Journal: Oncotarget

Article Title: Epithelial derived CTGF promotes breast tumor progression via inducing EMT and collagen I fibers deposition

doi:

Figure Lengend Snippet: Kaplan–Meier curves showing the overall survival and recurrence-free survival of patients with low or high tumor epithelial or stroma CTGF expression in the combined cohort of 103 breast cancer patients, P values were determined by log rank test.

Article Snippet: Overexpression of full-length CTGF gene was achieved by transfection of cDNA for CTGF (OriGene Technologies), which clones the open reading frame (ORF) of this gene into the pCMV6 Entry vector.

Techniques: Expressing

Journal: Oncotarget

Article Title: Epithelial derived CTGF promotes breast tumor progression via inducing EMT and collagen I fibers deposition

doi:

Figure Lengend Snippet: A–B. Quantitative graphs and representative CTGF immunohistochemistry staining of the breast tumor PDX tissue microarray. H score was calculated as described in Methods. P values were determined by student t -test. C. Representative images showing the Masson's trichrome staining and CTGF immunohistochemistry staining of the triple-negative (TNBC) and HER2-positive (HRE2+) breast tumor PDX tissue microarray. D. Correlation between the H scores of CTGF in tumor cells and % of stroma area in the TNBC tissue microarrays. R 2 = 0.66. E. Correlation between the H scores of CTGF in stromal cells and % of stroma area in the TNBC tissue microarrays. R 2 = 0.39. F. Correlation between the H scores of CTGF in stromal cells and % of stroma area in the HER2+ tissue microarrays. R 2 = 0.43.

Article Snippet: Overexpression of full-length CTGF gene was achieved by transfection of cDNA for CTGF (OriGene Technologies), which clones the open reading frame (ORF) of this gene into the pCMV6 Entry vector.

Techniques: Immunohistochemistry, Staining, Microarray

Journal: Oncotarget

Article Title: Epithelial derived CTGF promotes breast tumor progression via inducing EMT and collagen I fibers deposition

doi:

Figure Lengend Snippet: A–B. The co-expression analysis between CTGF and FN1, VIM, CDH1 in the n = 22 clinical breast tumor cohort . All gene probes were listed for FN1, VIM and CDH1 . The correlation co-efficient (cc) were calculated based on the log2 median-centered intensity of FN1, VIM, CDH1 and CTGF. C–D. The co-expression analysis between CTGF and FN1, VIM, CDH1 in the n = 66 clinical breast tumor cohort . E–H. CTGF gene expression in a series EMT-induced human mammary epithelial cell (HMLER) cell lines, including over-expression of EMT transcription factor Goosecoid (GSC), Snail, or Twist, and induction of EMT by TGFB1 transfection. P values were determined by student t -test. Fold change was log 2 scale.

Article Snippet: Overexpression of full-length CTGF gene was achieved by transfection of cDNA for CTGF (OriGene Technologies), which clones the open reading frame (ORF) of this gene into the pCMV6 Entry vector.

Techniques: Expressing, Gene Expression, Over Expression, Transfection

Journal: Oncotarget

Article Title: Epithelial derived CTGF promotes breast tumor progression via inducing EMT and collagen I fibers deposition

doi:

Figure Lengend Snippet: A. Representative phase-contrast images showing the elongated morphology of over-CTGF-HMLER cells which is typically associated with mesenchymal phenotype. B. RT-PCR analysis of the EMT genes and CTGF in the control and over-CTGF-HMLER cells. Relative gene expression level to GAPDH was plotted. C. Western blot analysis of the EMT markers and CTGF in the control and over-CTGF-HMLER cells. D–E. Quantitative graphs and representative membrane images showing the migration ability of the control and over-CTGF-HMLER cells measured by the Boyden Chamber assay. F. Quantitative graphs showing the invasion ability of the control and over-CTGF-HMLER cells measured by coating Matrigel in the Boyden Chamber assay. G–H. Enforced expression of CTGF in the HMLER cells promoted the number and size of mammospheres. I. Western blot analysis of enriched CD44 expression in the mammospheres. ** P < 0.01, vs control. P values were determined by student t -test.

Article Snippet: Overexpression of full-length CTGF gene was achieved by transfection of cDNA for CTGF (OriGene Technologies), which clones the open reading frame (ORF) of this gene into the pCMV6 Entry vector.

Techniques: Reverse Transcription Polymerase Chain Reaction, Control, Gene Expression, Western Blot, Membrane, Migration, Boyden Chamber Assay, Expressing

Journal: Oncotarget

Article Title: Epithelial derived CTGF promotes breast tumor progression via inducing EMT and collagen I fibers deposition

doi:

Figure Lengend Snippet: A. Representative phase-contrast images showing the epithelial morphology of the shCTGFs knockdown cells. B. Western blot analysis of the CTGF and N-cadherin, E-cadherin in the control and shCTGFs cells. C–D. Quantitative graphs and representative membrane images showing the migration ability of the control and shCTGFs cells measured by the Boyden Chamber assay. E. Quantitative graphs showing the invasion ability of the control and shCTGFs cells measured by coating Matrigel in the Boyden Chamber assay. F–G. CTGF depletion in the HMLER-snail cells reduced the number and size of mammospheres. H. Western blot analysis of CD44 in the mammospheres. * P < 0.05, vs control; ** P < 0.01, vs control. P values were determined by student t -test.

Article Snippet: Overexpression of full-length CTGF gene was achieved by transfection of cDNA for CTGF (OriGene Technologies), which clones the open reading frame (ORF) of this gene into the pCMV6 Entry vector.

Techniques: Knockdown, Western Blot, Control, Membrane, Migration, Boyden Chamber Assay

Journal: Oncotarget

Article Title: Epithelial derived CTGF promotes breast tumor progression via inducing EMT and collagen I fibers deposition

doi:

Figure Lengend Snippet: A–B. In vivo tumor growth of the over-CTGF-HMLER cells and control HMLER cells mixed with Matrigel (A) and PBS (B) C. Representative images showing the Masson's trichrome staining of the tumor connective tissue, and second harmonic generation (SHG) imaging of the intra-tumoral collagen I fibers, of the Matrigel-mixed over-CTGF-HMLER tumors. D–E. In vivo tumor growth of the shCTGF-HMLER cells (D), and representative Masson's trichrome staining and SHG imaging of the intra-tumoral collagen I fibers. F. Quantification of the percentage of collagen fiber and perpendicular collagen fiber to the overall connective tissue area. Mouse tumor sections (10 μm thick) were cut serially, and one section from every 100 μm was stained by Masson's trichrome staining, and the adjacent section was imaged by SHG imaging. In each section 5–10 fields were quantified. N = 4 mouse tumors in each group. * P < 0.05, vs control HMLER; ^ P < 0.05, vs HMLER-snail. P values were determined by student t -test.

Article Snippet: Overexpression of full-length CTGF gene was achieved by transfection of cDNA for CTGF (OriGene Technologies), which clones the open reading frame (ORF) of this gene into the pCMV6 Entry vector.

Techniques: In Vivo, Control, Staining, Imaging

Journal: Oncotarget

Article Title: Epithelial derived CTGF promotes breast tumor progression via inducing EMT and collagen I fibers deposition

doi:

Figure Lengend Snippet: A–B. A precedent treatment with anti-TNFR1 monoclonal antibody (10 μg/ml) more profoundly prevented the increase of CTGF-induced TNFR1 expression in the CTGF-highly expressed cells (over-CTGF-HMLER and HMLER-snail) than that in the CTGF-lowly expressed cells (control-HMLER and shCTGF2). C. Representative phase-contrast images showing the cell morphology changes in the indicated cell lines and treatments. D–E. The activation of IKBKA and IKBKB by CTGF could be specifically attenuated by the anti-TNFR1 antibody in the CTGF-high tumor cells. F. JUK but not ERK1/2 and p38 in the MAPK signaling pathway was up-regulated by exogenous CTGF in the CTGF-low tumor cells. G. Knockdown CTGF in the HMLER-snail cells down-regulated the IKBKA and IKBKB, and such down-regulation could be rescued by exogenous CTGF. All experimental data are representative of at least two independent experiments performed in triplicate. Data are expressed as mean ± s.e.m.

Article Snippet: Overexpression of full-length CTGF gene was achieved by transfection of cDNA for CTGF (OriGene Technologies), which clones the open reading frame (ORF) of this gene into the pCMV6 Entry vector.

Techniques: Expressing, Control, Activation Assay, Knockdown

Journal: Oncotarget

Article Title: Epithelial derived CTGF promotes breast tumor progression via inducing EMT and collagen I fibers deposition

doi:

Figure Lengend Snippet: A–B. Cell proliferation and mammosphere formation were significantly inhibited by the anti-TNFR1 treatment in over-CTGF-HMLER and HMLE-snail cells but not control-HMLER cells or shCTGF-HMLE-snail. C–D. Cell proliferation and mammosphere formation were significantly inhibited by resveratrol treatment in over-CTGF-HMLER and HMLE-snail cells but not control-HMLER cells or shCTGF-HMLE-snail. * P < 0.05, ** P < 0.01, vs anti-TNFR1(−) or resveratrol (−) group of the corresponding cell lines. P values were determined by student t -test.

Article Snippet: Overexpression of full-length CTGF gene was achieved by transfection of cDNA for CTGF (OriGene Technologies), which clones the open reading frame (ORF) of this gene into the pCMV6 Entry vector.

Techniques: Control

Journal: Oncotarget

Article Title: Epithelial derived CTGF promotes breast tumor progression via inducing EMT and collagen I fibers deposition

doi:

Figure Lengend Snippet: CTGF is mainly derived from epithelial tumor cells. Through the CTGF-TNFR1-IκB autocrine signaling, CTGF promotes tumor EMT which contributes to the stemness and metastasis. Through promoting the deposition and orientation of collagen I fibers in the primary tumor stroma, CTGF promotes stroma-tumor interaction which facilitates tumor progression.

Article Snippet: Overexpression of full-length CTGF gene was achieved by transfection of cDNA for CTGF (OriGene Technologies), which clones the open reading frame (ORF) of this gene into the pCMV6 Entry vector.

Techniques: Derivative Assay

Journal: Inflammation

Article Title: CTGF Triggers Rat Astrocyte Activation and Astrocyte-Mediated Inflammatory Response in Culture Conditions

doi: 10.1007/s10753-019-01029-7

Figure Lengend Snippet: CTGF activated rat astrocytes through an autocrine-dependent manner in vitro. RA cells were cultured in the presence of 10 ng/ml recombinant TGF-β1, 5 μg/ml Brefeldin A, 10 μg/ml CTGF neutralizing antibody, or recombinant CTGF with indicated concentrations for 24 h, respectively, and the mRNA expressions of CTGF ( a ) and GFAP ( b ) were examined by quantitative real-time polymerase chain reaction (qRT-PCR). Results are expressed as the mean ± S.E. * p < 0.05, ** p < 0.01, and *** p < 0.001. N.S. not significant.

Article Snippet: Recombinant rat CTGF was obtained from R&D Systems with purity > 85% and endotoxin level < 0.1 EU/ml.

Techniques: In Vitro, Cell Culture, Recombinant, Real-time Polymerase Chain Reaction, Quantitative RT-PCR

Journal: Inflammation

Article Title: CTGF Triggers Rat Astrocyte Activation and Astrocyte-Mediated Inflammatory Response in Culture Conditions

doi: 10.1007/s10753-019-01029-7

Figure Lengend Snippet: CTGF facilitated the production of cytokines and chemokines by astrocytes. RA cells were cultured in the presence of 10 ng/ml recombinant CTGF or TGF-β1 for 24 h, and supernatants were collected to measure the production of cytokines—TNF-α, IL-6, and IL-1β ( a ), and chemokines—MCP-1, RANTES, and CXCL1 ( b ). Results are expressed as the mean ± S.E. *: p < 0.05 as compared with solvent control group. * p < 0.05, ** p < 0.01, and *** p < 0.001 as compared with solvent control group.

Article Snippet: Recombinant rat CTGF was obtained from R&D Systems with purity > 85% and endotoxin level < 0.1 EU/ml.

Techniques: Cell Culture, Recombinant, Solvent, Control

Journal: Inflammation

Article Title: CTGF Triggers Rat Astrocyte Activation and Astrocyte-Mediated Inflammatory Response in Culture Conditions

doi: 10.1007/s10753-019-01029-7

Figure Lengend Snippet: CTGF enhanced the recruitment of PBMCs. a Schematic diagram of the Boyden chamber assay. b Effects of CTGF treatment on the chemotactic migration of PBMCs cocultured with RA. Number of invaded PBMCs during co-culture with RA was indicated on vertical axis. Results are expressed as the mean ± S.E. * p < 0.05 and ** p < 0.01.

Article Snippet: Recombinant rat CTGF was obtained from R&D Systems with purity > 85% and endotoxin level < 0.1 EU/ml.

Techniques: Boyden Chamber Assay, Migration, Co-Culture Assay

Journal: Inflammation

Article Title: CTGF Triggers Rat Astrocyte Activation and Astrocyte-Mediated Inflammatory Response in Culture Conditions

doi: 10.1007/s10753-019-01029-7

Figure Lengend Snippet: CTGF activated the NF-κB and AP-1 through ASK1-p38/JNK pathways. a RA cells were incubated with 10 ng/ml CTGF or TGF-β1 for 24 h, and total protein extracts were collected. The expression of key molecules in ASK, p38/JNK, and NF-κB/AP-1 pathways and their phosphorylation states were examined by Western blotting. b RA cells were treated with either 10 μM ASK1 inhibitor GS-4997, or solvent control as indicated, for 30 min prior to stimulation with 20 ng/ml CTGF for 24 h. The expression of ASK, p65, and c-Jun and their phosphorylation states were examined by Western blotting. c RA cells were treated with either 5 μM SB20358 or 25 μM SP600125, or DMSO as control, for 30 min prior to stimulation with CTGF for 24 h, and total protein extracts were collected. The expression of ASK, p65, c-Jun and their phosphorylation states were examined by Western blotting. d RA cells were treated with chemical inhibitors or solvent control for 30 min prior to stimulation with 10 ng/ml CTGF for 24 h. The mRNA expression of IL-6 was analyzed using qRT-PCR. * p < 0.05 and ** p < 0.01 as compared with CTGF group.

Article Snippet: Recombinant rat CTGF was obtained from R&D Systems with purity > 85% and endotoxin level < 0.1 EU/ml.

Techniques: Incubation, Expressing, Phospho-proteomics, Western Blot, Solvent, Control, Quantitative RT-PCR

Journal: Inflammation

Article Title: CTGF Triggers Rat Astrocyte Activation and Astrocyte-Mediated Inflammatory Response in Culture Conditions

doi: 10.1007/s10753-019-01029-7

Figure Lengend Snippet: Augmentation of inflammation by CTGF is abrogated by p38 or JNK inhibitors. RA cells were treated with either 5 μM SB20358 or 25 μM SP600125, or DMSO as control, for 30 min prior to stimulation with CTGF. Supernatants were collected to measure the production of TNF-α ( a ) and MCP-1 ( b ), 24 h later. c RA cells were seeded in the lower wells of the Boyden chamber, then treated with inhibitors or DMSO for 30 min, followed by the treatment with 10 ng/ml CTGF. Thereafter, the upper wells seeded with PBMCs were placed on top of the lower wells and cells were cocultured for 96 h, then the invaded PBMCs under the membrane were counted. Results are expressed as the mean ± S.E. * p < 0.05, ** p < 0.01, and *** p < 0.001 as compared with solvent control group. N.S. not significant.

Article Snippet: Recombinant rat CTGF was obtained from R&D Systems with purity > 85% and endotoxin level < 0.1 EU/ml.

Techniques: Control, Membrane, Solvent

Journal: Inflammation

Article Title: CTGF Triggers Rat Astrocyte Activation and Astrocyte-Mediated Inflammatory Response in Culture Conditions

doi: 10.1007/s10753-019-01029-7

Figure Lengend Snippet: Schematic presentation of the signaling pathways involved in CTGF-induced inflammatory augmentation of astrocytes.

Article Snippet: Recombinant rat CTGF was obtained from R&D Systems with purity > 85% and endotoxin level < 0.1 EU/ml.

Techniques: Protein-Protein interactions