C ) GP73. ( D ) DCP. ( E ) Representative image of each subgroup. * P <0.05, ** P <0.01. Abbreviations: HCC, hepatocellular carcinoma; LC, liver cirrhosis; CHB, chronic hepatitis B virus infection; HC, healthy controls; AFP, α-fetoprotein; GPC3, glypican 3; GP73, golgi protein 73; DCP, des-γ-carboxy prothrombin; ns, no significance; int, intensity.

Article Snippet: The following antibodies were spotted on aldehyde-coated microscope slides (Shanghai Baiao, China) by a GeSiM Nano-Plotter TM Micropipetting System (Radeberg, Germany): mouse monoclonal AFP antibody (1.25 mg/mL, Frpon Botech Inc, Shenzhen, China), mouse monoclonal GPC3 antibody (1 mg/mL, R&D Bio-techne, Minneapolis, USA), mouse monoclonal Golgi membrane protein 1 (GOLM1) antibody (0.5 mg/mL, Novus, Centennial, USA) and mouse monoclonal prothrombin factor II antibody (2.5mg/mL, Fujirebio Inc, Tokyo, Japan).

Techniques: Virus, Infection

Journal: OncoTargets and therapy

Article Title: Dynamic Changes in Serum Markers and Their Utility in the Early Diagnosis of All Stages of Hepatitis B-Associated Hepatocellular Carcinoma

doi: 10.2147/OTT.S229835

Figure Lengend Snippet: The Value of Serum AFP, GPC3, GP73 and DCP in the Diagnosis of HCC (Including All HCC Patients)

Article Snippet: The following antibodies were spotted on aldehyde-coated microscope slides (Shanghai Baiao, China) by a GeSiM Nano-Plotter TM Micropipetting System (Radeberg, Germany): mouse monoclonal AFP antibody (1.25 mg/mL, Frpon Botech Inc, Shenzhen, China), mouse monoclonal GPC3 antibody (1 mg/mL, R&D Bio-techne, Minneapolis, USA), mouse monoclonal Golgi membrane protein 1 (GOLM1) antibody (0.5 mg/mL, Novus, Centennial, USA) and mouse monoclonal prothrombin factor II antibody (2.5mg/mL, Fujirebio Inc, Tokyo, Japan).

Techniques:

Journal: OncoTargets and therapy

Article Title: Dynamic Changes in Serum Markers and Their Utility in the Early Diagnosis of All Stages of Hepatitis B-Associated Hepatocellular Carcinoma

doi: 10.2147/OTT.S229835

Figure Lengend Snippet: Assessment of the diagnostic value of serum AFP, GPC3, GP73 and DCP in differentiating HBV-related HCC from controls. ( A ) All HCC vs LC, CHB, HC. ( B ) All HCC vs LC, CHB. ( C ) All HCC v s LC. ( D ) Very early and early stage HCC vs LC, CHB, HC. ( E ) Very early and early stage HCC vs LC, CHB. ( F ) Very early and early stage HCC vs LC. Abbreviations: HCC, hepatocellular carcinoma; LC, liver cirrhosis; CHB, chronic hepatitis B virus infection; HC, healthy controls; AFP, α-fetoprotein; GPC3, glypican 3; GP73, golgi protein 73; DCP, des-γ-carboxy prothrombin.

Article Snippet: The following antibodies were spotted on aldehyde-coated microscope slides (Shanghai Baiao, China) by a GeSiM Nano-Plotter TM Micropipetting System (Radeberg, Germany): mouse monoclonal AFP antibody (1.25 mg/mL, Frpon Botech Inc, Shenzhen, China), mouse monoclonal GPC3 antibody (1 mg/mL, R&D Bio-techne, Minneapolis, USA), mouse monoclonal Golgi membrane protein 1 (GOLM1) antibody (0.5 mg/mL, Novus, Centennial, USA) and mouse monoclonal prothrombin factor II antibody (2.5mg/mL, Fujirebio Inc, Tokyo, Japan).

Techniques: Diagnostic Assay, Virus, Infection

Journal: OncoTargets and therapy

Article Title: Dynamic Changes in Serum Markers and Their Utility in the Early Diagnosis of All Stages of Hepatitis B-Associated Hepatocellular Carcinoma

doi: 10.2147/OTT.S229835

Figure Lengend Snippet: The Value of Serum AFP, GPC3, GP73 and DCP in the Early Diagnosis of HCC (Including the Very Early HCC)

Article Snippet: The following antibodies were spotted on aldehyde-coated microscope slides (Shanghai Baiao, China) by a GeSiM Nano-Plotter TM Micropipetting System (Radeberg, Germany): mouse monoclonal AFP antibody (1.25 mg/mL, Frpon Botech Inc, Shenzhen, China), mouse monoclonal GPC3 antibody (1 mg/mL, R&D Bio-techne, Minneapolis, USA), mouse monoclonal Golgi membrane protein 1 (GOLM1) antibody (0.5 mg/mL, Novus, Centennial, USA) and mouse monoclonal prothrombin factor II antibody (2.5mg/mL, Fujirebio Inc, Tokyo, Japan).

Techniques:

Journal: OncoTargets and therapy

Article Title: Dynamic Changes in Serum Markers and Their Utility in the Early Diagnosis of All Stages of Hepatitis B-Associated Hepatocellular Carcinoma

doi: 10.2147/OTT.S229835

Figure Lengend Snippet: The Value of Serum AFP, GPC3, GP73 and DCP in the Very Early Diagnosis of HCC

Article Snippet: The following antibodies were spotted on aldehyde-coated microscope slides (Shanghai Baiao, China) by a GeSiM Nano-Plotter TM Micropipetting System (Radeberg, Germany): mouse monoclonal AFP antibody (1.25 mg/mL, Frpon Botech Inc, Shenzhen, China), mouse monoclonal GPC3 antibody (1 mg/mL, R&D Bio-techne, Minneapolis, USA), mouse monoclonal Golgi membrane protein 1 (GOLM1) antibody (0.5 mg/mL, Novus, Centennial, USA) and mouse monoclonal prothrombin factor II antibody (2.5mg/mL, Fujirebio Inc, Tokyo, Japan).

Techniques:

Journal: OncoTargets and therapy

Article Title: Dynamic Changes in Serum Markers and Their Utility in the Early Diagnosis of All Stages of Hepatitis B-Associated Hepatocellular Carcinoma

doi: 10.2147/OTT.S229835

Figure Lengend Snippet: The value of serum AFP, GPC3, GP73 and DCP in the very early diagnosis of HBV-related HCC. ( A ) Very early stage HCC vs LC, CHB, HC. ( B ) Very early stage vs LC, CHB. ( C ) Very early stage vs LC. Abbreviations: HCC, hepatocellular carcinoma; LC, liver cirrhosis; CHB, chronic hepatitis B virus infection; HC, healthy controls; AFP, α-fetoprotein; GPC3, glypican 3; GP73, golgi protein 73; DCP, des-γ-carboxy prothrombin.

Article Snippet: The following antibodies were spotted on aldehyde-coated microscope slides (Shanghai Baiao, China) by a GeSiM Nano-Plotter TM Micropipetting System (Radeberg, Germany): mouse monoclonal AFP antibody (1.25 mg/mL, Frpon Botech Inc, Shenzhen, China), mouse monoclonal GPC3 antibody (1 mg/mL, R&D Bio-techne, Minneapolis, USA), mouse monoclonal Golgi membrane protein 1 (GOLM1) antibody (0.5 mg/mL, Novus, Centennial, USA) and mouse monoclonal prothrombin factor II antibody (2.5mg/mL, Fujirebio Inc, Tokyo, Japan).

Techniques: Virus, Infection

Journal: Cell Reports Medicine

Article Title: In vivo vulnerabilities to GPX4 and HDAC inhibitors in drug-persistent versus drug-resistant BRAF V600E lung adenocarcinoma

doi: 10.1016/j.xcrm.2024.101663

Figure Lengend Snippet:

Article Snippet: Rabbit monoclonal anti-CD200 (clone E519V) , Cell signaling Technology , Cat#23451; RRID: AB_3102027.

Techniques: Recombinant, Glutathione Assay, Flow Cytometry, shRNA, Control, Plasmid Preparation, Software

Journal: Heliyon

Article Title: CD200-CD200R affects cisplatin and paclitaxel sensitivity by regulating cathepsin K-mediated p65 NF-κB signaling in cervical cancer

doi: 10.1016/j.heliyon.2023.e19220

Figure Lengend Snippet: CD200 and CTSK expression in cervical cancer tissues. (A) The GEPIA online website was used to analyze CD200 and CTSK expression in cervical squamous cell carcinoma and endocervical adenocarcinoma (CESC) tissues and normal tissues. (B) The correlation of CD200 and CTSK in the CESC. CD200 (C) and CTSK (D) expression in tumor tissues and para-cancerous tissues was measured using immunohistochemistry. *P < 0.05, **P < 0.01. Representative images of CD200 and CTSK expression in tissues are provided in Supplemental and .

Article Snippet: Diluted primary CD200 rabbit antibody (#23451S, 1:200, Cell Signaling Technology, Shanghai, China), CD200R rabbit-antibody (1:400, #DF4715, Affinity, China), Ki-67 rabbit-antibody (1:400, #9027, Cell Signaling Technology) and CTSK rabbit antibody (1:400, #DF6614, Affinity, China) were incubated with the section at 4 °C for overnight.

Techniques: Expressing, Immunohistochemistry

Journal: Heliyon

Article Title: CD200-CD200R affects cisplatin and paclitaxel sensitivity by regulating cathepsin K-mediated p65 NF-κB signaling in cervical cancer

doi: 10.1016/j.heliyon.2023.e19220

Figure Lengend Snippet: Effects of CD200 on the viability and invasion of HeLa and Siha cells. HeLa and Siha cells were stably transfected with 2 μg of control plasmid or PUNO1-hCD200. (A) Cell viabilities were analyzed by CCK-8. (B) Cell invasion was measured by Transwell assay. Scale = 50 μm. The results of CD200 transfection in HeLa and SiHa cells are provided in Supplemental and . NS: no significance.

Article Snippet: Diluted primary CD200 rabbit antibody (#23451S, 1:200, Cell Signaling Technology, Shanghai, China), CD200R rabbit-antibody (1:400, #DF4715, Affinity, China), Ki-67 rabbit-antibody (1:400, #9027, Cell Signaling Technology) and CTSK rabbit antibody (1:400, #DF6614, Affinity, China) were incubated with the section at 4 °C for overnight.

Techniques: Stable Transfection, Transfection, Control, Plasmid Preparation, CCK-8 Assay, Transwell Assay

Journal: Heliyon

Article Title: CD200-CD200R affects cisplatin and paclitaxel sensitivity by regulating cathepsin K-mediated p65 NF-κB signaling in cervical cancer

doi: 10.1016/j.heliyon.2023.e19220

Figure Lengend Snippet: CD200-CD200R regulated the CTSK-mediated p65 NF-κB pathway in cervical cancer cells. Cancer cells were cocultured without or with M2 macrophage-like THP-1 cells. Expression levels of CTSK and p-p65 NF-κB/p65 NF-κB protein in HeLa (A) and Siha (B) were measured by Western blotting. The full length of blots was provided in Supplemental materials. *P < 0.05, **P < 0.01.

Article Snippet: Diluted primary CD200 rabbit antibody (#23451S, 1:200, Cell Signaling Technology, Shanghai, China), CD200R rabbit-antibody (1:400, #DF4715, Affinity, China), Ki-67 rabbit-antibody (1:400, #9027, Cell Signaling Technology) and CTSK rabbit antibody (1:400, #DF6614, Affinity, China) were incubated with the section at 4 °C for overnight.

Techniques: Expressing, Western Blot

Journal: Heliyon

Article Title: CD200-CD200R affects cisplatin and paclitaxel sensitivity by regulating cathepsin K-mediated p65 NF-κB signaling in cervical cancer

doi: 10.1016/j.heliyon.2023.e19220

Figure Lengend Snippet: CD200-CD200R is involved in the response to cisplatin or paclitaxel in cervical cancer. (A) Schematic diagram shows a coculture system of M2 macrophage-like THP-1 cells and cancer cells. Cell viabilities of HeLa (B) and Siha (C) cells were analyzed by CCK-8. Cell invasion of HeLa (D) and Siha (E) cells was analyzed by Transwell assay. The images of Transwell assay are provided in Supplemental . *P < 0.05, **P < 0.01.

Article Snippet: Diluted primary CD200 rabbit antibody (#23451S, 1:200, Cell Signaling Technology, Shanghai, China), CD200R rabbit-antibody (1:400, #DF4715, Affinity, China), Ki-67 rabbit-antibody (1:400, #9027, Cell Signaling Technology) and CTSK rabbit antibody (1:400, #DF6614, Affinity, China) were incubated with the section at 4 °C for overnight.

Techniques: CCK-8 Assay, Transwell Assay

Journal: Heliyon

Article Title: CD200-CD200R affects cisplatin and paclitaxel sensitivity by regulating cathepsin K-mediated p65 NF-κB signaling in cervical cancer

doi: 10.1016/j.heliyon.2023.e19220

Figure Lengend Snippet: CD200-CD200R affects the chemosensitivity of cervical cancer cells through regulating the CTSK-mediated p65 NF-κB pathway. Cancer cells were treated with 0.1 μg/mL rCTSK as a control to compare the effects of C200-CD200R on resistance to cisplatin or paclitaxel. Cell viabilities of HeLa (A) and Siha (B) were analyzed by CCK-8. Expression of p-p65 NF-κB/p65 NF-κB protein in HeLa (C) and Siha (D) was analyzed by Western blotting. The full length of blots was provided in Supplemental materials. **P < 0.01.

Article Snippet: Diluted primary CD200 rabbit antibody (#23451S, 1:200, Cell Signaling Technology, Shanghai, China), CD200R rabbit-antibody (1:400, #DF4715, Affinity, China), Ki-67 rabbit-antibody (1:400, #9027, Cell Signaling Technology) and CTSK rabbit antibody (1:400, #DF6614, Affinity, China) were incubated with the section at 4 °C for overnight.

Techniques: Control, CCK-8 Assay, Expressing, Western Blot

Journal: Heliyon

Article Title: CD200-CD200R affects cisplatin and paclitaxel sensitivity by regulating cathepsin K-mediated p65 NF-κB signaling in cervical cancer

doi: 10.1016/j.heliyon.2023.e19220

Figure Lengend Snippet: CD200-C200R affects the paclitaxel sensitivity in mouse xenografts through regulating CTSK. Stably transfected plasmid or CD200 HeLa cells were used to establish xenografts. After injection for 7 days, 3.6 mg/kg MK-0822 (CTSK inhibitor) was orally treated every week, and 5 mg/kg paclitaxel was administrated by tail vein twice a week. (A) Tumor size was measured every week. (B) Images of xenografts and tumor weight. (C) Positive expression of Ki-67 in tumor tissues of each group. Scale = 50 μm. NS: no significance. **P < 0.01.

Article Snippet: Diluted primary CD200 rabbit antibody (#23451S, 1:200, Cell Signaling Technology, Shanghai, China), CD200R rabbit-antibody (1:400, #DF4715, Affinity, China), Ki-67 rabbit-antibody (1:400, #9027, Cell Signaling Technology) and CTSK rabbit antibody (1:400, #DF6614, Affinity, China) were incubated with the section at 4 °C for overnight.

Techniques: Stable Transfection, Transfection, Plasmid Preparation, Injection, Expressing

Journal: Heliyon

Article Title: CD200-CD200R affects cisplatin and paclitaxel sensitivity by regulating cathepsin K-mediated p65 NF-κB signaling in cervical cancer

doi: 10.1016/j.heliyon.2023.e19220

Figure Lengend Snippet: A CD200-CD200R mediated feedback loop between cancer cells and M2-tumor-associated macrophages (TAMs) in cervical cancer.

Article Snippet: Diluted primary CD200 rabbit antibody (#23451S, 1:200, Cell Signaling Technology, Shanghai, China), CD200R rabbit-antibody (1:400, #DF4715, Affinity, China), Ki-67 rabbit-antibody (1:400, #9027, Cell Signaling Technology) and CTSK rabbit antibody (1:400, #DF6614, Affinity, China) were incubated with the section at 4 °C for overnight.

Techniques:

Journal: Clinical and Translational Medicine

Article Title: PD1 hi CD200 hi CD4 + exhausted T cell increase immunotherapy resistance and tumour progression by promoting epithelial–mesenchymal transition in bladder cancer

doi: 10.1002/ctm2.1303

Figure Lengend Snippet: Two subclusters of CD4 + exhausted T cells were defined in the tumour microenvironment using the pancancer CD4 + T cell atlas: (A) the UMAP plot of the subclusters of CD4 + T cells integrated from nine tumour types. Each dot indicated a single cell. Colour‐coded for the cell type; (B) the dot plot showing the particular genes for each subcluster of CD4 + T cells; (C) tissue prevalence estimated by Ro/e score in pan‐CD4 + T cells; (D) the violin plot showing exhausted‐related genes expression in each cluster; (E) volcano map showing differentially expressed genes between CD4_ex2 and CD4_ex1 subclusters (ex2: logFC > .3 and log p ‐value >2; ex1: logFC < −.3 and log p ‐value >2); (F) the UMAP plot showing the IFNG expression in pan‐CD4 + T cells; (G) the barplot showing the pathway enrichment of CD4_ex1 subcluster; (H) the UMAP plot showing the CD200 expression in pan‐CD4 + T cells; (I) Barplot** showing the correlation between T cell exhausted‐related genes and CD200 in The Cancer Genome Atlas (TCGA)‐BLCA; (J) multiplex immunofluorescence staining was performed for DAPI (blue), CD4 (white), PD1 (red) and CD200 (green); each stain was shown separately and merged. The circles represent the PD1 hi CD200 hi CD4 + exhausted T cells. The field of view in the square is the reduced field of view.

Article Snippet: Following those procedures, horseradish peroxidase (HRP)‐conjugated secondary antibodies were incubated, and tyramide signal amplification was carried out after sequentially applying different primary antibodies, including PD1 (CST, Boston, MA, USA), CD4 (CST, Boston, MA, USA), CD200 (CST, Boston, MA, USA), CD31 (CST, Boston, MA, USA), GSA6 (CST, Boston, MA, USA) and IFNG (Novus, USA).

Techniques: Expressing, Multiplex Assay, Immunofluorescence, Staining

Journal: Clinical and Translational Medicine

Article Title: PD1 hi CD200 hi CD4 + exhausted T cell increase immunotherapy resistance and tumour progression by promoting epithelial–mesenchymal transition in bladder cancer

doi: 10.1002/ctm2.1303

Figure Lengend Snippet: The PD‐1 hi CD200 hi CD4 + exhausted T cells predict a poor response to immunotherapy: (A) overall survival of patients with immunotherapy in PDCD1 low , PDCD1 hi CD200 low and PDCD1 hi CD200 hi groups in the GSE176307 and IMvigor210 cohorts; (B) barplots showing the proportion of responders among PDCD1 low , PDCD1 hi CD200 low and PDCD1 hi CD200 hi groups in the GSE176307 and IMvigor210 cohorts; (C) violin plots showing the estimation of the abundance of CD8 T cells and M1 macrophages in the PDCD1 low , PDCD1 hi CD200 low and PDCD1 hi CD200 hi groups using CIBERSORT algorithm; (D) the expression of IFN‐γ between the PD‐1 hi CD200 low CD4 − T cells and PD‐1 hi CD200 hi CD4 − T cells was measured by intracellular flow cytometry. Summary data of IFN‐γ staining (right) are presented as mean ± SD ( n = 3), ** p < 0.01, *** p <0.001; (E) multiplex immunofluorescence staining was performed for DAPI (blue), CD4 (white), PD1 (red), CD200 (green) and IFNG (yellow). The circles represent the PD1 hi CD200 hi CD4 + exhausted T cells. The arrows represent the IFNG.

Article Snippet: Following those procedures, horseradish peroxidase (HRP)‐conjugated secondary antibodies were incubated, and tyramide signal amplification was carried out after sequentially applying different primary antibodies, including PD1 (CST, Boston, MA, USA), CD4 (CST, Boston, MA, USA), CD200 (CST, Boston, MA, USA), CD31 (CST, Boston, MA, USA), GSA6 (CST, Boston, MA, USA) and IFNG (Novus, USA).

Techniques: Expressing, Flow Cytometry, Staining, Multiplex Assay, Immunofluorescence

Journal: Clinical and Translational Medicine

Article Title: PD1 hi CD200 hi CD4 + exhausted T cell increase immunotherapy resistance and tumour progression by promoting epithelial–mesenchymal transition in bladder cancer

doi: 10.1002/ctm2.1303

Figure Lengend Snippet: The PD‐1 hi CD200 hi CD4 exhausted T cells recruit tip cells to promote angiogenesis: (A) boxplot showing that estimation of the abundance of endothelial cells in the PDCD1 low , PDCD1 hi CD200 low and PDCD1 hi CD200 hi groups using xCell_counter algorithm; (B) volcano map showing differentially expressed genes between PDCD1 hi CD200 hi and PDCD1 hi CD200 low groups (PDCD1 hi CD200 hi : logFC >0.3 and log p ‐value >2; PDCD1 hi CD200 low : logFC < −0.3 and log p ‐value >2). Angiogenesis pathway‐related genes were highlighted; (C) gene set enrichment analysis (GSEA) showing that the angiogenesis pathway was overrepresented in PDCD1hi CD200hi groups; (D and E) fluorescence micrograph of tube formation by HMEC1 (top) and network mask (bottom). Branch points and capillary length were analysed to evaluate angiogenic activity. The bars represent mean ± SD ( n = 3), ** p <0.01, *** p < 0.001; (F) multiplex immunofluorescence staining was performed for DAPI (blue), CD4 (white), PD1 (red), CD200 (green) and CD31 (orange). The circles represent the PD1 hi CD200 hi CD4 + exhausted T cells. The arrows represent the CD31; (G) spatial distribution of CD31 + cells around PD‐1 hi CD200 hi CD4 − T cells. The histogram showing the distribution of distances from each the PD‐1 hi CD200 hi CD4 − T cells the nearest CD31 + cell; (H) the UMAP plot showing the clusters of endothelial cells. Each dot indicates a single cell; (I) the boxplot showing the angiogenesis‐related genes expression in subclusters of endothelial cells; (J) the aggregated cell–cell communication analysis of CD4_ex2 and endothelial cells in BLCA by CellChat; (K) a flow diagram showing the information flow of metabolite–sensor communications from CD4 + T cells to endothelial cells through metabolites and sensors. The size of dots represents the number of connections. The lines connect the sender, metabolite, sensor and receiver. The colour of the line indicates the −log10( p ‐value), and the width of lines represents the communication score.

Article Snippet: Following those procedures, horseradish peroxidase (HRP)‐conjugated secondary antibodies were incubated, and tyramide signal amplification was carried out after sequentially applying different primary antibodies, including PD1 (CST, Boston, MA, USA), CD4 (CST, Boston, MA, USA), CD200 (CST, Boston, MA, USA), CD31 (CST, Boston, MA, USA), GSA6 (CST, Boston, MA, USA) and IFNG (Novus, USA).

Techniques: Fluorescence, Activity Assay, Multiplex Assay, Immunofluorescence, Staining, Expressing

Journal: Clinical and Translational Medicine

Article Title: PD1 hi CD200 hi CD4 + exhausted T cell increase immunotherapy resistance and tumour progression by promoting epithelial–mesenchymal transition in bladder cancer

doi: 10.1002/ctm2.1303

Figure Lengend Snippet: The PD1 hi CD200 hi CD4 exhausted T cells promote epithelial–mesenchymal transition (EMT): (A) boxplot showing that estimation of the abundance of epithelial and fibroblast cells in the PDCD1 low , PDCD1 hi CD200 low and PDCD1 hi CD200 hi groups using xCel algorithm; (B) volcano map showing differentially expressed genes between PDCD1 hi CD200 hi and PDCD1 hi CD200 low groups (PDCD1 hi CD200 hi : logFC > 0.3 and log p ‐value >2; PDCD1 hi CD200 low : logFC < −0.3 and log p ‐value >2). The EMT pathway‐related genes were highlighted; (C) correlation of CD200 expression with the expression of transcription factor of EMT. The colour indicates the Spearman correlation coefficient; (D) gene set enrichment analysis (GSEA) showing that the EMT pathway was overrepresented in PDCD1 hi CD200 hi groups; (E) the invasion of T24 and UMUC3 cells was detected by transwell assay after co‐cultured with the PD1 hi CD200 hi CD4 + or PD1 hi CD200 low CD4 + exhausted T cells. Results are presented as mean ± SD ( n = 3), ** p < 0.01, *** p < 0.001; (F) the UMAP plot showing the clusters of epithelial cells. Each dot indicated a single cell. Colour‐coded for the cell type; (G) the boxplot showing the EMT‐related genes expression in clusters of epithelial cells; (H) analysis of simulated differentiation trajectories of three epithelial cell subclusters (Epi_CXCL1, Epi_OLFM4, and Epi_COL1A2) in BLCA. Each dot corresponds to a cell, and each colour represents an epithelial cell subcluster; (I) the RNA velocity analysis graph reflected the evolutionary relationship among the three epithelial cell subclusters; (J) the significantly enriched ligand–receptor pairs between epithelial cells and CD4_ex2 subcluster T cells in primary tumour; (K) multiplex immunofluorescence staining was performed for DAPI (blue), CD4 (white), PD1 (green), CD200 (red) and GAS6 (lawngreen). The circles represent the PD1 hi CD200 hi CD4 + exhausted T cells. The arrows represent the GAS6; (L) spatial distribution of GAS6 + cells around PD‐1 hi CD200 hi CD4 + T cells, DAPI (blue), CD4 (white), PD1 (red), CD200 (green) and GAS6 (lawngreen); (M) overall survival between the high and low expressions of GAS6 or AXL groups in The Cancer Genome Atlas (TCGA)‐BLCA; (N) scatter plot showing the correlation between the expression of CD200 and GAS6 in TCGA‐BLCA; (O) barplot showing the correlation between the expression of EMT transcription factor gene, EMT signature and GAS6 in TCGA‐BLCA.

Article Snippet: Following those procedures, horseradish peroxidase (HRP)‐conjugated secondary antibodies were incubated, and tyramide signal amplification was carried out after sequentially applying different primary antibodies, including PD1 (CST, Boston, MA, USA), CD4 (CST, Boston, MA, USA), CD200 (CST, Boston, MA, USA), CD31 (CST, Boston, MA, USA), GSA6 (CST, Boston, MA, USA) and IFNG (Novus, USA).

Techniques: Expressing, Transwell Assay, Cell Culture, Multiplex Assay, Immunofluorescence, Staining

Journal: Clinical and Translational Medicine

Article Title: PD1 hi CD200 hi CD4 + exhausted T cell increase immunotherapy resistance and tumour progression by promoting epithelial–mesenchymal transition in bladder cancer

doi: 10.1002/ctm2.1303

Figure Lengend Snippet: PD1 hi CD200 hi CD4 + exhausted T cells negatively correlate with the response to ICIs and positively recruit vascular cells in vivo: (A) schematic diagram of mice models; (B) schematic diagram of administration cycles in mice; (C) photographs of tumours dissected out from the three mice groups; (D) tumour volumes were measured once every days, and growth curves of tumours were shown; (E) multiplex immunofluorescence staining was performed for DAPI (blue), Cd4 (white), Pd1 (red) and Cd200 (green); each stain was shown separately and merged. Histograms show the percentages of Pd1hi Cd200hi Cd4 + exhausted T cells in anti‐PD1‐responsive and non‐responsive groups; (F) multiplex immunofluorescence staining was performed for DAPI (blue), Cd4 (white), Pd1 (red) and Cd200 (green); each stain was shown separately and merged. Histograms show the percentages of Pd1hi Cd200hi Cd4 + exhausted T cells in anti‐PD1‐responsive and non‐responsive groups; (G) multiplex immunofluorescence staining was performed for DAPI (blue), Cd4 (white), Pd1 (red), Cd200 (green) and Ifng (yellow); each stain was shown separately and merged. Histograms show the percentages of Ifng in anti‐PD1‐responsive and non‐responsive groups; (G) multiplex immunofluorescence staining was performed for DAPI (blue), Cd4 (white), Pd1 (red), Cd200 (green) and Cd31 (yellow); each stain was shown separately and merged. Histograms show the percentages of Cd31 in anti‐PD1‐responsive and non‐responsive groups; (H) multiplex immunofluorescence staining was performed for DAPI (blue), Cd4 (white), Pd1 (red), Cd200 (green) and Gas6 (yellow); each stain was shown separately and merged. Histograms show the percentages of Gas6 in anti‐PD1‐responsive and non‐responsive groups.

Article Snippet: Following those procedures, horseradish peroxidase (HRP)‐conjugated secondary antibodies were incubated, and tyramide signal amplification was carried out after sequentially applying different primary antibodies, including PD1 (CST, Boston, MA, USA), CD4 (CST, Boston, MA, USA), CD200 (CST, Boston, MA, USA), CD31 (CST, Boston, MA, USA), GSA6 (CST, Boston, MA, USA) and IFNG (Novus, USA).

Techniques: In Vivo, Multiplex Assay, Immunofluorescence, Staining