Journal: Journal of Cellular and Molecular Medicine

Article Title: Transcriptomic analysis reveals cell apoptotic signature modified by heparanase in melanoma cells

doi: 10.1111/jcmm.14349

Figure Lengend Snippet: Gene ontology classification of differentially expressed genes regulated by silencing of heparanase gene (HPSE) expression. (A) Gene ontology (GO) analysis of up‐ and (B) down‐regulated genes after silencing of HPSE in MDA‐MB‐435s cells by terms of biological process and cellular component. X ‐axis i ndicates functional fold enrichment calculated by binomial test, P < 0.01. (C) Listing of an array of 28 pro‐apoptotic genes classified by GO term positive regulation of cell death and apoptotic process. Y ‐axis indicates fold change comparing HPSE silenced cells with control cells. Dashed line indicates 1.5‐fold change. (D) Validation of expression of the 28 pro‐apoptotic genes by real‐time PCR. n = 3 biological repeats, * indicates the selected genes for further validation by Western blots. Dashed line indicates 1.5‐fold change. (E) Validation of up‐regulation of selected genes including CYR61, EGR1 and TNFRSF12A on protein level by Western blots. N = 3 biological repeats, representative blots are shown

Article Snippet: Anti‐EGR1 (AF2818), CYR61 (MAB4055), TNFRSF12 (MAB1199) were from R&D Systems (Abingdon, UK), and anti‐GAPDH (AM4300) from Ambion.

Techniques: Expressing, Functional Assay, Control, Biomarker Discovery, Real-time Polymerase Chain Reaction, Western Blot

Journal: Cancer Science

Article Title: Cysteine‐rich protein 61 regulates the chemosensitivity of chronic myeloid leukemia to imatinib mesylate through the nuclear factor kappa B/Bcl‐2 pathway

doi: 10.1111/cas.14083

Figure Lengend Snippet: Cysteine‐rich protein 61 (Cyr61) levels are upregulated in plasma and bone marrow ( BM ) samples from patients with CML . A, Left panel: Levels of Cyr61 in the plasma from CML patients (n = 36) and normal plasma from age‐matched healthy individuals ( CON ; n = 66) were detected by ELISA . Right panel: Levels of Cyr61 in the BM supernatant from CML patients (n = 33) and the normal BM supernatant from age‐matched healthy transplant donors (n = 11) were detected by ELISA . B, Levels of Cyr61 in the plasma from CML patients in blast crisis ( BC ) (n = 5) and in chronic phase ( CP ) (n = 31) were detected by ELISA . Right panel: Levels of Cyr61 in the marrow from CML patients in BC (n = 4) and in CP (n = 29) were detected by ELISA . C, Relative levels of Cyr61 mRNA in a T acute lymphoblastic leukemia ( ALL ) cell line (Jurkat), B ALL cell line (Nalm‐6), and CML cell line (K562) were detected by qRT ‐ PCR , and the level of Cyr61 mRNA in Nalm‐6 cells was taken as the control to calculate the relative expression of Cyr61 in Jurkat and K562 cells. D, Levels of Cyr61 protein in Jurkat, Nalm‐6, and K562 cells were detected by western blotting. Band intensity of Cyr61 was quantified by densitometry and normalized to GAPDH . E, Concentration of Cyr61 in the culture supernatant of Jurkat, Nalm‐6, and K562 cells was detected by ELISA . Data represent mean ± SEM of at least 3 independent experiments. *P < 0.05, ** P < 0.01

Article Snippet: Concentrations of Cyr61 in the plasma and BM from CML patients were quantitated using the human Cyr61 ELISA kit (R&D Systems, Minneapolis, MN, USA) according to the manufacturer's instructions.

Techniques: Clinical Proteomics, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR, Control, Expressing, Western Blot, Concentration Assay

Journal: Cancer Science

Article Title: Cysteine‐rich protein 61 regulates the chemosensitivity of chronic myeloid leukemia to imatinib mesylate through the nuclear factor kappa B/Bcl‐2 pathway

doi: 10.1111/cas.14083

Figure Lengend Snippet: Role of cysteine‐rich protein 61 (Cyr61) in the chemosensitivity of CML cells to imatinib mesylate ( IM ). A, K562 cells were treated with Cyr61 (125, 250, 500, 1000 ng/mL) for 24 h, and then treated with 0.5 μmol/L IM for 24 h; the percentages of apoptotic K562 cells were determined by flow cytometric analysis. Average percentage of apoptotic cells is shown. B, K562 cells were collected, incubated with Cyr61 (1000 ng/ mL ) preincubated with the antihuman Cyr61 093G9 monoclonal antibody (5000 pg/ mL ) or murine isotype‐matched antibody (Con‐IgG) (5000 pg/ mL ), and then treated with 0.5 μmol/L IM for 24 h. Cell apoptosis was determined by flow cytometric analysis. C, Cyr61 knockdown by shCyr61 or sh NC (negative control) in K562 cells. Endogenous Cyr61 expression is shown in the upper panel, whereas the secreted Cyr61 level in culture medium was determined by ELISA and shown in the lower panel. D, Ratio of apoptotic K562‐shCyr61 and K562‐sh NC cells was determined by flow cytometry at 24 h post‐treatment with or without 0.5 μmol/L IM . E, K562 cells were incubated with BM supernatants from a mixture of different CML patients (Cyr61 concentration was 243 pg/ mL ) with preincubation with 1000 pg/ mL 093G9 antibody or murine isotype‐matched antibody (Con‐IgG) for 2 h, and then treated with 0.5 μmol/L IM for 24 h. F, Human CML cell line KCL 22 cells were treated with Cyr61 (1000 ng/mL) for 24 h and then treated with 0.5 μmol/L IM for 24 h; the percentages of apoptotic cells were determined by flow cytometric analysis. G, Primary leukemic cells from three patients with CP CML were isolated and treated with exogenous recombinant human Cyr61 (1000 ng/mL) for 24 h, and then treated with 0.5 μmol/L IM for 24 h. Data represent mean ± SEM of at least 3 independent experiments. * P < 0.05, ** P < 0.01

Article Snippet: Concentrations of Cyr61 in the plasma and BM from CML patients were quantitated using the human Cyr61 ELISA kit (R&D Systems, Minneapolis, MN, USA) according to the manufacturer's instructions.

Techniques: Incubation, Knockdown, Negative Control, Expressing, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Concentration Assay, Isolation, Recombinant

Journal: Cancer Science

Article Title: Cysteine‐rich protein 61 regulates the chemosensitivity of chronic myeloid leukemia to imatinib mesylate through the nuclear factor kappa B/Bcl‐2 pathway

doi: 10.1111/cas.14083

Figure Lengend Snippet: Cysteine‐rich protein 61 (Cyr61) activates Bcl‐2 transcription in CML cells. A, Left panel: Bcl‐2, Bcl‐ xL , XIAP and Survivin mRNA expression in K562 cells treated by 1000 ng/mL Cyr61 for 8 h was detected by real‐time PCR . Right panel: Bcl‐2, Bcl‐ xL , XIAP and Survivin mRNA expression in K562‐shCyr61 cells and K562‐sh NC cells was detected by real‐time PCR . B, Left panel: Bcl‐2 protein in K562 cells stimulated by 1000 ng/mL Cyr61 for 48 h was detected by western blotting. Right panel: Bcl‐2 protein in K562‐shCyr61 cells and K562‐sh NC cells was detected by western blotting. The band intensity of Bcl‐2 was quantified by densitometry and normalized to GAPDH . C, K562 cells were treated with Cyr61 (1000 ng/mL), ABT 199 (1 μmol/L) (specific Bcl‐2 inhibitor), Cyr61 + ABT 199, or ABT 199 for 24 h, and then treated with 0.5 μmol/L imatinib mesylate ( IM ) for 24 h. Percentages of apoptotic K562 cells were determined by flow cytometric analysis. Data represent the mean ± SEM of at least 3 independent experiments. * P < 0.05, ** P < 0.01

Article Snippet: Concentrations of Cyr61 in the plasma and BM from CML patients were quantitated using the human Cyr61 ELISA kit (R&D Systems, Minneapolis, MN, USA) according to the manufacturer's instructions.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Western Blot

Journal: Cancer Science

Article Title: Cysteine‐rich protein 61 regulates the chemosensitivity of chronic myeloid leukemia to imatinib mesylate through the nuclear factor kappa B/Bcl‐2 pathway

doi: 10.1111/cas.14083

Figure Lengend Snippet: Cysteine‐rich protein 61 (Cyr61) inhibits imatinib mesylate ( IM )‐induced apoptosis through the nuclear factor kappa B ( NF ‐κB) signaling pathway. A, Effect of the inhibitors of signaling pathways on Cyr61 decreased CML cell apoptosis induced by IM . K562 cells were treated with 20 μmol/L LY 294002, 1 μmol/L PD 98059 or 4 μmol/L PDTC in combination with Cyr61 (1000 ng/mL) for 24 h and then treated with 0.5 μmol/L IM for 24 h; the percentages of apoptotic K562 cells were determined by flow cytometric analysis. B, NF ‐κB phosphorylation was detected by western blotting. Lane 1: stimulation of K562 cells with 0.5 μmol/L IM for 10 min; lane 2: stimulation of K562 cells with 1000 ng/ mL Cyr61 + 0.5 μmol/L IM for 10 min. C, K562 cells were treated with 1000 ng/ mL Cyr61 in combination with or without 4 μmol/L PDTC for 24 h, and then treated with 0.5 μmol/L IM for 24 h. Protein levels of Bcl‐2 in K562 cells were detected by western blotting. D, K562‐shCyr61 cells and K562‐sh NC cells were treated with 0.5 μmol/L IM for 24 h. Left panel: NF ‐κB phosphorylation was detected by western blotting. Right panel: Bcl‐2 protein levels in K562 cells were detected by western blotting. Band intensity of Bcl‐2 was quantified by densitometry and normalized to GAPDH . Data represent the mean ± SEM of at least 3 independent experiments. * P < 0.05, ** P < 0.01

Article Snippet: Concentrations of Cyr61 in the plasma and BM from CML patients were quantitated using the human Cyr61 ELISA kit (R&D Systems, Minneapolis, MN, USA) according to the manufacturer's instructions.

Techniques: Protein-Protein interactions, Phospho-proteomics, Western Blot

Journal: Cancer Science

Article Title: Cysteine‐rich protein 61 regulates the chemosensitivity of chronic myeloid leukemia to imatinib mesylate through the nuclear factor kappa B/Bcl‐2 pathway

doi: 10.1111/cas.14083

Figure Lengend Snippet: Inhibition of cysteine‐rich protein 61 (Cyr61) restores the chemosensitivity of CML cells to imatinib mesylate ( IM ) in vivo. NOD / SCID mice bearing s.c. K562‐shCyr61 or control K562‐sh NC cell xenografts (n = 6) were injected i.p. with IM or normal saline ( NS ) daily from 10 d after inoculation with 1.0 × 10 7 tumor cells for 20 d, and then the mice were killed. A, Representative images of tumors are shown. B, Tumor weight is shown. C, Average percentage of tumor volume is shown. * P < 0.05

Article Snippet: Concentrations of Cyr61 in the plasma and BM from CML patients were quantitated using the human Cyr61 ELISA kit (R&D Systems, Minneapolis, MN, USA) according to the manufacturer's instructions.

Techniques: Inhibition, In Vivo, Control, Injection, Saline

Journal: Cancer Science

Article Title: Cysteine‐rich protein 61 regulates the chemosensitivity of chronic myeloid leukemia to imatinib mesylate through the nuclear factor kappa B/Bcl‐2 pathway

doi: 10.1111/cas.14083

Figure Lengend Snippet: Proposed signaling pathway by which cysteine‐rich protein 61 (Cyr61) reduces imatinib mesylate ( IM )‐induced CML cell apoptosis. Increased Cyr61 in the bone marrow from CML patients stimulates nuclear factor kappa B ( NF ‐κB) phosphorylation, then upregulates Bcl‐2 production, finally leading to decrease of IM ‐induced CML cell apoptosis and insensitivity to IM

Article Snippet: Concentrations of Cyr61 in the plasma and BM from CML patients were quantitated using the human Cyr61 ELISA kit (R&D Systems, Minneapolis, MN, USA) according to the manufacturer's instructions.

Techniques: Phospho-proteomics

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Loss of Brg1 and Pten in Pancreatic Ductal Cells Forms Intraductal Tubulopapillary Neoplasm via the YAP/TAZ Pathway

doi: 10.1016/j.jcmgh.2025.101639

Figure Lengend Snippet: RNA-seq analysis of pancreatic ductal cells in HPB mice revealed the activation of the YAP/ TAZ pathway. ( A ) Experimental scheme for tamoxifen-induced genetic recombination and analysis. The arrowheads indicate tamoxifen administration. At 21 days after the first tamoxifen administration, pancreatic ductal cells were separated using ductal cell-specific Dolichos biflorus agglutinin lectin labeling, followed by magnetic bead separation. ( B ) Volcano plot showing the differentially expressed genes in HPB and HP mice. RNA-seq analysis of genes with high fold changes (>2 log or <2 log) revealed 417 upregulated and 609 downregulated genes in HPB mice compared with HP mice. ( C ) Representative results of gene set enrichment analysis comparing pancreatic ductal cells isolated from HP and HPB mice using “Hallmark gene sets.” NES, normalized enrichment score. ( D ) GSEA plot of the YAP-conserved signature comparing pancreatic ductal cells isolated from HP and HPB mice. FDR, false discovery rate. ( E ) qRT-PCR analysis of ductal cells isolated from HP or HPB mice. The expressions of Ankrd1 , Cyr61 , and Igfbp3 were significantly upregulated in HPB mice. The expressions of Ctgf and Birc5 were upregulated in HPB mice, although the difference was not statistically significant. Data are presented as the means ± SDs (n = 3 per group). The P values were evaluated using a 2-tailed Student’s t -test. ( F ) IHC for Taz, Cyr61, Birc5, and Ctgf in the pancreas of H, HB, HP, and HPB mice. Scale bar, 20 μm. n =5 per group.

Article Snippet: CYR61 , CST , 1:50 , #39382.

Techniques: RNA Sequencing, Activation Assay, Labeling, Isolation, Quantitative RT-PCR

Journal: Frontiers in Medicine

Article Title: Serum CYR61 Is Associated With Airway Inflammation and Is a Potential Biomarker for Severity in Chronic Obstructive Pulmonary Disease

doi: 10.3389/fmed.2021.781596

Figure Lengend Snippet: Serum CYR61 level and its association with severity of COPD. Sera were collected from COPD patients and controls. (A,B) Serum CYR61 level was detected using ELISA. (A) Serum CYR61 level was compared between COPD patients and controls. All data were represented as means ± S.E.M. ( N = 150). (B) Serum CYR61 level was compared among different grades of COPD patients. All data were represented as means ± S.E.M. ( N = 69 for G 1-2 patients, N = 48 for G 3 patients, N = 33 for G 4 patients). (C,D) Pulmonary CYR61 expression was compared between COPD patients and controls. (C) Three representative pictures. (D) Quantitative analysis of CYR61-positive cells in COPD patients and controls. (E,F) Pulmonary CYR61 was compared among different grades of COPD patients. (E) Three representative pictures: arrows indicate CYR61-positive cell; (F) Quantitative analysis of CYR61-positive cells in COPD patients with different grades. All data were represented as means ± S.E.M. ( N = 6). * P < 0.05, ** P < 0.01.

Article Snippet: CYR61 ELISA kits were from Cusabio (TX, USA).

Techniques: Enzyme-linked Immunosorbent Assay, Expressing

Journal: Frontiers in Medicine

Article Title: Serum CYR61 Is Associated With Airway Inflammation and Is a Potential Biomarker for Severity in Chronic Obstructive Pulmonary Disease

doi: 10.3389/fmed.2021.781596

Figure Lengend Snippet: The correlation between serum CYR61 and lung function indexes in COPD patients. Pulmonary function was measured in 150 COPD patients. Sera were collected and CYR61 was detected using ELISA. Correlation between serum CYR61 and lung function indexes was analyzed. (A) CYR61 vs. FVC(L); (B) CYR61 vs. FEV1(L); (C) CYR61 vs. FEV1/FVC (%); (D) CYR61 vs. FEV1(%).

Article Snippet: CYR61 ELISA kits were from Cusabio (TX, USA).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Medicine

Article Title: Serum CYR61 Is Associated With Airway Inflammation and Is a Potential Biomarker for Severity in Chronic Obstructive Pulmonary Disease

doi: 10.3389/fmed.2021.781596

Figure Lengend Snippet: Association of serum CYR61 with lung function.

Article Snippet: CYR61 ELISA kits were from Cusabio (TX, USA).

Techniques:

Journal: Frontiers in Medicine

Article Title: Serum CYR61 Is Associated With Airway Inflammation and Is a Potential Biomarker for Severity in Chronic Obstructive Pulmonary Disease

doi: 10.3389/fmed.2021.781596

Figure Lengend Snippet: The association between serum CYR61 and pulmonary NF-κB activation in COPD patients. Lung tissues were collected from COPD patients and controls. Pulmonary NF-κB p65 was detected using IHC. (A,B) Pulmonary NF-κB p65-positive nuclei were compared between COPD patients and control subjects. (A) Three representative pictures: arrows indicate p65-positive nuclei; (B) Quantitative analysis of p65-positive nuclei in COPD patients and controls. (C,D) Pulmonary NF-κB p65-positive nuclei were compared among COPD patients with different levels of CYR61. (C) Three representative pictures: arrows indicate p65-positive nuclei; (D) Quantitative analysis of p65-positive nuclei in COPD patients with different levels of CYR61. All data were represented as means ± S.E.M. ( N = 6). * P < 0.05, ** P < 0.01.

Article Snippet: CYR61 ELISA kits were from Cusabio (TX, USA).

Techniques: Activation Assay, Control

Journal: Frontiers in Medicine

Article Title: Serum CYR61 Is Associated With Airway Inflammation and Is a Potential Biomarker for Severity in Chronic Obstructive Pulmonary Disease

doi: 10.3389/fmed.2021.781596

Figure Lengend Snippet: The association of serum CYR61 level with serum inflammatory cytokines in COPD patients. Sera were collected from 150 COPD patients and 150 control subjects. Serum CYR61 and inflammatory cytokines were detected using ELISA. (A,D) Association between serum CYR61 and MCP-1 was analyzed among all subjects. (A) Serum MCP-1 was compared among subjects with different levels of CYR61. (D) Correlation analysis between serum CYR61 and MCP-1. (B,E) Association between serum CYR61 and MCP-1 was analyzed among COPD patients. (B) Serum MCP-1 was compared among COPD patients with different levels of CYR61. (E) Correlation analysis between serum CYR61 and MCP-1. (C,F) Association between serum CYR61 and MCP-1 was analyzed among control subjects. (C) Serum MCP-1 was compared among control subjects with different levels of CYR61. (F) Correlation analysis between serum CYR61 and MCP-1. (G,J) Association between serum CYR61 and TNF-α was analyzed among all subjects. (G) Serum TNF-α was compared among subjects with different levels of CYR61. (J) Correlation analysis between serum CYR61 and TNF-α. (H,K) Association between serum CYR61 and TNF-α was analyzed among COPD patients. (H) Serum TNF-α was compared among COPD patients with different levels of CYR61. (K) Correlation analysis between serum CYR61 and TNF-α. (I,L) Association between serum CYR61 and TNF-α was analyzed among control subjects. (I) Serum TNF-α was compared among control subjects with different levels of CYR61. (L) Correlation analysis between serum CYR61 and TNF-α. * P < 0.05, ** P < 0.01.

Article Snippet: CYR61 ELISA kits were from Cusabio (TX, USA).

Techniques: Control, Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Medicine

Article Title: Serum CYR61 Is Associated With Airway Inflammation and Is a Potential Biomarker for Severity in Chronic Obstructive Pulmonary Disease

doi: 10.3389/fmed.2021.781596

Figure Lengend Snippet: Associations of serum CYR61 with MCP-1 and TNF-α.

Article Snippet: CYR61 ELISA kits were from Cusabio (TX, USA).

Techniques: Control

Journal: Frontiers in Medicine

Article Title: Serum CYR61 Is Associated With Airway Inflammation and Is a Potential Biomarker for Severity in Chronic Obstructive Pulmonary Disease

doi: 10.3389/fmed.2021.781596

Figure Lengend Snippet: Association of serum CYR61 and hospital stays in COPD patients.

Article Snippet: CYR61 ELISA kits were from Cusabio (TX, USA).

Techniques:

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: CCN1 Enhances Tumor Immunosuppression through Collagen-Mediated Chemokine Secretion in Pancreatic Cancer.

doi: 10.1002/advs.202500589

Figure Lengend Snippet: Figure 1. CCN1 is markedly upregulated in pancreatic cancer and associated with tumor progression. A) mRNA expression of CCN1 in human pancreatic cancer tissues (n = 182) and normal tissues (n = 165) were evaluated using the TCGA-PAAD (pancreatic adenocarcinoma) and GTEx datasets. B) Kaplan–Meier analysis of overall survival in PAAD patients based on CCN1 expression. CCN1 high expression (n = 91), CCN low expression (n = 91). C)

Article Snippet: The sections were then stained with antibodies against CCN1 (26689-1-AP, Proteintech), AACT (RAB-0011, MXB), AAT (RAB-0012, MXB), CA199 (MAB-0778, MXB), CD3 (MAB-0740, MXB), CD20 (Kit-0001, MXB), CD34 (Kit-0004, MXB), CD68 (Kit-0026, MXB), CD163 (MAB-0869, MXB), CD3 (MAB-0740, MXB), Ki-67 (MAB-0672, MXB), F4/80 (SC-377009, Santa Cruz), CD8 (SC-7970, Santa Cruz), CD3 (SC-20047, Santa Cruz), CD45 (SC-1178, Santa Cruz), CD86 (SC-28347, Santa Cruz), Gr (SC-393232, Santa Cruz), and GranzymeB (SC-8022, Santa Cruz) followed by incubation with secondary antibodies (SD3100, Celnovte) for 20 min at 37 °C in the dark.

Techniques: Expressing

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: CCN1 Enhances Tumor Immunosuppression through Collagen-Mediated Chemokine Secretion in Pancreatic Cancer.

doi: 10.1002/advs.202500589

Figure Lengend Snippet: Figure 2. Ccn1 expression in PDAC correlates with immunosuppression. A) Tumor growth curves of sgCtrl and sgCcn1 KPC cells subcutaneously in- oculated in C57B/L6 mouse. n = 6 mice per group. B) Representative images of tumors (left) and quantification of tumor weight (right) in sgCtrl and sgCcn1 KPC cell-inoculated mice. C) Representative images of H&E, and immunohistochemistry analysis for Ki67, CCN1, CD45, F4/80, CD4, CD8, and

Article Snippet: The sections were then stained with antibodies against CCN1 (26689-1-AP, Proteintech), AACT (RAB-0011, MXB), AAT (RAB-0012, MXB), CA199 (MAB-0778, MXB), CD3 (MAB-0740, MXB), CD20 (Kit-0001, MXB), CD34 (Kit-0004, MXB), CD68 (Kit-0026, MXB), CD163 (MAB-0869, MXB), CD3 (MAB-0740, MXB), Ki-67 (MAB-0672, MXB), F4/80 (SC-377009, Santa Cruz), CD8 (SC-7970, Santa Cruz), CD3 (SC-20047, Santa Cruz), CD45 (SC-1178, Santa Cruz), CD86 (SC-28347, Santa Cruz), Gr (SC-393232, Santa Cruz), and GranzymeB (SC-8022, Santa Cruz) followed by incubation with secondary antibodies (SD3100, Celnovte) for 20 min at 37 °C in the dark.

Techniques: Expressing, Immunohistochemistry

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: CCN1 Enhances Tumor Immunosuppression through Collagen-Mediated Chemokine Secretion in Pancreatic Cancer.

doi: 10.1002/advs.202500589

Figure Lengend Snippet: Figure 3. Ccn1 loss reduces the expression of chemokines and collagens in PDAC. A) Volcano plot illustrates the distribution of differential genes between sgCtrl and sgCcn1 KPC cells. B) Heatmap showing the mRNA expression of chemokines and collagens between sgCtrl and sgCcn1 KPC cells. C) mRNA levels of chemokines, including Ccl2, Ccl7, Ccl20, Cxcl1, Cxcl3, Cxcl5, Csf1, Csf2, and Csf3 between sgCtrl and sgCcn1 in KPC cells. D) Spearman

Article Snippet: The sections were then stained with antibodies against CCN1 (26689-1-AP, Proteintech), AACT (RAB-0011, MXB), AAT (RAB-0012, MXB), CA199 (MAB-0778, MXB), CD3 (MAB-0740, MXB), CD20 (Kit-0001, MXB), CD34 (Kit-0004, MXB), CD68 (Kit-0026, MXB), CD163 (MAB-0869, MXB), CD3 (MAB-0740, MXB), Ki-67 (MAB-0672, MXB), F4/80 (SC-377009, Santa Cruz), CD8 (SC-7970, Santa Cruz), CD3 (SC-20047, Santa Cruz), CD45 (SC-1178, Santa Cruz), CD86 (SC-28347, Santa Cruz), Gr (SC-393232, Santa Cruz), and GranzymeB (SC-8022, Santa Cruz) followed by incubation with secondary antibodies (SD3100, Celnovte) for 20 min at 37 °C in the dark.

Techniques: Expressing

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: CCN1 Enhances Tumor Immunosuppression through Collagen-Mediated Chemokine Secretion in Pancreatic Cancer.

doi: 10.1002/advs.202500589

Figure Lengend Snippet: Figure 5. Ccn1 regulates the TME in PDAC. A) Schematic representation of macrophage isolation from mouse bone marrow. B) Macrophage invasion evaluated by the chemotactic effect of sgCtrl and sgCcn1 KPC cells in a Transwell invasion assay. C) Macrophage polarization assessed by flow cytometry in macrophages cocultured with sgCtrl and sgCcn1 KPC cells. D) Macrophage polarization assessed by flow cytometry in macrophages cocultured with

Article Snippet: The sections were then stained with antibodies against CCN1 (26689-1-AP, Proteintech), AACT (RAB-0011, MXB), AAT (RAB-0012, MXB), CA199 (MAB-0778, MXB), CD3 (MAB-0740, MXB), CD20 (Kit-0001, MXB), CD34 (Kit-0004, MXB), CD68 (Kit-0026, MXB), CD163 (MAB-0869, MXB), CD3 (MAB-0740, MXB), Ki-67 (MAB-0672, MXB), F4/80 (SC-377009, Santa Cruz), CD8 (SC-7970, Santa Cruz), CD3 (SC-20047, Santa Cruz), CD45 (SC-1178, Santa Cruz), CD86 (SC-28347, Santa Cruz), Gr (SC-393232, Santa Cruz), and GranzymeB (SC-8022, Santa Cruz) followed by incubation with secondary antibodies (SD3100, Celnovte) for 20 min at 37 °C in the dark.

Techniques: Isolation, Transwell Invasion Assay, Cytometry

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: CCN1 Enhances Tumor Immunosuppression through Collagen-Mediated Chemokine Secretion in Pancreatic Cancer.

doi: 10.1002/advs.202500589

Figure Lengend Snippet: Figure 6. Targeting Ccn1 enhances the efficacy of gemcitabine and anti-PD-1 in PDAC. A) Cell viability of sgCtrl and sgCcn1 KPC cells after treatment with indicated concentrations of gemcitabine for 48 h. B) ROS levels in sgCtrl and sgCcn1 KPC cells with gemcitabine for 48 h by Fluorescence-activated cell sorting (FACS) analysis. C) Representative images (left) and quantification (right) of ROS level in sgCtrl and sgCcn1 KPC cells treated with gemcitabine for 48 h. D,E) C57B/L6 mice were inoculated subcutaneously with sgCtrl and sgCcn1 KPC cells. Tumor-bearing mice were treated with isotype control, anti-PD1 mAb, gemcitabine, or a combination of both treatments (n = 5 mice per group). Tumor volume was monitored (D), and representative tumor images (left) were acquired with final tumor weights (right) shown (E). F) C57BL/6 mice were orthotopically implanted with sgCtrl or sgCcn1 KPC cells. Tumor-bearing mice were treated with isotype control, anti-PD1 monoclonal antibody, gemcitabine, or a combination of both (n = 5 mice per group). Representative tumor images (top) and final tumor weights (bottom) are shown. G) Representative images of Masson’s trichrome staining and immunohistochemical analysis for F4/80, CD8, CD45, GranzymeB (GrzB), and Gr in sgCtrl and sgCcn1 KPC orthotopic tumors.

Article Snippet: The sections were then stained with antibodies against CCN1 (26689-1-AP, Proteintech), AACT (RAB-0011, MXB), AAT (RAB-0012, MXB), CA199 (MAB-0778, MXB), CD3 (MAB-0740, MXB), CD20 (Kit-0001, MXB), CD34 (Kit-0004, MXB), CD68 (Kit-0026, MXB), CD163 (MAB-0869, MXB), CD3 (MAB-0740, MXB), Ki-67 (MAB-0672, MXB), F4/80 (SC-377009, Santa Cruz), CD8 (SC-7970, Santa Cruz), CD3 (SC-20047, Santa Cruz), CD45 (SC-1178, Santa Cruz), CD86 (SC-28347, Santa Cruz), Gr (SC-393232, Santa Cruz), and GranzymeB (SC-8022, Santa Cruz) followed by incubation with secondary antibodies (SD3100, Celnovte) for 20 min at 37 °C in the dark.

Techniques: Fluorescence, FACS, Control, Staining, Immunohistochemical staining

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: CCN1 Enhances Tumor Immunosuppression through Collagen-Mediated Chemokine Secretion in Pancreatic Cancer.

doi: 10.1002/advs.202500589

Figure Lengend Snippet: Figure 7. Proposed model for CCN1-induced tumor immunosuppression in pancreatic cancer. This schematic diagram illustrates the role of CCN1 in pancreatic cancer. In control pancreatic cancer cells, the tumor cells exhibit resistance to TNF𝛼-mediated cell death. In contrast, CCN1- KO pancreatic tumor cells are more sensitive to TNF𝛼-induced apoptosis. Loss of CCN1 leads to reduced collagen expression, which further downregulates the secretion of chemokines, ultimately promoting immune cell infiltration into the TME.

Article Snippet: The sections were then stained with antibodies against CCN1 (26689-1-AP, Proteintech), AACT (RAB-0011, MXB), AAT (RAB-0012, MXB), CA199 (MAB-0778, MXB), CD3 (MAB-0740, MXB), CD20 (Kit-0001, MXB), CD34 (Kit-0004, MXB), CD68 (Kit-0026, MXB), CD163 (MAB-0869, MXB), CD3 (MAB-0740, MXB), Ki-67 (MAB-0672, MXB), F4/80 (SC-377009, Santa Cruz), CD8 (SC-7970, Santa Cruz), CD3 (SC-20047, Santa Cruz), CD45 (SC-1178, Santa Cruz), CD86 (SC-28347, Santa Cruz), Gr (SC-393232, Santa Cruz), and GranzymeB (SC-8022, Santa Cruz) followed by incubation with secondary antibodies (SD3100, Celnovte) for 20 min at 37 °C in the dark.

Techniques: Control, Expressing

Journal: Neoplasia (New York, N.Y.)

Article Title: mTORC2-mediated direct phosphorylation regulates YAP activity promoting glioblastoma growth and invasive characteristics.

doi: 10.1016/j.neo.2021.07.005

Figure Lengend Snippet: Fig. 6. mTORC2-mediated YAP S436 phosphorylation can occur independent of Hippo pathway signaling in GBM. (A) Inhibition of YAP S436 phosphorylation by JR-AB2-011 (1 μM, 8 h) in the indicated lines. Immunoblots were probed for phospho-S436 YAP, total YAP and actin as shown. (B) CTGF and Cyr61 mRNA expression in LN229 shYAP1 cells expressing native YAP or LATS-resistant YAP-5SA mutant (C) following treatment with either JR-AB2-011 (1 μM, 8 h) or insulin (100 nM, 4 h) relative to control untreated cells. mRNA was isolated and subjected to qRT-PCR analyses. qRT-PCR measurements were performed in quadruplicate and the mean and + S.D. are shown. ∗P < 0.05.

Article Snippet: Antibodies to the following proteins were used: phospho-S 473 -AKT (#9271, CST), phospho-S 127 -YAP (#ab76252, Abcam), AKT (#9272, CST), Rictor (#A300-459A, Bethyl Laboratories), Raptor (A300-553A, Bethyl Laboratories), actin (#ab3280, Abcam), YAP1 (#12395S, CST), α-Flag (#TA50011, Origene), TEAD1 (#12292S, CST), TEAD2 (#ab92279, Abcam), TEAD3 (#13224S, CST), TEAD4 (#ab137833, Abcam), SMAD1 (#9743S, CST), p73 (#14620S, CST), FOS (#4384S, CST), TBX5 (#ab137833, Abcam), Mnk1 (#sc-133107, Santa Cruz Biotechnology), CTGF (#HPA031075, Sigma), Cyr61 (#NB100-356SS, Novus), Hsp90 (#SMC149B, StressMarq Biosciences), Lamin B2 (#12255S, CST) and mSin1 (#07-2276, MilliporeSigma).

Techniques: Phospho-proteomics, Inhibition, Western Blot, Expressing, Mutagenesis, Control, Isolation, Quantitative RT-PCR

Journal: Neoplasia (New York, N.Y.)

Article Title: mTORC2-mediated direct phosphorylation regulates YAP activity promoting glioblastoma growth and invasive characteristics.

doi: 10.1016/j.neo.2021.07.005

Figure Lengend Snippet: Fig. 7. Growth of YAP S436 mutant expressing GBM cells in vivo . (A) LN229 shYAP1 cells stably expressing native YAP1, nonphosphorylatable YAP1 S436A, or the phosphomimetic YAP1 S436E alleles were monitored for tumor growth for up to 52 days following establishment of ~200 cm 3 subcutaneous tumors in SCID mice ( n = 4-5 per group). (B) Overall survival of mice with harboring the indicated S436 YAP mutant subcutaneous implanted LN229 tumors. ∗, P < 0.05, n = 4-5 mice per group. (C) Weight of tumors harvested at autopsy from xenografted mice implanted with the indicated GBM cells. ∗P < 0.05. (D) CTGF and Cyr61 mRNA expression from harvested tumors cells expressing the indicated YAP1 alleles. mRNA was isolated and subjected to qRT-PCR analyses. qRT-PCR measurements were performed in quadruplicate and the mean and + S.D. are shown. ∗P < 0.05. (E) mTORC2 mediated phosphorylation of serine 436 of YAP1 leads to increases in protein stability, nuclear localization and TEAD association resulting in enhancement of YAP1 target gene expression.

Article Snippet: Antibodies to the following proteins were used: phospho-S 473 -AKT (#9271, CST), phospho-S 127 -YAP (#ab76252, Abcam), AKT (#9272, CST), Rictor (#A300-459A, Bethyl Laboratories), Raptor (A300-553A, Bethyl Laboratories), actin (#ab3280, Abcam), YAP1 (#12395S, CST), α-Flag (#TA50011, Origene), TEAD1 (#12292S, CST), TEAD2 (#ab92279, Abcam), TEAD3 (#13224S, CST), TEAD4 (#ab137833, Abcam), SMAD1 (#9743S, CST), p73 (#14620S, CST), FOS (#4384S, CST), TBX5 (#ab137833, Abcam), Mnk1 (#sc-133107, Santa Cruz Biotechnology), CTGF (#HPA031075, Sigma), Cyr61 (#NB100-356SS, Novus), Hsp90 (#SMC149B, StressMarq Biosciences), Lamin B2 (#12255S, CST) and mSin1 (#07-2276, MilliporeSigma).

Techniques: Mutagenesis, Expressing, In Vivo, Stable Transfection, Isolation, Quantitative RT-PCR, Phospho-proteomics, Targeted Gene Expression

Journal: International journal of molecular sciences

Article Title: Cilostazol Stimulates Angiogenesis and Accelerates Fracture Healing in Aged Male and Female Mice by Increasing the Expression of PI3K and RUNX2.

doi: 10.3390/ijms25020755

Figure Lengend Snippet: Figure 6. (a) Representative Western blots of BMP-2 and BMP-4 expression within the callus tissue of controls and cilostazol-treated mice at 2 weeks after fracture. (b,c) Expression of BMP-2 (b) and BMP-4 (c) within the callus tissue of controls (white bars, n = 4) and cilostazol-treated mice (black bars, n = 4) at 2 weeks after fracture. Mean ± SEM. (d) Representative Western blots of CYR61 and CD31 expression within the callus tissue of controls and cilostazol-treated mice at 2 weeks after fracture. (e,f) Expression of CYR61 (e) and CD31 (f) within the callus tissue of controls (white bars, n = 4) and cilostazol-treated mice (black bars, n = 4) at 2 weeks after fracture. Mean ± SEM; * p < 0.05 vs. control. (g) Representative Western blots of PI3K and RUNX2 expression within the callus tissue of controls and cilostazol-treated mice at 2 weeks after fracture. (h,i) Expression of PI3K (h) and RUNX2 (i) within the callus tissue of controls (white bars, n = 4) and cilostazol-treated mice (black bars, n = 4) at 2 weeks after fracture. Mean ± SEM; * p < 0.05 vs. control. (b,f) Non-parametric data; analysis performed by Mann–Whitney U-test. (c,e,h,i) Parametric data; analysis performed by unpaired Student’s t-test.

Article Snippet: After saving the whole protein fraction, the analysis was performed using the following monoclonal antibodies: goat anti-mouse BMP2 and BMP4 (1:300, R&D Systems, Wiesbaden, Germany), sheep anti-mouse CYR61 (1:300, R&D Systems), rabbit anti-mouse CD31 (1:300, Cell Signaling Technology, Danvers, MA, USA), mouse anti-mouse PI3K (1:100, Santa Cruz Biotechnology, Heidelberg, Germany), and rabbit anti-mouse RUNX2 (1:300, Abcam).

Techniques: Western Blot, Expressing, Control, MANN-WHITNEY