Journal: Theranostics

Article Title: Blockade of phosphotyrosine pathways suggesting SH2 superbinder as a novel therapy for pulmonary fibrosis

doi: 10.7150/thno.72269

Figure Lengend Snippet: High pY levels might affect the proliferation, migration and differentiation of lung fibroblasts. A , Changes in cell proliferation between hnLFs and hfLFs determined by CCK-8 kit (n = 7). *** P < 0.001. B-C , Assessing cell proliferation by EdU assay between hnLFs and hfLFs (n = 6). *** P < 0.001. D-G , Comparison of migration between hnLFs and hfLFs (n = 6). *** P < 0.001. H , Representative images of hnLFs and hfLFs under TEM. Arrows pointed to the lipid droplets in cytoplasm. I , Representative fluorescence images of Nile Red in hnLFs and hfLFs. J-K , The contractility of hnLFs and hfLFs in 3D collagen matrices (n = 6). *** P < 0.001. L , Phospho-RTK array analysis with 200 μg of lysates from hnLFs and hfLFs, which were serum-starved for 6 h. M , Quantification of phospho-RTK levels (average of two drops fold average of positive control). N-O , Western blot and quantification of p-VEGR2, p-PDGFRβ, p-EGFR and p-FGFR1 in hnLFs and hfLFs (n = 6). * P < 0.05, ** P < 0.01, *** P < 0.001. P-Q , Western blot and quantification of p-PLCγ1, p-GAB1, p-SHC and p-SRC in hnLFs and hfLFs (n = 6). ** P < 0.01, *** P < 0.001. R-S , Western blot and quantification of p-AKT, p-ERK1/2 and p-STAT3 in hnLFs and hfLFs (n = 6). ** P < 0.01, *** P < 0.001.

Article Snippet: Antibodies against FN (Cat#15613-1-AP), COL1A1 (Cat#67288-1-lg), α-SMA (Cat#55135-1-AP, Cat#67735-1-AP), FGFR1 (Cat#60325-1-lg), CD31 (Cat#60287-1-lg), CD45 (Cat#11265-1-lg), EpCAM (Cat#60287-1-lg), Vimentin (Cat#10366-1-AP) and GAPDH (Cat#60004-1-lg) were purchased from Proteintech (Rosemont, IL, USA).

Techniques: Migration, CCK-8 Assay, EdU Assay, Comparison, Fluorescence, Positive Control, Western Blot

Journal: Theranostics

Article Title: Blockade of phosphotyrosine pathways suggesting SH2 superbinder as a novel therapy for pulmonary fibrosis

doi: 10.7150/thno.72269

Figure Lengend Snippet: SH2 superbinder blocked multiple fibrosis associated pathways through interrupting pY-SH2 combination in pY mediated signal transmission. A-B , Phospho-RTK array analysis with 200 μg of lysates from hfLFs treated with 1 μM GST-SH2 WT or GST-SH2 TrM for 24 h. C , Western blot of p-VEGR2, p-PDGFRβ, p-EGFR and p-FGFR1 in hnLFs and hfLFs after GST, GST-SH2 WT and GST-SH2 TrM incubation. D , Western blot of p-PLCγ1, p-GAB1, p-SHC and p-SRC in hnLFs and hfLFs after GST, GST-SH2 WT or GST-SH2 TrM incubation. E , Western blot of p-AKT, p-ERK1/2 and p-STAT3 in hnLFs and hfLFs after GST, GST-SH2 WT or GST-SH2 TrM incubation. F , hnLFs and hfLFs were incubated with 1 μM GST, GST-SH2 WT or GST-SH2 TrM for 24 h. The immunoprecipitation of EGFR and SHC was detected. G , Representative fluorescence images of EGFR and SHC colocalization in hnLFs and hfLFs after GST, GST-SH2 WT or GST-SH2 TrM treatment. H , Immunoprecipitations of GRB2 with GAB1, SHC, EGFR and PDGFRβ. I , GST pull down assay showed the binding capacity of SH2 superbinder with pY in hnLFs and hfLFs. J , Representative fluorescence images of GST tag and pY colocalization in hnLFs and hfLFs after incubation with GST, GST-SH2 WT or GST-SH2 TrM. K-L , GST pull down assay showed the binding capacity of SH2 superbinder with RTKs (such as VEGFR2, PDGFRβ, EGFR and FGFR1) and adaptor proteins (such as GAB1, SHC and SRC) in hfLFs and hfLFs.

Article Snippet: Antibodies against FN (Cat#15613-1-AP), COL1A1 (Cat#67288-1-lg), α-SMA (Cat#55135-1-AP, Cat#67735-1-AP), FGFR1 (Cat#60325-1-lg), CD31 (Cat#60287-1-lg), CD45 (Cat#11265-1-lg), EpCAM (Cat#60287-1-lg), Vimentin (Cat#10366-1-AP) and GAPDH (Cat#60004-1-lg) were purchased from Proteintech (Rosemont, IL, USA).

Techniques: Transmission Assay, Western Blot, Incubation, Immunoprecipitation, Fluorescence, Pull Down Assay, Binding Assay

Journal: Scientific Reports

Article Title: Up-regulation of pro-angiogenic pathways and induction of neovascularization by an acute retinal light damage

doi: 10.1038/s41598-020-63449-y

Figure Lengend Snippet: bFGF and FGFR1 analysis. ( A ) bFGF (left) and FGFR1 (right) analysis by Western Blot. Statistical analysis was performed by one-way ANOVA test followed by Tukey test. Data are shown as mean ± SE (n = 4). *p < 0,05; **p < 0,01; ***p < 0,001 versus Control. # p < 0,05; ## p < 0,01; ### p < 0,001 versus LD. Original western blots presented are available in Supplementary Fig. . ( B ) Representative confocal images of retinal cryosections immunolabelled for bFGF (green) and FGFR1 (red). Nuclei were stained with Bisbenzimide (blue). (a–e) High magnification of a random area of the sections showing co-localization in yellow (white arrows) of bFGF and FGFR1. Both bFGF and FGFR1 increased in the rat retina after light damage over time. CTRL (Control); LD (Light damage); LD + 7rec (Light damage + 7 days of recovery); LD + 60rec (Light damage + 60 days of recovery); LD + 120rec (Light damage + 120 days of recovery); ONL (Outer nuclear layer); INL (Inner nuclear layer); GCL (Ganglion cell layer).

Article Snippet: Sections were incubated overnight at 4 °C with primary antibodies: polyclonal anti-VEGFA (Santa Cruz, sc-7269) (1:250 in 1% BSA), polyclonal anti-VEGFR2 (Invitrogen, AHR5102) (1:250 in 1% BSA), polyclonal anti IBA-1 (Wako Pure Chemical Industries, 019-19741) (1:200 in 1% GS), monoclonal bFGF (Millipore 2718303) (1:200 in 0,75% HS), monoclonal FGFR1 (OriGene TA324059) (1:250 in 1% GS) and plyclonal anti-vWF (Chemicon, AB7356) (1:250 in 1% BSA).

Techniques: Western Blot, Staining

Journal: Journal of Biological Chemistry

Article Title: Acidic Fibroblast Growth Factor (FGF) Potentiates Glial-mediated Neurotoxicity by Activating FGFR2 IIIb Protein

doi: 10.1074/jbc.m111.270470

Figure Lengend Snippet: FIGURE 1. A, expression of mRNAs of the four FGF receptor genes (FGFR1–4) in various cells. The lanes are: 1, 100-bp ladder; 2, microglia; 3, astrocytes; 4, THP-1 cells; 5, U373 cells; and 6, SH-SY5Y cells. GAPDH loading controls are shown in the lower panel. Expression of mRNAs (B) and proteins (C) for the two spliced variants of FGFR1 and -2 (FGFR1 IIIb, FGFR1 IIIc, FGFR2 IIIb, FGFR2 IIIc), in glial cells are indicated. B, lane 1, 100-bp ladder; 2, microglia; 3, astrocytes; 4, THP-1 cells; and 5, U373 cells. GAPDH loading controls are shown in the lower panel. Equalization of sample loading was assessed independently using GAPDH as the housekeeping protein. Three independent experiments were performed and these were representatives. C, 1, microglia; 2, astrocytes; 3, THP-1 cells; and 4, U373 cells. Equalization of sample loading was assessed independently using -tubulin as the housekeeping protein. Three independent experiments were performed and these were representatives. D and E, quantitative results. Values are mean S.E., n 3. D, notice that FGFR2 is the major FGFR in glial cells, but SH-SY5Y cells express small amounts of all four FGFR genes. E, FGFR2 IIIb is the major receptor in all the glial cells examined.

Article Snippet: For protein expression of FGFR1 IIIb, FGFR1 IIIc, FGFR2 IIIb, and FGFR2 IIIc, the following antibodies were utilized: monoclonal anti-FGFR1 antibody (MAB658, R&D Systems, 1/2,000), monoclonal anti-FGFR2 (MAB6843, R&D Systems, 1/2,000) and, as the secondary antibody, anti-mouse IgG (A3682, Sigma, 1:3000).

Techniques: Expressing

Journal: Journal of Advanced Research

Article Title: Efficient improvement of the proliferation, differentiation, and anti-arthritic capacity of mesenchymal stem cells by simply culturing on the immobilized FGF2 derived peptide, 44-ERGVVSIKGV-53

doi: 10.1016/j.jare.2023.09.041

Figure Lengend Snippet: Antibody list.

Article Snippet: T-FGFR1 , , 1:1,000 , Novus Biologicals , NBP2-33784.

Techniques: Staining, Plasmid Preparation

Journal: Molecular Cancer

Article Title: Targeting the heparin-binding domain of fibroblast growth factor receptor 1 as a potential cancer therapy

doi: 10.1186/s12943-015-0391-4

Figure Lengend Snippet: Expression of FGFR1 in cells from breast and bone tissue. a , FGFR mRNA transcript levels in normal and cancer cells. b , Fold change in FGFR mRNA based on expression in cancers cells relative to normal cells. c , FGFR protein levels in normal versus cancer cells. Results are from triplicate experiments and the Western blot is representative of the triplicates. (* p < 0.05)

Article Snippet: The anti-FGFR1 antibody (clone # MAB765, R&D Systems) is one of the very few neutralizing antibodies against FGFR1 that have reached the market.

Techniques: Expressing, Western Blot

Journal: Molecular Cancer

Article Title: Targeting the heparin-binding domain of fibroblast growth factor receptor 1 as a potential cancer therapy

doi: 10.1186/s12943-015-0391-4

Figure Lengend Snippet: Specificity of IMB-R1 for FGFR1. a , Binding of IMB-R1 and MAB765 to FGFR1 isoforms (representative blot from triplicate experiments). b , Affinity of IMB-R1 for FGFR isoforms (results are from triplicate experiments). c , Binding of FGFR to heparin in the presence or absence of IMB-R1 (results are from triplicate experiments). d , Binding of FGF2 to FGFR in the presence or absence of IMB-R1 (results are from triplicate experiments). e , FGFR phosphorylation stimulated by FGF2 in the presence or absence of IMB-R1 including the fold change of FGFR phosphorylation as determined by a comparison of means

Article Snippet: The anti-FGFR1 antibody (clone # MAB765, R&D Systems) is one of the very few neutralizing antibodies against FGFR1 that have reached the market.

Techniques: Binding Assay, Phospho-proteomics, Comparison

Journal: Molecular Cancer

Article Title: Targeting the heparin-binding domain of fibroblast growth factor receptor 1 as a potential cancer therapy

doi: 10.1186/s12943-015-0391-4

Figure Lengend Snippet: Effects of FGFR1 inhibitors on cell growth. a , Fold change in cell number 48 h after treatment with IMB-R1. b , Temporal change in MG63 cell number following treatment with IMB-R1 (1:250). c , Cell number following FGF2 treatment (20 ng/ml for MG63, 5 ng/ml for MDAMB468 and T47D) for 48 h. Cells were pre-treatment with IMB-R1 for 1 h before FGF2 treatment. d , Cell number (MG63) following 48 h treatment with IMB-R1 or antigen-purified IMB-R1. e , Cell numbers following 48 h treatment with varying doses of FGFR inhibitors. f , Cell numbers following 48 h treatment with the FGFR1 antibody (MAB765) at varying doses. Unless otherwise stated, IMB-R1 was applied at a dilution of 1:250. All experiments were performed in triplicate

Article Snippet: The anti-FGFR1 antibody (clone # MAB765, R&D Systems) is one of the very few neutralizing antibodies against FGFR1 that have reached the market.

Techniques: Purification

Journal: Molecular Cancer

Article Title: Targeting the heparin-binding domain of fibroblast growth factor receptor 1 as a potential cancer therapy

doi: 10.1186/s12943-015-0391-4

Figure Lengend Snippet: The effect of IMB-R1 on gene expression. Cells were treated with IMB-R1 (1:250 dilution) for 48 h and RNA extracted for microarray analysis. a , The heatmaps: Red, up-regulation; Green, down-regulation. b , Top 10 affected cellular functions by IMB-R1 as determined by Ingenuity Pathway Analysis. c , The number of common genes affected across the different cells. d , The antioxidant genes significantly regulated by IMB-R1. e , The proposed signaling mechanisms disrupted by IMB-R1 during FGF2/FGFR1 dependent cell growth and survival in cancer cells

Article Snippet: The anti-FGFR1 antibody (clone # MAB765, R&D Systems) is one of the very few neutralizing antibodies against FGFR1 that have reached the market.

Techniques: Gene Expression, Microarray

Journal: Molecular Cancer

Article Title: Targeting the heparin-binding domain of fibroblast growth factor receptor 1 as a potential cancer therapy

doi: 10.1186/s12943-015-0391-4

Figure Lengend Snippet: IMB-R1 targets FGFR1 in multiple human cancers. a , IMB-R1 histochemical staining from a human cancer tissue array. Right panel , enlarged image of boxed area highlighting increased FGFR1 expression (detected by IMB-R1) in breast cancer tissues from twenty separate donors compared with adjacent healthy breast tissue. b , The intensity of FGFR1 expression (detected by IMB-R1) was scored and the average scores for the various cancer tissues compared with those from adjacent healthy tissues

Article Snippet: The anti-FGFR1 antibody (clone # MAB765, R&D Systems) is one of the very few neutralizing antibodies against FGFR1 that have reached the market.

Techniques: Staining, Expressing