human tgf β1  (R&D Systems)

 
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  • 98
    Name:
    Human TGF beta 1 Protein
    Description:
    The Human TGF beta 1 Protein from R D Systems is derived from Human Platelets The Human TGF beta 1 Protein has been validated for the following applications Bioactivity
    Catalog Number:
    100-B-001
    Price:
    289
    Category:
    Proteins and Enzymes
    Source:
    Human Platelets
    Applications:
    Bioactivity
    Purity:
    >97%, by SDS-PAGE under reducing conditions and visualized by silver stain.
    Conjugate:
    Unconjugated
    Size:
    1 ug
    Buy from Supplier


    Structured Review

    R&D Systems human tgf β1
    Human TGF beta 1 Protein
    The Human TGF beta 1 Protein from R D Systems is derived from Human Platelets The Human TGF beta 1 Protein has been validated for the following applications Bioactivity
    https://www.bioz.com/result/human tgf β1/product/R&D Systems
    Average 98 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    human tgf β1 - by Bioz Stars, 2021-05
    98/100 stars

    Images

    1) Product Images from "Clinical and biochemical profiles suggest fibromuscular dysplasia is a systemic disease with altered TGF-β expression and connective tissue features"

    Article Title: Clinical and biochemical profiles suggest fibromuscular dysplasia is a systemic disease with altered TGF-β expression and connective tissue features

    Journal: The FASEB Journal

    doi: 10.1096/fj.14-251207

    Western blot analysis of TGF-β pathway biomarkers. Whole-cell lysates from FMD case subject ( n =18) and control fibroblasts ( n =14) were probed for canonical pathway markers pSmad2 and pSmad1/5/8, treated with TGF-β1 or BMP-4, respectively,
    Figure Legend Snippet: Western blot analysis of TGF-β pathway biomarkers. Whole-cell lysates from FMD case subject ( n =18) and control fibroblasts ( n =14) were probed for canonical pathway markers pSmad2 and pSmad1/5/8, treated with TGF-β1 or BMP-4, respectively,

    Techniques Used: Western Blot

    Circulating and secreted TGF-β in FMD. A ) Elevated levels of total TGF-β1 and -β2 in platelet-poor EDTA-plasma from FMD case subjects vs. age- and sex-matched healthy controls. Samples were acid-activated prior to assaying by ELISA.
    Figure Legend Snippet: Circulating and secreted TGF-β in FMD. A ) Elevated levels of total TGF-β1 and -β2 in platelet-poor EDTA-plasma from FMD case subjects vs. age- and sex-matched healthy controls. Samples were acid-activated prior to assaying by ELISA.

    Techniques Used: Enzyme-linked Immunosorbent Assay

    2) Product Images from "Smad7 suppresses renal fibrosis via altering expression of TGF-?/Smad3-regulated microRNAs"

    Article Title: Smad7 suppresses renal fibrosis via altering expression of TGF-?/Smad3-regulated microRNAs

    Journal: Molecular Therapy

    doi: 10.1038/mt.2012.251

    Delivery of miR-21 mimic re-induces transforming growth factor-β1 (TGF-β1)-induced expression of fibrotic markers in Smad7 overexpressing tubular epithelial cells (TECs) . ( a ) Real-time PCR analysis of miR-21 in TECs at 24 hours after TGF-β1
    Figure Legend Snippet: Delivery of miR-21 mimic re-induces transforming growth factor-β1 (TGF-β1)-induced expression of fibrotic markers in Smad7 overexpressing tubular epithelial cells (TECs) . ( a ) Real-time PCR analysis of miR-21 in TECs at 24 hours after TGF-β1

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction

    3) Product Images from "Epithelial contribution to the profibrotic stiff microenvironment and myofibroblast population in lung fibrosis"

    Article Title: Epithelial contribution to the profibrotic stiff microenvironment and myofibroblast population in lung fibrosis

    Journal: Molecular Biology of the Cell

    doi: 10.1091/mbc.E17-01-0026

    Expression of fibrillar collagens in EMT-competent lung epithelial cells and primary lung fibroblasts on stimulation with TGF-β1 for 3 d. (A, B) mRNA levels of COL1A1 (A) and COL3A1 (B) assessed by qRT-PCR using three technical replicates. POLR2A was used as endogenous gene. (C) Total secreted collagens of primary myofibroblasts assessed with the Sircol assay. Statistical analysis as in Figure 2 . Note the vertical log scale in A and B.
    Figure Legend Snippet: Expression of fibrillar collagens in EMT-competent lung epithelial cells and primary lung fibroblasts on stimulation with TGF-β1 for 3 d. (A, B) mRNA levels of COL1A1 (A) and COL3A1 (B) assessed by qRT-PCR using three technical replicates. POLR2A was used as endogenous gene. (C) Total secreted collagens of primary myofibroblasts assessed with the Sircol assay. Statistical analysis as in Figure 2 . Note the vertical log scale in A and B.

    Techniques Used: Expressing, Quantitative RT-PCR

    Assembly of α-SMA into stress fibers in EMT-competent lung epithelial cells and primary lung fibroblasts on stimulation with TGF-β1 for 3 d. (A) Quantitative colocalization analysis of the images in Figure 3 through the Manders colocalization coefficient M(α-SMA), which corresponds to the fraction (between 0 and 1) of α-SMA co-occurring with F-actin. (B) Colocalization analysis of the images in Figure 3 in terms of the percentage of cells stimulated with TGF-β1 that exhibited at least three straight and continuous white lines (green bars) or three straight and discontinuous (but not speckled) white lines (red bars), which were interpreted as apparent (well-differentiated) myofibroblasts or proto-myofibroblasts, respectively. (C) Absolute average F-actin fluorescence intensity per cell obtained on stimulation with TGF-β1 in epithelial cells undergoing weak or strong EMT and in primary fibroblasts from either control or IPF patients. Statistical analysis as in Figure 2 .
    Figure Legend Snippet: Assembly of α-SMA into stress fibers in EMT-competent lung epithelial cells and primary lung fibroblasts on stimulation with TGF-β1 for 3 d. (A) Quantitative colocalization analysis of the images in Figure 3 through the Manders colocalization coefficient M(α-SMA), which corresponds to the fraction (between 0 and 1) of α-SMA co-occurring with F-actin. (B) Colocalization analysis of the images in Figure 3 in terms of the percentage of cells stimulated with TGF-β1 that exhibited at least three straight and continuous white lines (green bars) or three straight and discontinuous (but not speckled) white lines (red bars), which were interpreted as apparent (well-differentiated) myofibroblasts or proto-myofibroblasts, respectively. (C) Absolute average F-actin fluorescence intensity per cell obtained on stimulation with TGF-β1 in epithelial cells undergoing weak or strong EMT and in primary fibroblasts from either control or IPF patients. Statistical analysis as in Figure 2 .

    Techniques Used: Fluorescence

    Immunofluorescence analysis of epithelial (E-cadherin) and mesenchymal (vimentin) markers in EMT-competent lung epithelial cells and primary lung fibroblasts. (A) Representative immunofluorescence images of E-cadherin (left) and vimentin (right) in epithelial cell lines (H441, A549) and primary fibroblasts obtained from randomly selected control (fibrosis-free) or IPF patients (referred by the three-digit number following # symbol) cultured for 3 d in the absence or presence of 5 ng/ml TGF-β1. Images were obtained with a ×20 objective (seven images/condition). Blue indicates DAPI staining here and thereafter. Scale bar here, 50 μm. Representative images from other epithelial cell lines (PC9, H1975) and patients are shown in Supplemental Figure S1. (B) Ratio of the average vimentin fluorescence intensity per cell obtained in the presence or absence of TGF-β1 in a panel of epithelial cell lines and primary fibroblasts from randomly selected control and IPF patients. Horizontal gray dashed line corresponds to no fold change (ratio = 1), whereas horizontal solid blue lines correspond to the average values for each cellular model (i.e., epithelial cells exhibiting weak or strong EMT, n = 2; and primary fibroblasts from control or IPF patients, n = 3). (C) Absolute average vimentin fluorescence intensity per cell obtained on stimulation with TGF-β1 in the same cell models shown in B. * p ≤ 0.05 with respect to 1 were determined by Student’s t test (here and hereafter).
    Figure Legend Snippet: Immunofluorescence analysis of epithelial (E-cadherin) and mesenchymal (vimentin) markers in EMT-competent lung epithelial cells and primary lung fibroblasts. (A) Representative immunofluorescence images of E-cadherin (left) and vimentin (right) in epithelial cell lines (H441, A549) and primary fibroblasts obtained from randomly selected control (fibrosis-free) or IPF patients (referred by the three-digit number following # symbol) cultured for 3 d in the absence or presence of 5 ng/ml TGF-β1. Images were obtained with a ×20 objective (seven images/condition). Blue indicates DAPI staining here and thereafter. Scale bar here, 50 μm. Representative images from other epithelial cell lines (PC9, H1975) and patients are shown in Supplemental Figure S1. (B) Ratio of the average vimentin fluorescence intensity per cell obtained in the presence or absence of TGF-β1 in a panel of epithelial cell lines and primary fibroblasts from randomly selected control and IPF patients. Horizontal gray dashed line corresponds to no fold change (ratio = 1), whereas horizontal solid blue lines correspond to the average values for each cellular model (i.e., epithelial cells exhibiting weak or strong EMT, n = 2; and primary fibroblasts from control or IPF patients, n = 3). (C) Absolute average vimentin fluorescence intensity per cell obtained on stimulation with TGF-β1 in the same cell models shown in B. * p ≤ 0.05 with respect to 1 were determined by Student’s t test (here and hereafter).

    Techniques Used: Immunofluorescence, Cell Culture, Staining, Fluorescence

    Visualization of the actin cytoskeleton, α-SMA, and their colocalization on stimulation with TGF-β1 for 3 d. Selected regions from representative immunofluorescence images of F-actin (left), α-SMA (middle), and their corresponding colocalization (right) in epithelial cells (H441, A549) and primary fibroblasts obtained from randomly selected control or IPF patients cultured in the absence or presence of 5 ng/ml TGF-β1. The colocalization of F-actin and α-SMA was labeled in white. Images were obtained with a ×20 objective (seven images/condition). Scale bar, 50 μm. Full images as well as additional representative images from other epithelial cell lines (PC9, H1975) and patients are shown in Supplemental Figure S4.
    Figure Legend Snippet: Visualization of the actin cytoskeleton, α-SMA, and their colocalization on stimulation with TGF-β1 for 3 d. Selected regions from representative immunofluorescence images of F-actin (left), α-SMA (middle), and their corresponding colocalization (right) in epithelial cells (H441, A549) and primary fibroblasts obtained from randomly selected control or IPF patients cultured in the absence or presence of 5 ng/ml TGF-β1. The colocalization of F-actin and α-SMA was labeled in white. Images were obtained with a ×20 objective (seven images/condition). Scale bar, 50 μm. Full images as well as additional representative images from other epithelial cell lines (PC9, H1975) and patients are shown in Supplemental Figure S4.

    Techniques Used: Immunofluorescence, Cell Culture, Labeling

    Nanoindentation elasticity measurements of single cells carried out with AFM on lung epithelial cells and primary lung fibroblasts. (A) Representative phase contrast image of an AFM force sensor (cantilever) on top of a single lung epithelial cell (A549). AFM force sensors were used to indent locally the cell surface and assess the corresponding opposing force to compute the Young’s modulus ( E ) as described in Material and Methods , using three technical replicates. (B) Ratio of the average E values obtained on single cells cultured in the presence or absence of TGF-β1 in a panel of epithelial cell lines ( n = 4) and primary fibroblasts from randomly selected control ( n = 5) and IPF ( n = 5) patients. On average 10 cells were measured per experimental condition (two independent experiments). Horizontal gray dashed line corresponds to no fold change (ratio = 1), whereas horizontal solid blue lines correspond to the average values for each cellular model (i.e., epithelial cells exhibiting weak or strong EMT, and primary fibroblasts from control or IPF patients) as in Figure 1B . The E data for each cell line and patient are shown in Supplemental Figure S5. Statistical analysis as in Figure 1B . (C) E values obtained on single cells on TGF-β1 stimulation from each cellular model. Red horizontal lines correspond to the average E values for each model as in Figure 1C . The semi-transparent orange horizontal band corresponds to the E values assessed on normal pulmonary tissue elsewhere (see the main text for references) and was added to illustrate the aberrant mechanical behavior of IPF fibroblasts. Statistical analysis as in Figure 2 .
    Figure Legend Snippet: Nanoindentation elasticity measurements of single cells carried out with AFM on lung epithelial cells and primary lung fibroblasts. (A) Representative phase contrast image of an AFM force sensor (cantilever) on top of a single lung epithelial cell (A549). AFM force sensors were used to indent locally the cell surface and assess the corresponding opposing force to compute the Young’s modulus ( E ) as described in Material and Methods , using three technical replicates. (B) Ratio of the average E values obtained on single cells cultured in the presence or absence of TGF-β1 in a panel of epithelial cell lines ( n = 4) and primary fibroblasts from randomly selected control ( n = 5) and IPF ( n = 5) patients. On average 10 cells were measured per experimental condition (two independent experiments). Horizontal gray dashed line corresponds to no fold change (ratio = 1), whereas horizontal solid blue lines correspond to the average values for each cellular model (i.e., epithelial cells exhibiting weak or strong EMT, and primary fibroblasts from control or IPF patients) as in Figure 1B . The E data for each cell line and patient are shown in Supplemental Figure S5. Statistical analysis as in Figure 1B . (C) E values obtained on single cells on TGF-β1 stimulation from each cellular model. Red horizontal lines correspond to the average E values for each model as in Figure 1C . The semi-transparent orange horizontal band corresponds to the E values assessed on normal pulmonary tissue elsewhere (see the main text for references) and was added to illustrate the aberrant mechanical behavior of IPF fibroblasts. Statistical analysis as in Figure 2 .

    Techniques Used: Cell Culture

    Analysis of FAK Y397 activity in primary fibroblasts on TGF-β1 stimulation, and effect of loss of FAK activity in the stiffness, cytoskeletal organization, and collagen expression of fibroblasts. (A) Representative Western blot showing the time-course expression of FAK Y397 , total FAK, and β-actin in primary control fibroblasts stimulated with TGF-β1. (B) Representative Western blot showing FAK pY397 and total FAK of control fibroblasts and IPF fibroblasts stimulated with TGF-β1 for 6 h; right: corresponding densitometry analysis of FAK pY397 /FAK in control ( n = 4) and IPF fibroblasts ( n = 4). (C) Representative Western blot showing FAK pY397 , total FAK and α-tubulin of FAK wild-type (FAK +/+ ), or FAK null (FAK –/– ) mouse embryonic fibroblasts stimulated with TGF-β1 for 6 h; right: corresponding densitometry analysis of FAK pY397 /α-tubulin ( n = 3). (D) E of FAK +/+ and FAK –/– fibroblasts stimulated with TGF-β1 for 3 d assessed by AFM as in Figure 5 . (E) Selected regions from representative immunofluorescence images of F-actin (left), α-SMA (middle), and their corresponding colocalization (right) of FAK +/+ and FAK –/– fibroblasts stimulated with TGF-β1 for 3 d. Images were obtained as in Figure 3 . Scale bar, 50 μm. (F, G) mRNA levels of Col1a1 (F) and Col3a1 (G) of FAK +/+ and FAK –/– fibroblasts stimulated with TGF-β1 for 3 d assessed as in Figure 6 . Statistical analysis as in Figure 2 ( n = 2, unless otherwise indicated).
    Figure Legend Snippet: Analysis of FAK Y397 activity in primary fibroblasts on TGF-β1 stimulation, and effect of loss of FAK activity in the stiffness, cytoskeletal organization, and collagen expression of fibroblasts. (A) Representative Western blot showing the time-course expression of FAK Y397 , total FAK, and β-actin in primary control fibroblasts stimulated with TGF-β1. (B) Representative Western blot showing FAK pY397 and total FAK of control fibroblasts and IPF fibroblasts stimulated with TGF-β1 for 6 h; right: corresponding densitometry analysis of FAK pY397 /FAK in control ( n = 4) and IPF fibroblasts ( n = 4). (C) Representative Western blot showing FAK pY397 , total FAK and α-tubulin of FAK wild-type (FAK +/+ ), or FAK null (FAK –/– ) mouse embryonic fibroblasts stimulated with TGF-β1 for 6 h; right: corresponding densitometry analysis of FAK pY397 /α-tubulin ( n = 3). (D) E of FAK +/+ and FAK –/– fibroblasts stimulated with TGF-β1 for 3 d assessed by AFM as in Figure 5 . (E) Selected regions from representative immunofluorescence images of F-actin (left), α-SMA (middle), and their corresponding colocalization (right) of FAK +/+ and FAK –/– fibroblasts stimulated with TGF-β1 for 3 d. Images were obtained as in Figure 3 . Scale bar, 50 μm. (F, G) mRNA levels of Col1a1 (F) and Col3a1 (G) of FAK +/+ and FAK –/– fibroblasts stimulated with TGF-β1 for 3 d assessed as in Figure 6 . Statistical analysis as in Figure 2 ( n = 2, unless otherwise indicated).

    Techniques Used: Activity Assay, Expressing, Western Blot, Immunofluorescence

    Morphometric analysis of EMT-competent lung epithelial cells and primary lung fibroblasts stimulated with the profibrotic cytokine TGF-β1 for 3 d. (A) Representative phase contrast images of epithelial cells (H441, A549) and primary fibroblasts obtained from randomly selected control or IPF patients in the absence or presence of TGF-β1. Images were obtained with a ×20 objective (two to three images/condition). Scale bar, 50 μm. Representative images from other epithelial cell lines (PC9, H1975) and patients are shown in Supplemental Figure S3. (B–D) Average cell values of the circularity (B), elongation (C), and spreading (D) assessed on stimulation with TGF-β1 in epithelial cells undergoing weak or strong EMT and in primary fibroblasts from control or IPF patients. On average, 30 cells were analyzed per experimental condition. * p ≤ 0.05, ** p ≤ 0.01, and *** p ≤ 0.005 with respect to IPF fibroblasts; # p ≤ 0.05, ## p ≤ 0.01, and ### p ≤ 0.005 with respect to control fibroblasts. All comparisons were determined by Student’s t test.
    Figure Legend Snippet: Morphometric analysis of EMT-competent lung epithelial cells and primary lung fibroblasts stimulated with the profibrotic cytokine TGF-β1 for 3 d. (A) Representative phase contrast images of epithelial cells (H441, A549) and primary fibroblasts obtained from randomly selected control or IPF patients in the absence or presence of TGF-β1. Images were obtained with a ×20 objective (two to three images/condition). Scale bar, 50 μm. Representative images from other epithelial cell lines (PC9, H1975) and patients are shown in Supplemental Figure S3. (B–D) Average cell values of the circularity (B), elongation (C), and spreading (D) assessed on stimulation with TGF-β1 in epithelial cells undergoing weak or strong EMT and in primary fibroblasts from control or IPF patients. On average, 30 cells were analyzed per experimental condition. * p ≤ 0.05, ** p ≤ 0.01, and *** p ≤ 0.005 with respect to IPF fibroblasts; # p ≤ 0.05, ## p ≤ 0.01, and ### p ≤ 0.005 with respect to control fibroblasts. All comparisons were determined by Student’s t test.

    Techniques Used:

    Comprehensive comparison of the phenotype of lung cells undergoing EMT on stimulation with TGF-β1 and that of nonactivated primary fibroblasts from control donors cultured in the absence of TGF-β1 in terms of vimentin (A), F-actin (B), % of apparent proto-myofibroblasts/myofibroblasts (C), spreading (D), circularity (E), elongation (F), stiffness (G), and expression of COL1A1 (H) and COL3A1 (I). Statistical analysis as in Figure 2 .
    Figure Legend Snippet: Comprehensive comparison of the phenotype of lung cells undergoing EMT on stimulation with TGF-β1 and that of nonactivated primary fibroblasts from control donors cultured in the absence of TGF-β1 in terms of vimentin (A), F-actin (B), % of apparent proto-myofibroblasts/myofibroblasts (C), spreading (D), circularity (E), elongation (F), stiffness (G), and expression of COL1A1 (H) and COL3A1 (I). Statistical analysis as in Figure 2 .

    Techniques Used: Cell Culture, Expressing

    4) Product Images from "Transforming growth factor-? expression in human retinal pigment epithelial cells is enhanced by Toxoplasma gondii: a possible role in the immunopathogenesis of retinochoroiditis"

    Article Title: Transforming growth factor-? expression in human retinal pigment epithelial cells is enhanced by Toxoplasma gondii: a possible role in the immunopathogenesis of retinochoroiditis

    Journal: Clinical and Experimental Immunology

    doi: 10.1046/j.1365-2249.2002.01815.x

    Effect of TGF-β on T. gondii replication in HRPE. HRPE were infected with T. gondii and treated with human recombinant TGF-β1,TGF-β2 or other growth factors as described in Material and methods section. After 5 days, T. gondii replication in HRPE was quantified by counting the parasites using a haemocytometer. Results are the means ± s.e. for four experiments each with duplicate samples.
    Figure Legend Snippet: Effect of TGF-β on T. gondii replication in HRPE. HRPE were infected with T. gondii and treated with human recombinant TGF-β1,TGF-β2 or other growth factors as described in Material and methods section. After 5 days, T. gondii replication in HRPE was quantified by counting the parasites using a haemocytometer. Results are the means ± s.e. for four experiments each with duplicate samples.

    Techniques Used: Infection, Recombinant

    Time-course of total and activeTGF-β1, and TGF-β2 secretion by HRPE. Cultures were serum starved for 24 h and incubated in SFM for 1, 2, 3 or 4 days and culture supernatants were collected and the levels of active (mature) and total TGF-β1 and TGF-β2 were determined by ELISA. Results are the means ± s.e. for four experiments each with duplicate samples. ▪, TGF-β1(total); □, TGF-β1 (active); •,TGF-β2 (total); ○, TGF-β2 (active).
    Figure Legend Snippet: Time-course of total and activeTGF-β1, and TGF-β2 secretion by HRPE. Cultures were serum starved for 24 h and incubated in SFM for 1, 2, 3 or 4 days and culture supernatants were collected and the levels of active (mature) and total TGF-β1 and TGF-β2 were determined by ELISA. Results are the means ± s.e. for four experiments each with duplicate samples. ▪, TGF-β1(total); □, TGF-β1 (active); •,TGF-β2 (total); ○, TGF-β2 (active).

    Techniques Used: Incubation, Enzyme-linked Immunosorbent Assay

    RT-PCR analyses of TGF-β1 (a) and TGF-β2 (b) mRNA expression in T. gondii -infected HRPE. Total RNA was prepared from control (uninfected) and T. gondii -infected HRPE and 1 u g of RNA was used for reverse transcription and amplification (30 cycles) in a single tube using an RNA PCR kit as described in the Methods section. Control, d1; Control, d2; Control, d3 and Control, d4=uninfected cultures incubated for 1, 2, 3 or 4 days in serum free medium (SFM). T. gondii , d1; T. gondii , d2; T. gondii , d3 and T. gondii , d4=cultures infected with T. gondii for 1, 2, 3 or 4 days post-inoculation in SFM. The position of TGF-β1, TGF-β2 and GAPDH PCR products, 186, 529 and 600 bp, respectively, are indicated by arrows.
    Figure Legend Snippet: RT-PCR analyses of TGF-β1 (a) and TGF-β2 (b) mRNA expression in T. gondii -infected HRPE. Total RNA was prepared from control (uninfected) and T. gondii -infected HRPE and 1 u g of RNA was used for reverse transcription and amplification (30 cycles) in a single tube using an RNA PCR kit as described in the Methods section. Control, d1; Control, d2; Control, d3 and Control, d4=uninfected cultures incubated for 1, 2, 3 or 4 days in serum free medium (SFM). T. gondii , d1; T. gondii , d2; T. gondii , d3 and T. gondii , d4=cultures infected with T. gondii for 1, 2, 3 or 4 days post-inoculation in SFM. The position of TGF-β1, TGF-β2 and GAPDH PCR products, 186, 529 and 600 bp, respectively, are indicated by arrows.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Expressing, Infection, Amplification, Polymerase Chain Reaction, Incubation

    5) Product Images from "Transient Tumor-Fibroblast Interactions Increase Tumor Cell Malignancy by a TGF-? Mediated Mechanism in a Mouse Xenograft Model of Breast Cancer"

    Article Title: Transient Tumor-Fibroblast Interactions Increase Tumor Cell Malignancy by a TGF-? Mediated Mechanism in a Mouse Xenograft Model of Breast Cancer

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0009832

    TGF-β signaling induces scattering of CA1a cells. A. Exogenous TGF-β and fibroblast-released TGF-β induces cell scattering in dot assays. Cells were plated into dot assays, incubated over night, and then stimulated with TGF-β (5 ng/ml), EGF (100 ng/ml), or vehicle (Control) for 4 d. Use of CoCM that was derived from cocultures of CA1a cells and TGFβ1 knockout fibroblasts (CoCM(KO)) did not stimulate cell scattering while conditioned medium from cocultures employing TGF-β1 wildtype fibroblasts (CoCM(WT)) did induce cell scattering. B. CoCM as compared to TuCM and FbCM induces higher levels of CAGA (Smad3) and ARE (Smad2) mediated luciferase activity in CA1a cells (n = 3, ANOVA/Dunnett's Multiple Comparison, p = 0.0002 (ARE) and p
    Figure Legend Snippet: TGF-β signaling induces scattering of CA1a cells. A. Exogenous TGF-β and fibroblast-released TGF-β induces cell scattering in dot assays. Cells were plated into dot assays, incubated over night, and then stimulated with TGF-β (5 ng/ml), EGF (100 ng/ml), or vehicle (Control) for 4 d. Use of CoCM that was derived from cocultures of CA1a cells and TGFβ1 knockout fibroblasts (CoCM(KO)) did not stimulate cell scattering while conditioned medium from cocultures employing TGF-β1 wildtype fibroblasts (CoCM(WT)) did induce cell scattering. B. CoCM as compared to TuCM and FbCM induces higher levels of CAGA (Smad3) and ARE (Smad2) mediated luciferase activity in CA1a cells (n = 3, ANOVA/Dunnett's Multiple Comparison, p = 0.0002 (ARE) and p

    Techniques Used: Incubation, Derivative Assay, Knock-Out, Luciferase, Activity Assay

    6) Product Images from "Transforming Growth Factor ?1 (TGF-?1) Promotes Endothelial Cell Survival during In Vitro Angiogenesis via an Autocrine Mechanism Implicating TGF-? Signaling"

    Article Title: Transforming Growth Factor ?1 (TGF-?1) Promotes Endothelial Cell Survival during In Vitro Angiogenesis via an Autocrine Mechanism Implicating TGF-? Signaling

    Journal: Molecular and Cellular Biology

    doi: 10.1128/MCB.21.21.7218-7230.2001

    Inhibition of TGF-α signaling blocks tube formation and cell survival induced by TGF-β1 in collagen gels. (Top) 1G11 endothelial cells grown in collagen gels were cultured for 24 h in the presence of DMEM alone (basal) or 10 ng of TGF-β1/ml either alone or supplemented with 1 μM tyrphostin AG1478. After this time, cells were examined by phase-contrast microscopy. Magnifications: left, ×90; right, ×180. (Bottom) 1G11 endothelial cells grown in collagen gels were cultured for 24 h in the presence of DMEM either alone (lane B) or with 1 μM tyrphostin AG1478 (B+AG), in 10 ng of TGF-β1/ml either alone (TGFβ) or with 1 μM tyrphostin AG1478 (TGFβ+AG), in 50 ng of TGF-α/ml either alone (TGFα) or with tyrphostin AG1478 (TGFα+AG), or in 100 ng of FGF-2/ml either alone (FGF) or with 1 μM tyrphostin AG1478 (FGF+AG). Dead and living cells were counted as described in Materials and Methods. Results are averages of three different experiments.
    Figure Legend Snippet: Inhibition of TGF-α signaling blocks tube formation and cell survival induced by TGF-β1 in collagen gels. (Top) 1G11 endothelial cells grown in collagen gels were cultured for 24 h in the presence of DMEM alone (basal) or 10 ng of TGF-β1/ml either alone or supplemented with 1 μM tyrphostin AG1478. After this time, cells were examined by phase-contrast microscopy. Magnifications: left, ×90; right, ×180. (Bottom) 1G11 endothelial cells grown in collagen gels were cultured for 24 h in the presence of DMEM either alone (lane B) or with 1 μM tyrphostin AG1478 (B+AG), in 10 ng of TGF-β1/ml either alone (TGFβ) or with 1 μM tyrphostin AG1478 (TGFβ+AG), in 50 ng of TGF-α/ml either alone (TGFα) or with tyrphostin AG1478 (TGFα+AG), or in 100 ng of FGF-2/ml either alone (FGF) or with 1 μM tyrphostin AG1478 (FGF+AG). Dead and living cells were counted as described in Materials and Methods. Results are averages of three different experiments.

    Techniques Used: Inhibition, Cell Culture, Microscopy

    TGF-α stimulates tube formation in collagen gels in the long term and potentiates TGF-β1 action in the short term. 1G11 cells grown in collagen gels were cultured for 24 h in the presence of DMEM alone or different concentrations of TGF-β1 (2 and 5 ng/ml) in the presence or the absence of TGF-α (50 ng/ml). In parallel, 1G11 cells were grown in collagen gels for 5 days in the presence or the absence of TGF-β1 (10 ng/ml) or TGF-α (50 ng/ml). Cells were examined by phase-contrast microscopy.
    Figure Legend Snippet: TGF-α stimulates tube formation in collagen gels in the long term and potentiates TGF-β1 action in the short term. 1G11 cells grown in collagen gels were cultured for 24 h in the presence of DMEM alone or different concentrations of TGF-β1 (2 and 5 ng/ml) in the presence or the absence of TGF-α (50 ng/ml). In parallel, 1G11 cells were grown in collagen gels for 5 days in the presence or the absence of TGF-β1 (10 ng/ml) or TGF-α (50 ng/ml). Cells were examined by phase-contrast microscopy.

    Techniques Used: Cell Culture, Microscopy

    A soluble EGF receptor (IgB-1) blocks TGF-β1 stimulation of Akt and p42/p44 MAPK. Quiescent 1G11 cells were preincubated for 1 h with medium from control or soluble extracellular EGF receptor IgB-1-transfected HEK293 cells. After this period, cells were stimulated with TGF-β1 for 4 h. Cells were then lysed, and phospho-Akt, phospho p42/p44 MAPK, and p42/p44 MAPK were immunodetected as described in Materials and Methods. A representative Western blot from three different experiments is shown.
    Figure Legend Snippet: A soluble EGF receptor (IgB-1) blocks TGF-β1 stimulation of Akt and p42/p44 MAPK. Quiescent 1G11 cells were preincubated for 1 h with medium from control or soluble extracellular EGF receptor IgB-1-transfected HEK293 cells. After this period, cells were stimulated with TGF-β1 for 4 h. Cells were then lysed, and phospho-Akt, phospho p42/p44 MAPK, and p42/p44 MAPK were immunodetected as described in Materials and Methods. A representative Western blot from three different experiments is shown.

    Techniques Used: Transfection, Western Blot

    TGF-β1 induces TGF-α protein expression in 1G11 cells. Quiescent 1G11 cells were stimulated for 2 h with 10 ng of TGF-β1/ml or were not stimulated (lane B). After this time, medium was collected and proteins were precipitated and loaded on a 15% gel. As a control, 300 ng of human TGF-α was also loaded. In parallel, cells were lysed and 100 μg was loaded on the gel. TGF-α was immunodetected by using a specific antibody. A Western blot representative of three different experiments is shown.
    Figure Legend Snippet: TGF-β1 induces TGF-α protein expression in 1G11 cells. Quiescent 1G11 cells were stimulated for 2 h with 10 ng of TGF-β1/ml or were not stimulated (lane B). After this time, medium was collected and proteins were precipitated and loaded on a 15% gel. As a control, 300 ng of human TGF-α was also loaded. In parallel, cells were lysed and 100 μg was loaded on the gel. TGF-α was immunodetected by using a specific antibody. A Western blot representative of three different experiments is shown.

    Techniques Used: Expressing, Western Blot

    Tyrphostin AG1478, a specific inhibitor of the EGF receptor, blocks TGF-β1 stimulation of p42/p44 MAPK and Akt in the absence of changes in TGF-β signaling. (A) Quiescent 1G11 cells were preincubated for 15 min in the presence (+) or the absence (−) of tyrphostin AG1478 (1 μM). After this time, cells were stimulated for 4 h with 10 ng of TGF-β1/ml (in duplicate in the presence of tyrphostin AG1478) or for 1 h with 50 ng of TGF-α/ml, 10 ng of PDGF-BB/ml, 1 μM insulin, or 10% FCS or were left unstimulated (lane B). Cells were lysed, and phospho-Akt, Akt, and phospho-p42/p44 MAPK were immunodetected as described in Materials and Methods. A Western blot representative of four different experiments is shown. (B) Quiescent 1G11 cells were preincubated for 15 min in the presence (+) or the absence (−) of tyrphostin AG1478 (1 μM). After this time, cells were stimulated for the times indicated with 10 ng of TGF-β1/ml or for 15 min with 50 ng of TGF-α/ml. After lysis, phospho-Smad2, phospho-Akt, Akt, and phospho-p42/p44 MAPK were immunodetected as described in Materials and Methods. A representative Western blot is shown.
    Figure Legend Snippet: Tyrphostin AG1478, a specific inhibitor of the EGF receptor, blocks TGF-β1 stimulation of p42/p44 MAPK and Akt in the absence of changes in TGF-β signaling. (A) Quiescent 1G11 cells were preincubated for 15 min in the presence (+) or the absence (−) of tyrphostin AG1478 (1 μM). After this time, cells were stimulated for 4 h with 10 ng of TGF-β1/ml (in duplicate in the presence of tyrphostin AG1478) or for 1 h with 50 ng of TGF-α/ml, 10 ng of PDGF-BB/ml, 1 μM insulin, or 10% FCS or were left unstimulated (lane B). Cells were lysed, and phospho-Akt, Akt, and phospho-p42/p44 MAPK were immunodetected as described in Materials and Methods. A Western blot representative of four different experiments is shown. (B) Quiescent 1G11 cells were preincubated for 15 min in the presence (+) or the absence (−) of tyrphostin AG1478 (1 μM). After this time, cells were stimulated for the times indicated with 10 ng of TGF-β1/ml or for 15 min with 50 ng of TGF-α/ml. After lysis, phospho-Smad2, phospho-Akt, Akt, and phospho-p42/p44 MAPK were immunodetected as described in Materials and Methods. A representative Western blot is shown.

    Techniques Used: Western Blot, Lysis

    Stimulation with TGF-β1 causes EGF receptor activation. (A) Quiescent 1G11 cells were stimulated or not (lane B) for the indicated times with 10 ng of TGF-β1/ml and for 10 min with 25 ng of FGF-2/ml, 10 ng of EGF/ml, or 10 ng of PDGF-BB/ml. Cells were lysed, and proteins were incubated with Sepharose-WGL for 1 h. After being washed, the final pellet was resuspended in Laemmli sample buffer and loaded on a SDS–7.5% polyacrylamide gel. Phosphotyrosine-containing proteins were immunodetected by using a specific antibody. Arrow, phosphotyrosine-containing protein that appeared after TGF-β1 treatment. A representative Western blot is shown. (B) Cells were treated as for panel A and lysed, and the EGF receptor (EGFR) was immunoprecipitated (IP) by incubation with a specific anti-EGF receptor antibody preadsorbed to protein A-Sepharose beads. After being washed the pellet was treated as for panel A. WB, Western blot.
    Figure Legend Snippet: Stimulation with TGF-β1 causes EGF receptor activation. (A) Quiescent 1G11 cells were stimulated or not (lane B) for the indicated times with 10 ng of TGF-β1/ml and for 10 min with 25 ng of FGF-2/ml, 10 ng of EGF/ml, or 10 ng of PDGF-BB/ml. Cells were lysed, and proteins were incubated with Sepharose-WGL for 1 h. After being washed, the final pellet was resuspended in Laemmli sample buffer and loaded on a SDS–7.5% polyacrylamide gel. Phosphotyrosine-containing proteins were immunodetected by using a specific antibody. Arrow, phosphotyrosine-containing protein that appeared after TGF-β1 treatment. A representative Western blot is shown. (B) Cells were treated as for panel A and lysed, and the EGF receptor (EGFR) was immunoprecipitated (IP) by incubation with a specific anti-EGF receptor antibody preadsorbed to protein A-Sepharose beads. After being washed the pellet was treated as for panel A. WB, Western blot.

    Techniques Used: Activation Assay, Incubation, Western Blot, Immunoprecipitation

    TGF-β1 and complete medium stimulate 1G11 capillary endothelial cells to form a tubular network in type I collagen gels. 1G11 cells were mixed with type I collagen and placed on culture plates to polymerize. After gel formation, DMEM alone (basal), complete medium (20% FCS and 150 μg of endothelial cell growth supplement/ml), or TGF-β1 (10 ng/ml) was added for 48 h and gels were examined by phase-contrast microscopy.
    Figure Legend Snippet: TGF-β1 and complete medium stimulate 1G11 capillary endothelial cells to form a tubular network in type I collagen gels. 1G11 cells were mixed with type I collagen and placed on culture plates to polymerize. After gel formation, DMEM alone (basal), complete medium (20% FCS and 150 μg of endothelial cell growth supplement/ml), or TGF-β1 (10 ng/ml) was added for 48 h and gels were examined by phase-contrast microscopy.

    Techniques Used: Microscopy

    LY-294002 and PD-98059 prevent TGF-β1-induced tube formation and Ets-1 induction. (A) 1G11 endothelial cells immersed in collagen gels were cultured for 24 h in the presence of DMEM alone (basal) or 10 ng of TGF-β1/ml either alone or with 10 μM SB202190, 15 μM LY-294002 (Ly), 10 nM rapamycin (rapa), or 30 μM PD-98059 (PD). Cells were examined by phase-contrast microscopy. Magnification, ×128. (B) 1G11 cells grown in collagen gels for 24 h in the presence of 10 ng of TGF-β1/ml either alone (TGF-β1 lane −) or with 15 μM LY-294002 (lane +Ly) or 30 μM PD-98059 (lane +PD) or in the absence of TGF-β (B lane −) were lysed, and Ets-1 was detected by immunoblotting with a specific antibody. Identical amounts of protein were loaded on the gel. The decrease in p42 MAPK content in lanes +Ly and +PD reflects partial cell death. A representative Western blot of three different experiments is shown.
    Figure Legend Snippet: LY-294002 and PD-98059 prevent TGF-β1-induced tube formation and Ets-1 induction. (A) 1G11 endothelial cells immersed in collagen gels were cultured for 24 h in the presence of DMEM alone (basal) or 10 ng of TGF-β1/ml either alone or with 10 μM SB202190, 15 μM LY-294002 (Ly), 10 nM rapamycin (rapa), or 30 μM PD-98059 (PD). Cells were examined by phase-contrast microscopy. Magnification, ×128. (B) 1G11 cells grown in collagen gels for 24 h in the presence of 10 ng of TGF-β1/ml either alone (TGF-β1 lane −) or with 15 μM LY-294002 (lane +Ly) or 30 μM PD-98059 (lane +PD) or in the absence of TGF-β (B lane −) were lysed, and Ets-1 was detected by immunoblotting with a specific antibody. Identical amounts of protein were loaded on the gel. The decrease in p42 MAPK content in lanes +Ly and +PD reflects partial cell death. A representative Western blot of three different experiments is shown.

    Techniques Used: Cell Culture, Microscopy, Western Blot

    TGF-β1 stimulates endothelial cell survival in collagen gels. (A) 1G11 cells immersed in collagen gels were cultured for 24 h in the presence of DMEM alone (B), 15 μM LY-294002 alone (B+Ly), 30 μM PD-98059 alone (B+PD), 10 ng of TGF-β1/ml (TGFβ1), and TGF-β1 in the presence of 15 μM LY-294002 (TGFβ1+Ly) or 30 μM PD-98059 (TGFβ1+PD). After this time, live and dead cells were counted as described in Materials and Methods. Results are expressed as the ratios of live cells/dead cells and are averages of eight different experiments. (B) Proliferative 1G11 cells (0) or cells immersed in collagen gels for 8 h in the absence or presence of TGF-β1 (10 ng/ml), LY-294002 (15 μM), or PD-98059 (30 μM) were lysed, and PARP was detected by immunoblotting with a specific antibody. A representative Western blot is shown. The observed difference in the mobility of PARP between proliferative cells and cells immersed in collagen gels is due to the presence of collagen in the SDS-polyacrylamide gel electrophoresis.
    Figure Legend Snippet: TGF-β1 stimulates endothelial cell survival in collagen gels. (A) 1G11 cells immersed in collagen gels were cultured for 24 h in the presence of DMEM alone (B), 15 μM LY-294002 alone (B+Ly), 30 μM PD-98059 alone (B+PD), 10 ng of TGF-β1/ml (TGFβ1), and TGF-β1 in the presence of 15 μM LY-294002 (TGFβ1+Ly) or 30 μM PD-98059 (TGFβ1+PD). After this time, live and dead cells were counted as described in Materials and Methods. Results are expressed as the ratios of live cells/dead cells and are averages of eight different experiments. (B) Proliferative 1G11 cells (0) or cells immersed in collagen gels for 8 h in the absence or presence of TGF-β1 (10 ng/ml), LY-294002 (15 μM), or PD-98059 (30 μM) were lysed, and PARP was detected by immunoblotting with a specific antibody. A representative Western blot is shown. The observed difference in the mobility of PARP between proliferative cells and cells immersed in collagen gels is due to the presence of collagen in the SDS-polyacrylamide gel electrophoresis.

    Techniques Used: Cell Culture, Western Blot, Polyacrylamide Gel Electrophoresis

    TGF-β1 stimulates PI3K, p70 S6K, and p42/p44 MAPK activities in endothelial cells grown in collagen gels and in two-dimensional cultures. (A) 1G11 cells were grown in collagen gels for the periods of time indicated. Cells were lysed, and phospho-Akt, phospho-p42/p44 MAPK (pp42/pp44 MAPK), and p42 MAPK were detected by immunoblotting with specific antibodies. A representative Western blot is shown. (B) 1G11 cells were grown on collagen- or gelatin-coated plates until confluence. After depletion of growth factors, cells were stimulated with 10 ng of TGF-β1/ml for the periods of time indicated or with 20 ng of EGF for 30 min. Cells were lysed, and Western blotting was performed using anti-phospho-Akt, anti-phospho-p42/p44 MAPK, or p42/p44 MAPK. The same extracts were loaded on an SDS–9% polyacrylamide gel (shift-up) and blotted with an anti-p70 S6K antibody. Hyperphosphorylated and active forms of p70 S6K (arrows) migrated more slowly than hypophosphorylated forms. The Western blots are representative of three independent experiments.
    Figure Legend Snippet: TGF-β1 stimulates PI3K, p70 S6K, and p42/p44 MAPK activities in endothelial cells grown in collagen gels and in two-dimensional cultures. (A) 1G11 cells were grown in collagen gels for the periods of time indicated. Cells were lysed, and phospho-Akt, phospho-p42/p44 MAPK (pp42/pp44 MAPK), and p42 MAPK were detected by immunoblotting with specific antibodies. A representative Western blot is shown. (B) 1G11 cells were grown on collagen- or gelatin-coated plates until confluence. After depletion of growth factors, cells were stimulated with 10 ng of TGF-β1/ml for the periods of time indicated or with 20 ng of EGF for 30 min. Cells were lysed, and Western blotting was performed using anti-phospho-Akt, anti-phospho-p42/p44 MAPK, or p42/p44 MAPK. The same extracts were loaded on an SDS–9% polyacrylamide gel (shift-up) and blotted with an anti-p70 S6K antibody. Hyperphosphorylated and active forms of p70 S6K (arrows) migrated more slowly than hypophosphorylated forms. The Western blots are representative of three independent experiments.

    Techniques Used: Western Blot, Electrophoretic Mobility Shift Assay

    TGF-β1 induces TGF-α mRNA expression in endothelial cells. (A) 1G11 and H5V cells stimulated with 10 ng of TGF-β1/ml for the indicated times or not stimulated were lysed, and poly(A) + mRNA was isolated. Gels were loaded with 2 μg of mRNA, and Northern blotting was performed using a 190-bp PCR-amplified fragment as a TGF-α-specific probe. Rat GAPDH (glyceraldehyde-3-phosphate dehydrogenase) was used as a control. A representative result is shown. (B) HUVEC stimulated with 10 ng of TGF-β1/ml for 2 h or not stimulated were lysed, and poly(A) + mRNA was isolated. After cDNA was obtained, PCR was performed using specific primers for human TGF-α or actin (as a control). Samples were loaded on a 2% agarose gel. 1 and 2, two independent preparations of HUVEC poly(A) + for unstimulated and stimulated cells.
    Figure Legend Snippet: TGF-β1 induces TGF-α mRNA expression in endothelial cells. (A) 1G11 and H5V cells stimulated with 10 ng of TGF-β1/ml for the indicated times or not stimulated were lysed, and poly(A) + mRNA was isolated. Gels were loaded with 2 μg of mRNA, and Northern blotting was performed using a 190-bp PCR-amplified fragment as a TGF-α-specific probe. Rat GAPDH (glyceraldehyde-3-phosphate dehydrogenase) was used as a control. A representative result is shown. (B) HUVEC stimulated with 10 ng of TGF-β1/ml for 2 h or not stimulated were lysed, and poly(A) + mRNA was isolated. After cDNA was obtained, PCR was performed using specific primers for human TGF-α or actin (as a control). Samples were loaded on a 2% agarose gel. 1 and 2, two independent preparations of HUVEC poly(A) + for unstimulated and stimulated cells.

    Techniques Used: Expressing, Isolation, Northern Blot, Polymerase Chain Reaction, Amplification, Agarose Gel Electrophoresis

    Effect of TGF-β1 on Akt and p42/p44 MAPK activation depends on mRNA and protein synthesis and vesicular secretion. Quiescent 1G11 cells were preincubated for 15 min in the presence of 5 μg of actinomycin D/ml, 10 μg of cycloheximide/ml, or 1 μg of brefeldin A/ml or in the absence of inhibitors (−), followed by a 4-h stimulation with 10 ng of TGF-β1/ml, a 30-min stimulation with 10 ng of PDGF-BB/ml, or no stimulation (basal). Cells were lysed, and phospho-Akt and Akt were immunodetected as previously (76) described. A Western blot representative of three different experiments is shown.
    Figure Legend Snippet: Effect of TGF-β1 on Akt and p42/p44 MAPK activation depends on mRNA and protein synthesis and vesicular secretion. Quiescent 1G11 cells were preincubated for 15 min in the presence of 5 μg of actinomycin D/ml, 10 μg of cycloheximide/ml, or 1 μg of brefeldin A/ml or in the absence of inhibitors (−), followed by a 4-h stimulation with 10 ng of TGF-β1/ml, a 30-min stimulation with 10 ng of PDGF-BB/ml, or no stimulation (basal). Cells were lysed, and phospho-Akt and Akt were immunodetected as previously (76) described. A Western blot representative of three different experiments is shown.

    Techniques Used: Activation Assay, Western Blot

    7) Product Images from "Non–fibro-adipogenic pericytes from human embryonic stem cells attenuate degeneration of the chronically injured mouse muscle"

    Article Title: Non–fibro-adipogenic pericytes from human embryonic stem cells attenuate degeneration of the chronically injured mouse muscle

    Journal: JCI Insight

    doi: 10.1172/jci.insight.125334

    Human embryonic stem cell derivatives are lineage-restricted pericytes. ( A ) Representative flow cytometry dot plot of CD34, CD45, and CD146 expression by hES cell derivatives expanded in EGM-2 medium at passage 0. Flow cytometry analysis of expression of characteristic cell surface markers: PDGFR-β, CD73, and CD146 (pericytic); CD45 (hematopoietic); and CD34 (endothelial, hematopoietic progenitors and adventitial cells) by expanded hES derivatives at passages 2 and 9. ( B ) Vasculogenic features of CD146 + PC-like cells from hES are demonstrated by tube formation on Matrigel. ( C ) Representative dot plots of flow cytometry analysis of CD34, PDGFR-α, CD146, and CD56 expression by expanded hES pericytic derivatives. ( D – K ) hES PCs lack fibro-adipogenic differentiation capability. Adipocytes are not detected in control ( D ) and adipogenic cultures ( E ). Negative Oil Red O staining of non-induced control ( F ) and adipogenic induced cultures ( G ). Representative images of picrosirius red staining for collagen of control ( H ) and TGF-β1–treated ( I ) hES PC cultures. Spectrophotometric quantification of collagen production (mean ± SEM) by non-induced and TGF-β1–induced hES PC cultures at the indicated time points ( J ). # P
    Figure Legend Snippet: Human embryonic stem cell derivatives are lineage-restricted pericytes. ( A ) Representative flow cytometry dot plot of CD34, CD45, and CD146 expression by hES cell derivatives expanded in EGM-2 medium at passage 0. Flow cytometry analysis of expression of characteristic cell surface markers: PDGFR-β, CD73, and CD146 (pericytic); CD45 (hematopoietic); and CD34 (endothelial, hematopoietic progenitors and adventitial cells) by expanded hES derivatives at passages 2 and 9. ( B ) Vasculogenic features of CD146 + PC-like cells from hES are demonstrated by tube formation on Matrigel. ( C ) Representative dot plots of flow cytometry analysis of CD34, PDGFR-α, CD146, and CD56 expression by expanded hES pericytic derivatives. ( D – K ) hES PCs lack fibro-adipogenic differentiation capability. Adipocytes are not detected in control ( D ) and adipogenic cultures ( E ). Negative Oil Red O staining of non-induced control ( F ) and adipogenic induced cultures ( G ). Representative images of picrosirius red staining for collagen of control ( H ) and TGF-β1–treated ( I ) hES PC cultures. Spectrophotometric quantification of collagen production (mean ± SEM) by non-induced and TGF-β1–induced hES PC cultures at the indicated time points ( J ). # P

    Techniques Used: Flow Cytometry, Expressing, Staining

    8) Product Images from "miR-192 Mediates TGF-?/Smad3-Driven Renal Fibrosis"

    Article Title: miR-192 Mediates TGF-?/Smad3-Driven Renal Fibrosis

    Journal: Journal of the American Society of Nephrology : JASN

    doi: 10.1681/ASN.2010020134

    TGF-β induces miR-192 expression in a time- and dosage-dependent manner in NRK52E cells. (A) Real-time PCR shows that TGF-β1 (2 ng/ml), not PBS, induces miR-192 expression in a time-dependant manner. (B) Real-time PCR demonstrates that
    Figure Legend Snippet: TGF-β induces miR-192 expression in a time- and dosage-dependent manner in NRK52E cells. (A) Real-time PCR shows that TGF-β1 (2 ng/ml), not PBS, induces miR-192 expression in a time-dependant manner. (B) Real-time PCR demonstrates that

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction

    Overexpression of miR-192 promotes but inhibition of miR-192 blocks TGF-β1–induced collagen I mRNA and protein expression in rat TECs. (A and B) Real-time PCR (A) and Western blot (B) analysis. Real-time PCR and Western blot analyses show
    Figure Legend Snippet: Overexpression of miR-192 promotes but inhibition of miR-192 blocks TGF-β1–induced collagen I mRNA and protein expression in rat TECs. (A and B) Real-time PCR (A) and Western blot (B) analysis. Real-time PCR and Western blot analyses show

    Techniques Used: Over Expression, Inhibition, Expressing, Real-time Polymerase Chain Reaction, Western Blot

    9) Product Images from "Activin-Like Kinase 5 (ALK5) Mediates Abnormal Proliferation of Vascular Smooth Muscle Cells from Patients with Familial Pulmonary Arterial Hypertension and Is Involved in the Progression of Experimental Pulmonary Arterial Hypertension Induced by Monocrotaline"

    Article Title: Activin-Like Kinase 5 (ALK5) Mediates Abnormal Proliferation of Vascular Smooth Muscle Cells from Patients with Familial Pulmonary Arterial Hypertension and Is Involved in the Progression of Experimental Pulmonary Arterial Hypertension Induced by Monocrotaline

    Journal: The American Journal of Pathology

    doi: 10.2353/ajpath.2009.080565

    PASMCs derived from iPAH patients were plated at equal cell densities in 96-well plates. Cells were starved for 48 hours before treatment with 0.625 ng/ml of TGF-β1 in growth media containing 10% (v/v) fetal calf serum. One μmol/L of SB525334
    Figure Legend Snippet: PASMCs derived from iPAH patients were plated at equal cell densities in 96-well plates. Cells were starved for 48 hours before treatment with 0.625 ng/ml of TGF-β1 in growth media containing 10% (v/v) fetal calf serum. One μmol/L of SB525334

    Techniques Used: Derivative Assay

    Enhanced transcriptional responses to TGF-β1 on iPAH PASMCs. iPAH and control PASMCs were cultured to confluence, serum-starved, and treated for 0, 1, 4, and 12 hours in the presence or absence of TGF-β1 (2 ng/ml). Cells were cultured
    Figure Legend Snippet: Enhanced transcriptional responses to TGF-β1 on iPAH PASMCs. iPAH and control PASMCs were cultured to confluence, serum-starved, and treated for 0, 1, 4, and 12 hours in the presence or absence of TGF-β1 (2 ng/ml). Cells were cultured

    Techniques Used: Cell Culture

    PASMCs derived from iPAH or normotensive patients were plated at equal cell densities in 96-well plates. Cells were serum-starved for 48 hours before treatment with 0.625 ng/ml of TGF-β1. Proliferation was measured by BrdU incorporation after
    Figure Legend Snippet: PASMCs derived from iPAH or normotensive patients were plated at equal cell densities in 96-well plates. Cells were serum-starved for 48 hours before treatment with 0.625 ng/ml of TGF-β1. Proliferation was measured by BrdU incorporation after

    Techniques Used: Derivative Assay, BrdU Incorporation Assay

    10) Product Images from "Overexpression of cathepsin K during silica-induced lung fibrosis and control by TGF-?"

    Article Title: Overexpression of cathepsin K during silica-induced lung fibrosis and control by TGF-?

    Journal: Respiratory Research

    doi: 10.1186/1465-9921-6-84

    Cat K expression is reduced in response to TGF-β1 in control and silicotic mouse lung fibroblasts. Cat K mRNA quantification in pulmonary fibroblasts of C57BL/6 mice. (A) Control fibroblasts were incubated with 1 or 10 ng cytokine/ml. Bars represent the mean of triplicate measurements of Cat K expression on the same sample. The Cat K downregulation by TGF-β was reproduced in 4 independent experiments. (B) Fibroblasts from control (pool of 10 animals) and silicotic (pool of 7 animals, silica) mice collected 2 months after instillation and incubated at least in duplicates without (non-treated) or with 10 ng TGF-β1/ml (TGF-beta). The results are representative of 2 independent experiments ( P
    Figure Legend Snippet: Cat K expression is reduced in response to TGF-β1 in control and silicotic mouse lung fibroblasts. Cat K mRNA quantification in pulmonary fibroblasts of C57BL/6 mice. (A) Control fibroblasts were incubated with 1 or 10 ng cytokine/ml. Bars represent the mean of triplicate measurements of Cat K expression on the same sample. The Cat K downregulation by TGF-β was reproduced in 4 independent experiments. (B) Fibroblasts from control (pool of 10 animals) and silicotic (pool of 7 animals, silica) mice collected 2 months after instillation and incubated at least in duplicates without (non-treated) or with 10 ng TGF-β1/ml (TGF-beta). The results are representative of 2 independent experiments ( P

    Techniques Used: Expressing, Mouse Assay, Incubation

    TGF-β1 downregulates Cat K mRNA expression in human lung fibroblasts. Quantification of Cat K mRNA in human pulmonary fibroblasts incubated with 10 ng TGF-β1/ml (TGF-beta) or non-treated. Values are presented as means ± SEM (5 culture-wells for each condition, P = 0.053).
    Figure Legend Snippet: TGF-β1 downregulates Cat K mRNA expression in human lung fibroblasts. Quantification of Cat K mRNA in human pulmonary fibroblasts incubated with 10 ng TGF-β1/ml (TGF-beta) or non-treated. Values are presented as means ± SEM (5 culture-wells for each condition, P = 0.053).

    Techniques Used: Expressing, Incubation

    11) Product Images from "Induction of Colonic Regulatory T Cells by Mesalamine by Activating the Aryl Hydrocarbon Receptor"

    Article Title: Induction of Colonic Regulatory T Cells by Mesalamine by Activating the Aryl Hydrocarbon Receptor

    Journal: Cellular and Molecular Gastroenterology and Hepatology

    doi: 10.1016/j.jcmgh.2017.03.010

    Mesalamine increases the level of the active form of TGF-β1, which is necessary for Treg induction. ( A ) Levels of the active form of TGF-β1 (acid-) or total TGF-β1 (acid+) in colons isolated from WT mice orally treated with mesalamine for 2 weeks (n = 5 mice per group). ( B ) Colon tissue samples (1.6 mg) from WT mice orally treated with mesalamine for 2 weeks were added into the culture of TGF-β reporter cells (MFB-F11) for 24 hours with or without HTS466284 (a chemical inhibitor of TGF-β signaling); then, SEAP activities of the supernatants were measured. 10 ng/mL human TGF-β1 was used as a positive control (n = 5). ( C ) TGF-β1 mRNA levels in the colon isolated from WT mice orally treated with mesalamine for 2 weeks (n = 8 mice per group). ( D ) Levels of active TGF-β1 (acid-) or of total TGF-β1 (acid+) in the colon isolated from AhR -/- mice treated orally with mesalamine for 2 weeks (n = 4 mice per group). ( E ) Levels of active TGF-β1 (acid-) or of total TGF-β1 (acid+) in the colon isolated from bone marrow chimeric mice (BM-AhR +/+ mice and BM-AhR -/- mice) treated orally with mesalamine for 2 weeks (n = 5 mice per group). ( F , G ) TSP-1 and Integrin αV mRNA levels in the colon isolated from WT mice or AhR -/- mice orally treated with mesalamine for 2 weeks (n = 8 mice per group). ( H ) WT mice were orally treated with mesalamine with or without GW788388, an inhibitor of TGF-β type I receptor kinase, in drinking water for 2 weeks, and the percentages of Foxp3 + cells within the CD3 + CD4 + cell population isolated from the colon LP were determined (n = 8 mice per group). Error bars represent mean ± SEM. * P
    Figure Legend Snippet: Mesalamine increases the level of the active form of TGF-β1, which is necessary for Treg induction. ( A ) Levels of the active form of TGF-β1 (acid-) or total TGF-β1 (acid+) in colons isolated from WT mice orally treated with mesalamine for 2 weeks (n = 5 mice per group). ( B ) Colon tissue samples (1.6 mg) from WT mice orally treated with mesalamine for 2 weeks were added into the culture of TGF-β reporter cells (MFB-F11) for 24 hours with or without HTS466284 (a chemical inhibitor of TGF-β signaling); then, SEAP activities of the supernatants were measured. 10 ng/mL human TGF-β1 was used as a positive control (n = 5). ( C ) TGF-β1 mRNA levels in the colon isolated from WT mice orally treated with mesalamine for 2 weeks (n = 8 mice per group). ( D ) Levels of active TGF-β1 (acid-) or of total TGF-β1 (acid+) in the colon isolated from AhR -/- mice treated orally with mesalamine for 2 weeks (n = 4 mice per group). ( E ) Levels of active TGF-β1 (acid-) or of total TGF-β1 (acid+) in the colon isolated from bone marrow chimeric mice (BM-AhR +/+ mice and BM-AhR -/- mice) treated orally with mesalamine for 2 weeks (n = 5 mice per group). ( F , G ) TSP-1 and Integrin αV mRNA levels in the colon isolated from WT mice or AhR -/- mice orally treated with mesalamine for 2 weeks (n = 8 mice per group). ( H ) WT mice were orally treated with mesalamine with or without GW788388, an inhibitor of TGF-β type I receptor kinase, in drinking water for 2 weeks, and the percentages of Foxp3 + cells within the CD3 + CD4 + cell population isolated from the colon LP were determined (n = 8 mice per group). Error bars represent mean ± SEM. * P

    Techniques Used: Isolation, Mouse Assay, Positive Control

    12) Product Images from "Profibrotic up-regulation of glucose transporter 1 by TGF-β involves activation of MEK and mammalian target of rapamycin complex 2 pathways"

    Article Title: Profibrotic up-regulation of glucose transporter 1 by TGF-β involves activation of MEK and mammalian target of rapamycin complex 2 pathways

    Journal: The FASEB Journal

    doi: 10.1096/fj.201600428R

    TGF-β stimulates GLUT1 up-regulation and glucose uptake in fibroblasts. A ) AKR-2B cells were treated in the absence (−) or presence (+) of TGF-β1 (10 ng/ml), and 2-deoxyglucose uptake was determined at the indicated times ( n = 3). B ) Glucose uptake assay in TGF-β1–treated (12 h) AKR-2B fibroblast cells. Results show an increase in 2-deoxyglucose uptake, which is inhibited by the GLUT-specific inhibitor, phloretin (100 μM; n = 3). C ) Quantitative RT-PCR analyses of GLUT induction by TGF-β1 in AKR-2B cells ( n = 3). GLUT7 expression was not examined, whereas GLUTs 2 and 12 displayed no detectable expression. Dotted line indicates 1 unit on y axis. GLUTs 11 and 14 were not included, as no corresponding murine genes have been reported. The profiles of GLUT expression indicate that GLUT1 and GLUT3 exhibit > 2-fold induction by TGF-β1, although only GLUT1 protein showed a parallel increase (data not shown for GLUT3 protein). D ) AKR-2B cells were treated for the indicated times with TGF-β1 (10 ng/ml) or solvent (4 mM HCl, 1.0 mg/ml bovine serum albumin), and at the indicated times whole-cell lysates were prepared for quantitative PCR (top) or Western blot analysis (bottom) and blotted for GLUT1 or glyceraldehyde 3-phosphate dehydrogenase (GAPDH). Data show a time-dependent up-regulation of GLUT1 mRNA and protein by TGF-β1 ( n = 3). E ) Subcellular localization of GLUT1 in AKR-2B fibroblasts. Cell-surface biotinylation reveals the presence of GLUT1 in the membrane fraction under basal and TGF-β–stimulated (12 h) conditions. F provide P values for indicated data points.
    Figure Legend Snippet: TGF-β stimulates GLUT1 up-regulation and glucose uptake in fibroblasts. A ) AKR-2B cells were treated in the absence (−) or presence (+) of TGF-β1 (10 ng/ml), and 2-deoxyglucose uptake was determined at the indicated times ( n = 3). B ) Glucose uptake assay in TGF-β1–treated (12 h) AKR-2B fibroblast cells. Results show an increase in 2-deoxyglucose uptake, which is inhibited by the GLUT-specific inhibitor, phloretin (100 μM; n = 3). C ) Quantitative RT-PCR analyses of GLUT induction by TGF-β1 in AKR-2B cells ( n = 3). GLUT7 expression was not examined, whereas GLUTs 2 and 12 displayed no detectable expression. Dotted line indicates 1 unit on y axis. GLUTs 11 and 14 were not included, as no corresponding murine genes have been reported. The profiles of GLUT expression indicate that GLUT1 and GLUT3 exhibit > 2-fold induction by TGF-β1, although only GLUT1 protein showed a parallel increase (data not shown for GLUT3 protein). D ) AKR-2B cells were treated for the indicated times with TGF-β1 (10 ng/ml) or solvent (4 mM HCl, 1.0 mg/ml bovine serum albumin), and at the indicated times whole-cell lysates were prepared for quantitative PCR (top) or Western blot analysis (bottom) and blotted for GLUT1 or glyceraldehyde 3-phosphate dehydrogenase (GAPDH). Data show a time-dependent up-regulation of GLUT1 mRNA and protein by TGF-β1 ( n = 3). E ) Subcellular localization of GLUT1 in AKR-2B fibroblasts. Cell-surface biotinylation reveals the presence of GLUT1 in the membrane fraction under basal and TGF-β–stimulated (12 h) conditions. F provide P values for indicated data points.

    Techniques Used: Quantitative RT-PCR, Expressing, Real-time Polymerase Chain Reaction, Western Blot

    Autocrine activation of receptor tyrosine kinases (RTKs) is required for induction of GLUT1 by TGF-β. Quantitative RT-PCR analysis of 10 ng/ml TGF-β1–stimulated GLUT1 expression (12 h post-treatment) in AKR-2B cells in the presence of the PDGFR-specific inhibitor, CP673451 ( A ; 2 µM), the ErbB1/2-specific inhibitor, lapatinib ( B ; 5 µM), shRNA specific for PDGFRα+β ( C ), or shRNA specific for ErbB1+2 ( D ; n provide P values for indicated data points.
    Figure Legend Snippet: Autocrine activation of receptor tyrosine kinases (RTKs) is required for induction of GLUT1 by TGF-β. Quantitative RT-PCR analysis of 10 ng/ml TGF-β1–stimulated GLUT1 expression (12 h post-treatment) in AKR-2B cells in the presence of the PDGFR-specific inhibitor, CP673451 ( A ; 2 µM), the ErbB1/2-specific inhibitor, lapatinib ( B ; 5 µM), shRNA specific for PDGFRα+β ( C ), or shRNA specific for ErbB1+2 ( D ; n provide P values for indicated data points.

    Techniques Used: Activation Assay, Quantitative RT-PCR, Expressing, shRNA

    TGF-β induces GLUT1 expression dependent upon Smad pathway activation of MEK. A ) Quantitative RT-PCR analysis of GLUT1 and profibrotic gene induction by TGF-β1 (12 h post-treatment) in AKR-2B cells (pools) that stably express control (nontargeting; ctrl) or shRNA targeting Smad2 and/or Smad3 ( n = 3). Results show that both Smad2 and Smad3 are required for TGF-β1–dependent GLUT1, PAI-1, and CTGF up-regulation. Western blot analyses confirm efficient knockdown of Smad2/Smad3 and uniform activation of Smad2 and/or Smad3 in appropriate controls (5 h). B ) Western blot analysis of ERK1/2 activation by TGF-β1 (5 h post-treatment) in AKR-2B cells (pools) that stably express control (nontargeting) or shRNA targeting Smad2 and/or Smad3. Results indicate that TGF-β1 activates ERK1/2 downstream of the Smad2/3 pathway. C ) Quantitative RT-PCR analysis of GLUT1 induction by TGF-β1 (12 h post-treatment) in AKR-2B cells in the absence (−) or presence (+) of the MEK-ERK1/2 inhibitor U0126 (3 μM; n = 3). Prevention of ERK1/2 phosphorylation (5 h) confirms the efficacy of U0126, whereas the lack of effect on pSmad3 documents specificity. D ) Quantitative RT-PCR analysis of GLUT1 induction by ectopic expression of a constitutively active MEK1 (54 h post-transfection) in AKR-2B cells ( n provide P values for indicated data points.
    Figure Legend Snippet: TGF-β induces GLUT1 expression dependent upon Smad pathway activation of MEK. A ) Quantitative RT-PCR analysis of GLUT1 and profibrotic gene induction by TGF-β1 (12 h post-treatment) in AKR-2B cells (pools) that stably express control (nontargeting; ctrl) or shRNA targeting Smad2 and/or Smad3 ( n = 3). Results show that both Smad2 and Smad3 are required for TGF-β1–dependent GLUT1, PAI-1, and CTGF up-regulation. Western blot analyses confirm efficient knockdown of Smad2/Smad3 and uniform activation of Smad2 and/or Smad3 in appropriate controls (5 h). B ) Western blot analysis of ERK1/2 activation by TGF-β1 (5 h post-treatment) in AKR-2B cells (pools) that stably express control (nontargeting) or shRNA targeting Smad2 and/or Smad3. Results indicate that TGF-β1 activates ERK1/2 downstream of the Smad2/3 pathway. C ) Quantitative RT-PCR analysis of GLUT1 induction by TGF-β1 (12 h post-treatment) in AKR-2B cells in the absence (−) or presence (+) of the MEK-ERK1/2 inhibitor U0126 (3 μM; n = 3). Prevention of ERK1/2 phosphorylation (5 h) confirms the efficacy of U0126, whereas the lack of effect on pSmad3 documents specificity. D ) Quantitative RT-PCR analysis of GLUT1 induction by ectopic expression of a constitutively active MEK1 (54 h post-transfection) in AKR-2B cells ( n provide P values for indicated data points.

    Techniques Used: Expressing, Activation Assay, Quantitative RT-PCR, Stable Transfection, shRNA, Western Blot, Transfection

    Up-regulation of GLUT1 mediates TGF-β–induced profibrotic phenotype. A ) AKR-2B cells were treated in the presence (+) or absence (−) of TGF-β1 (10 ng/ml) and/or the GLUT-specific GLUT inhibitor II (Inh II; 10 μM) for 12 h, and quantitative RT-PCR was performed for PAI-1, CTGF, or α−SMA ( n = 3). Western blot of the indicated proteins (bottom). B ) TGF-β1–stimulated 2-deoxyglucose uptake in presence or absence of GLUT Inh II ( n = 3). C ) Analogous study as in panel A , except AKR-2B cells were infected with nontargeting control (ctrl) or shRNA (1 or 2) to GLUT1 ( n = 3). D ) As in panel C , 2-deoxyglucose uptake in presence or absence of control or GLUT1 shRNA ( n = 3). E ) Similar study as in panel A , using the GLUT inhibitor phloretin (100 μM; n = 3). F , G ) GLUT Inh II ( F ) or knockdown of GLUT1 by RNA interference ( G ) reduces soft agar colony formation induced by TGF-β1 ( n provide P values for indicated data points.
    Figure Legend Snippet: Up-regulation of GLUT1 mediates TGF-β–induced profibrotic phenotype. A ) AKR-2B cells were treated in the presence (+) or absence (−) of TGF-β1 (10 ng/ml) and/or the GLUT-specific GLUT inhibitor II (Inh II; 10 μM) for 12 h, and quantitative RT-PCR was performed for PAI-1, CTGF, or α−SMA ( n = 3). Western blot of the indicated proteins (bottom). B ) TGF-β1–stimulated 2-deoxyglucose uptake in presence or absence of GLUT Inh II ( n = 3). C ) Analogous study as in panel A , except AKR-2B cells were infected with nontargeting control (ctrl) or shRNA (1 or 2) to GLUT1 ( n = 3). D ) As in panel C , 2-deoxyglucose uptake in presence or absence of control or GLUT1 shRNA ( n = 3). E ) Similar study as in panel A , using the GLUT inhibitor phloretin (100 μM; n = 3). F , G ) GLUT Inh II ( F ) or knockdown of GLUT1 by RNA interference ( G ) reduces soft agar colony formation induced by TGF-β1 ( n provide P values for indicated data points.

    Techniques Used: Quantitative RT-PCR, Western Blot, Infection, shRNA

    13) Product Images from "Heat Shock Protein 90 Inhibitors Protect and Restore Pulmonary Endothelial Barrier Function"

    Article Title: Heat Shock Protein 90 Inhibitors Protect and Restore Pulmonary Endothelial Barrier Function

    Journal: American Journal of Respiratory Cell and Molecular Biology

    doi: 10.1165/rcmb.2007-0324OC

    Effects of TGF-β1 and radicicol on BPAEC cytoskeletal rearrangement. BPAEC grown on glass coverslips for 10 days were treated with ( a , e , h , l ) vehicle (0.1% DMSO), ( b , f , i , m ) RA (1 μg/ml) for 6 hours, ( c , g , j , n ) TGF-β1 (10
    Figure Legend Snippet: Effects of TGF-β1 and radicicol on BPAEC cytoskeletal rearrangement. BPAEC grown on glass coverslips for 10 days were treated with ( a , e , h , l ) vehicle (0.1% DMSO), ( b , f , i , m ) RA (1 μg/ml) for 6 hours, ( c , g , j , n ) TGF-β1 (10

    Techniques Used:

    Effect of radicicol on TGF-β1–induced p38 MAPK activation. BPAEC were treated with ( a ) vehicle (0.1% DMSO), TGF-β1 (10 ng/ml), TGF-β1 plus RA (1 μg/ml), given together, or ( b ) RA alone for the indicated periods
    Figure Legend Snippet: Effect of radicicol on TGF-β1–induced p38 MAPK activation. BPAEC were treated with ( a ) vehicle (0.1% DMSO), TGF-β1 (10 ng/ml), TGF-β1 plus RA (1 μg/ml), given together, or ( b ) RA alone for the indicated periods

    Techniques Used: Activation Assay

    Effects of TGF-β1 and hsp90 inhibitors on BPAEC transendothelial resistance (TER). BPAEC were plated on gold microelectrodes and TER was measured as described in M aterials and M ethods . ( a ) EC were treated with either vehicle (0.1% DMSO) or TGF-β1,
    Figure Legend Snippet: Effects of TGF-β1 and hsp90 inhibitors on BPAEC transendothelial resistance (TER). BPAEC were plated on gold microelectrodes and TER was measured as described in M aterials and M ethods . ( a ) EC were treated with either vehicle (0.1% DMSO) or TGF-β1,

    Techniques Used:

    Effects of TGF-β1 and radicicol on MLC and MYPT1 phosphorylation. Confluent BPAEC monolayers were incubated with vehicle (0.1% DMSO), TGF-β1 (10 ng/ml) alone or together with RA (1 μg/ml) for the indicated periods of time. ( a )
    Figure Legend Snippet: Effects of TGF-β1 and radicicol on MLC and MYPT1 phosphorylation. Confluent BPAEC monolayers were incubated with vehicle (0.1% DMSO), TGF-β1 (10 ng/ml) alone or together with RA (1 μg/ml) for the indicated periods of time. ( a )

    Techniques Used: Incubation

    Effects of TGF-β1 and radicicol on endothelial monolayer integrity. ( a–d ) bovine pulmonary arterial endothelial cells (BPAEC) were cultured in 35-mm dishes for 10 days to allow a confluent monolayer to form. Cells were then exposed for
    Figure Legend Snippet: Effects of TGF-β1 and radicicol on endothelial monolayer integrity. ( a–d ) bovine pulmonary arterial endothelial cells (BPAEC) were cultured in 35-mm dishes for 10 days to allow a confluent monolayer to form. Cells were then exposed for

    Techniques Used: Cell Culture

    Effects of TGF-β1 and radicicol on hsp27 phosphorylation and hsp90/hsp27 complex formation. BPAEC were treated with vehicle (0.1% DMSO), TGF-β1 (10 ng/ml), TGF-β1 plus RA (1 μg/ml), given together, or RA alone for 6 hours.
    Figure Legend Snippet: Effects of TGF-β1 and radicicol on hsp27 phosphorylation and hsp90/hsp27 complex formation. BPAEC were treated with vehicle (0.1% DMSO), TGF-β1 (10 ng/ml), TGF-β1 plus RA (1 μg/ml), given together, or RA alone for 6 hours.

    Techniques Used:

    14) Product Images from "Glyoxalase 1 sustains the metastatic phenotype of prostate cancer cells via EMT control. Glyoxalase 1 sustains the metastatic phenotype of prostate cancer cells via EMT control"

    Article Title: Glyoxalase 1 sustains the metastatic phenotype of prostate cancer cells via EMT control. Glyoxalase 1 sustains the metastatic phenotype of prostate cancer cells via EMT control

    Journal: Journal of Cellular and Molecular Medicine

    doi: 10.1111/jcmm.13581

    Glyoxalase 1 (Glo1) sustains the metastatic phenotype of prostate cancer cells via epithelial‐to‐mesenchymal transition (EMT) control: involvement of miR‐101, hydroimidazolone (MG‐H1), argpyrimidine (AP) and TGF‐β1/Smads signalling pathway. Glo1 up‐regulation, maintained by the decreased expression (X mark) of the tumour suppressor miR‐101, leads to MG‐H1‐AP depletion. In turn, MG‐H1‐AP depletion contributes to keep activated TGF‐β1/Smads signalling pathway that promotes EMT by inhibiting the epithelial markers E‐cadherin (E‐cad) and zonula occludens‐ (ZO‐1) and activating the mesenchymal markers vimentin (Vim) and N‐cadherin (N‐cad), together with MMP‐2 and MMP‐9, directly via Snail and/or indirectly via other TGF‐β1/Smad‐dependent EMT‐associated transcription factors. Altogether, these events sustain the invasive and migrating metastatic phenotype of prostate cancer cells
    Figure Legend Snippet: Glyoxalase 1 (Glo1) sustains the metastatic phenotype of prostate cancer cells via epithelial‐to‐mesenchymal transition (EMT) control: involvement of miR‐101, hydroimidazolone (MG‐H1), argpyrimidine (AP) and TGF‐β1/Smads signalling pathway. Glo1 up‐regulation, maintained by the decreased expression (X mark) of the tumour suppressor miR‐101, leads to MG‐H1‐AP depletion. In turn, MG‐H1‐AP depletion contributes to keep activated TGF‐β1/Smads signalling pathway that promotes EMT by inhibiting the epithelial markers E‐cadherin (E‐cad) and zonula occludens‐ (ZO‐1) and activating the mesenchymal markers vimentin (Vim) and N‐cadherin (N‐cad), together with MMP‐2 and MMP‐9, directly via Snail and/or indirectly via other TGF‐β1/Smad‐dependent EMT‐associated transcription factors. Altogether, these events sustain the invasive and migrating metastatic phenotype of prostate cancer cells

    Techniques Used: Expressing

    Glyoxalase 1 (Glo1)‐dependent MG‐H1 and argpyrimidine (AP) depletion triggers epithelial‐to‐mesenchymal transition (EMT) via TGF‐β1/SMAD4 signalling pathway in PC3 cells. (A) Glo1 silencing (siGlo1) in PC3 cells significantly affected TGF‐β1 mRNA levels, evaluated by qRT‐PCR, (B) TGF‐β1 protein intracellular levels, evaluated by Western blot, and (C) TGF‐β1 secreted levels, evaluated by a specific ELISA kit. Pre‐treatment with aminoguanidine (AG) proved Glo1‐dependent Hsp40‐modified AP‐mediated role in controlling TGF‐β1 expression (A, B, C). (D) Glo1 silencing (siGlo1) and AG pre‐treatment significantly affected Smad4 activation, evaluated both in nuclear and in cytoplasmic fractions of PC3 cells by Western blot, proving Glo1‐dependent Hsp40‐modified AP‐mediated role in controlling TGF‐β1/Smad4 signalling pathway. Inhibition of TGF‐β1 signalling by SB431542 treatment demonstrated TGF‐β1 role in controlling (E) Smad4 activation, evaluated at nuclear and cytoplasmic levels by Western blot, (F) EMT, evaluated by Western blot analysis of the epithelial markers E‐cadherin (E‐cad) and zonula occludens‐1 (ZO‐1) or mesenchymal markers vimentin (Vim), N‐cadherin (N‐cad) and Snail, (G) MMP‐2 and MMP‐9 expression, evaluated by Western blotting or activity, evaluated by the gelatin zymography, and (H) migration and invasion capabilities, evaluated by specific assays. The Western blots were obtained by the appropriate Abs. The blots were stripped off the bound Abs and reprobed with anti‐β‐actin or lamin β1 to confirm equal loading. The Western blots shown are representative of three independent experiments. The histograms indicate mean ± SD of three different cultures, and each was tested in triplicate. siCtr: control (non‐specific siRNA). (−) untreated and (+) treated cells. * P
    Figure Legend Snippet: Glyoxalase 1 (Glo1)‐dependent MG‐H1 and argpyrimidine (AP) depletion triggers epithelial‐to‐mesenchymal transition (EMT) via TGF‐β1/SMAD4 signalling pathway in PC3 cells. (A) Glo1 silencing (siGlo1) in PC3 cells significantly affected TGF‐β1 mRNA levels, evaluated by qRT‐PCR, (B) TGF‐β1 protein intracellular levels, evaluated by Western blot, and (C) TGF‐β1 secreted levels, evaluated by a specific ELISA kit. Pre‐treatment with aminoguanidine (AG) proved Glo1‐dependent Hsp40‐modified AP‐mediated role in controlling TGF‐β1 expression (A, B, C). (D) Glo1 silencing (siGlo1) and AG pre‐treatment significantly affected Smad4 activation, evaluated both in nuclear and in cytoplasmic fractions of PC3 cells by Western blot, proving Glo1‐dependent Hsp40‐modified AP‐mediated role in controlling TGF‐β1/Smad4 signalling pathway. Inhibition of TGF‐β1 signalling by SB431542 treatment demonstrated TGF‐β1 role in controlling (E) Smad4 activation, evaluated at nuclear and cytoplasmic levels by Western blot, (F) EMT, evaluated by Western blot analysis of the epithelial markers E‐cadherin (E‐cad) and zonula occludens‐1 (ZO‐1) or mesenchymal markers vimentin (Vim), N‐cadherin (N‐cad) and Snail, (G) MMP‐2 and MMP‐9 expression, evaluated by Western blotting or activity, evaluated by the gelatin zymography, and (H) migration and invasion capabilities, evaluated by specific assays. The Western blots were obtained by the appropriate Abs. The blots were stripped off the bound Abs and reprobed with anti‐β‐actin or lamin β1 to confirm equal loading. The Western blots shown are representative of three independent experiments. The histograms indicate mean ± SD of three different cultures, and each was tested in triplicate. siCtr: control (non‐specific siRNA). (−) untreated and (+) treated cells. * P

    Techniques Used: Quantitative RT-PCR, Western Blot, Enzyme-linked Immunosorbent Assay, Modification, Expressing, Activation Assay, Inhibition, Activity Assay, Zymography, Migration

    15) Product Images from "Cytosolic Phospholipase A2? and PPAR-? Signaling Pathway Counteracts TGF-?-mediated Inhibition of Primary and Transformed Hepatocyte Growth"

    Article Title: Cytosolic Phospholipase A2? and PPAR-? Signaling Pathway Counteracts TGF-?-mediated Inhibition of Primary and Transformed Hepatocyte Growth

    Journal: Hepatology (Baltimore, Md.)

    doi: 10.1002/hep.23703

    Overexpression of cPLA 2 α in hepatocytes prevents TGF-β1-induced mitoinhibition
    Figure Legend Snippet: Overexpression of cPLA 2 α in hepatocytes prevents TGF-β1-induced mitoinhibition

    Techniques Used: Over Expression

    cPLA 2 α signaling prevents TGF-β1-induced inhibition of cell growth
    Figure Legend Snippet: cPLA 2 α signaling prevents TGF-β1-induced inhibition of cell growth

    Techniques Used: Inhibition

    AA and PPARγ ligands block TGF-β1-induced Smad 3 binding to its DNA response element. Comparison of cPLA 2 α and PPARγ expression in different cell lines
    Figure Legend Snippet: AA and PPARγ ligands block TGF-β1-induced Smad 3 binding to its DNA response element. Comparison of cPLA 2 α and PPARγ expression in different cell lines

    Techniques Used: Blocking Assay, Binding Assay, Expressing

    TGF-β1 activates arachidonic acid signaling cascade through cPLA 2 α phosphorylation in transformed human hepatocytes
    Figure Legend Snippet: TGF-β1 activates arachidonic acid signaling cascade through cPLA 2 α phosphorylation in transformed human hepatocytes

    Techniques Used: Transformation Assay

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    Enzyme-linked Immunosorbent Assay:

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    Recombinant:

    Article Title: Aortic Valve Endothelial Cells Undergo Transforming Growth Factor-?-Mediated and Non-Transforming Growth Factor-?-Mediated Transdifferentiation in Vitro
    Article Snippet: .. Material used were endothelial basal medium (EBM) (CC-3121; Clonetics, San Diego, CA); fetal bovine serum (FBS) (Hyclone, Logan, UT); 100× GPS (29.2 mg/ml l -glutamine, 10,000 U/ml penicillin G, 10,000 μg/ml streptomycin sulfate); gentamicin sulfate and 100× PSF (10,000 U/ml penicillin G, 10,000 μg/ml streptomycin sulfate, 25 μg/ml amphotericin B) (Life Technologies, Inc., Grand Island, NY); collagenase A (Boehringer Mannheim, Indianapolis, IN); Immobilon-P membrane (Millipore, Bedford, MA); Hyperfilm ECL, fluorescein-streptavidin, and Texas Red-streptavidin (Amersham Life Sciences, Arlington Heights, IL); Lumiglo (KPL); human TGF-β1, recombinant human TGF-β2 and -β3, recombinant human platelet-derived growth factor (PDGF)-BB, and anti-PDFG-BB (R & D Systems, Minneapolis, MN); Vectastain Elite ABC kit, avidin/biotin blocking kit, fluorescein anti-mouse IgG, Texas red anti-rabbit IgG, peroxidase-conjugated anti-goat IgG, biotinylated horse anti-mouse IgG, avidin-peroxidase, peroxidase-conjugated anti-mouse IgG, and 3,3′5,5′-tetramethylbenzidine (Vector Laboratories, Burlingame, CA); 3-amino-9-ethyl carbazol and mouse anti-human α-SMA (clone 1A4 ) (Sigma Chemical Co., St. Louis, MO); goat anti-human CD31/PECAM-1 IgG (Santa Cruz Biotechnology, Santa Cruz, CA); rabbit anti-human von Willebrand factor (vWF) and mouse anti-human CD31/PECAM-1 (DAKO, Carpinteria, CA); polycarbonate PVP-F membranes (Neuro Probe, Inc, Gaithersburg, MD). .. Recombinant human bFGF was kindly provided by Scios Nova Inc., Mountain View, CA; soluble recombinant TGF-β type II receptor, prepared as described, was kindly provided by Philip Gotwals, Biogen, Cambridge, MA; SM1 antibody was kindly provided by Masanori Aikawa, Brigham and Women’s Hospital, Boston; rabbit anti-bovine CD31/PECAM-1 was kindly provided by Steven Albelda, University of Pennsylvania.

    Avidin-Biotin Assay:

    Article Title: Aortic Valve Endothelial Cells Undergo Transforming Growth Factor-?-Mediated and Non-Transforming Growth Factor-?-Mediated Transdifferentiation in Vitro
    Article Snippet: .. Material used were endothelial basal medium (EBM) (CC-3121; Clonetics, San Diego, CA); fetal bovine serum (FBS) (Hyclone, Logan, UT); 100× GPS (29.2 mg/ml l -glutamine, 10,000 U/ml penicillin G, 10,000 μg/ml streptomycin sulfate); gentamicin sulfate and 100× PSF (10,000 U/ml penicillin G, 10,000 μg/ml streptomycin sulfate, 25 μg/ml amphotericin B) (Life Technologies, Inc., Grand Island, NY); collagenase A (Boehringer Mannheim, Indianapolis, IN); Immobilon-P membrane (Millipore, Bedford, MA); Hyperfilm ECL, fluorescein-streptavidin, and Texas Red-streptavidin (Amersham Life Sciences, Arlington Heights, IL); Lumiglo (KPL); human TGF-β1, recombinant human TGF-β2 and -β3, recombinant human platelet-derived growth factor (PDGF)-BB, and anti-PDFG-BB (R & D Systems, Minneapolis, MN); Vectastain Elite ABC kit, avidin/biotin blocking kit, fluorescein anti-mouse IgG, Texas red anti-rabbit IgG, peroxidase-conjugated anti-goat IgG, biotinylated horse anti-mouse IgG, avidin-peroxidase, peroxidase-conjugated anti-mouse IgG, and 3,3′5,5′-tetramethylbenzidine (Vector Laboratories, Burlingame, CA); 3-amino-9-ethyl carbazol and mouse anti-human α-SMA (clone 1A4 ) (Sigma Chemical Co., St. Louis, MO); goat anti-human CD31/PECAM-1 IgG (Santa Cruz Biotechnology, Santa Cruz, CA); rabbit anti-human von Willebrand factor (vWF) and mouse anti-human CD31/PECAM-1 (DAKO, Carpinteria, CA); polycarbonate PVP-F membranes (Neuro Probe, Inc, Gaithersburg, MD). .. Recombinant human bFGF was kindly provided by Scios Nova Inc., Mountain View, CA; soluble recombinant TGF-β type II receptor, prepared as described, was kindly provided by Philip Gotwals, Biogen, Cambridge, MA; SM1 antibody was kindly provided by Masanori Aikawa, Brigham and Women’s Hospital, Boston; rabbit anti-bovine CD31/PECAM-1 was kindly provided by Steven Albelda, University of Pennsylvania.

    Blocking Assay:

    Article Title: Aortic Valve Endothelial Cells Undergo Transforming Growth Factor-?-Mediated and Non-Transforming Growth Factor-?-Mediated Transdifferentiation in Vitro
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    other:

    Article Title: Dendritic cell derived exosomes loaded with immunoregulatory cargo reprogram local immune responses and inhibit degenerative bone disease in vivo
    Article Snippet: The presence of TGFB1 and EXO in intraluminal vesicles of MVB for presumptive release was determined by immunogold-TEM in regDC ( ).

    Article Title: Cytosolic Phospholipase A2? and PPAR-? Signaling Pathway Counteracts TGF-?-mediated Inhibition of Primary and Transformed Hepatocyte Growth
    Article Snippet: Human TGF-β1 (transforming growth factor beta 1) was purchased from R & D Systems, Inc. (Minneapolis, MN).

    Isolation:

    Article Title: Dendritic cell derived exosomes loaded with immunoregulatory cargo reprogram local immune responses and inhibit degenerative bone disease in vivo
    Article Snippet: The relative levels of TGFB1 and IL-10 in both lumen and transmembrane domain of regDC EXO was determined as in materials and methods, and as shown in ( and ). .. Neither TGFB1 nor IL10 were detectable in iDC EXO and stimDC EXO, nor in the PBS supernatant of regDC EXO after isolation, ruling out soluble carry over. .. Higher magnification TEM of regDC EXO confirms ELISA data, with TGFB1 inside and attached to outer EXO membrane ( ).

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    R&D Systems tgf β1
    Effects of MAPKs inhibitors on <t>TGF-β1</t> induced increase of proliferation in ASMCs. Confluent and growth-arrested ASMCs were pretreated for 1 hour with SB 203580, PD 98059, or SP 600125, prior to 24-hour treatment with 1 ng/ml of TGF-β1 (T). DNA synthesis was measured by [ 3 H]-thymidine incorporation assay. Inhibition of phosphorylated p38 and ERK1/2 reduced TGF-β1 induced DNA synthesis. ## p
    Tgf β1, supplied by R&D Systems, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    R&D Systems anti transforming growth factor β1
    Overexpression of KLF15 regulates endothelial cell associated factors in Eahy926 cells. (A) mRNA expression levels of eNOS, MCP-1, ICAM-1 and <t>TGF-β1</t> were determined via reverse transcription-quantitative polymerase chain reaction. (B) The protein expression levels of eNOS, MCP-1, ICAM-1 and TGF-β1 were investigated via western blotting. β-actin was used as an internal control. *P
    Anti Transforming Growth Factor β1, supplied by R&D Systems, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    R&D Systems reagents tgf β1
    EGCG reduced the expression of liver fibrogenic genes and the activity of the <t>TGF-β/Smad</t> signaling pathway in a dose-dependent manner in human hepatic stellate LX-2 cells. (A) mRNA expression of COL1A1, MMP-2, TIMP-1, <t>TGF-β1,</t> TGFBR2, and SMAD4 in LX-2 cells. b P
    Reagents Tgf β1, supplied by R&D Systems, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    R&D Systems recombinant human rh tgf β1
    <t>TGF-</t> <t>β1</t> induced dose-dependent changes in endothelial cell receptors. a Representative western blots of endothelial cells exposed to exogenous TGF-β1 (0.1–10.0 ng/ml) for 24 h. Densitometry showed significantly reduced VEGFR2 expression ( b ), but no change in VEGFR1 ( c ) in cells treated with higher doses of TGF-β1. The TGF-β1 co-receptor endoglin was significantly upregulated in a dose dependent fashion ( e ). Notch1 was significantly downregulated in endothelial cells treated with 5.0 and 10.0 ng/ml TGF-β1 ( d ), and its ligand Dll4 showed a trend towards reduced expression with higher doses ( f ). TGF-β1 signaling through ALK5/SMAD2-dependent pathways, as revealed by phosphorylation of SMAD2, was maximal at doses of 5.0 ng/ml and higher ( g ). * p ≤ 0.05; ** p ≤ 0.01; N = 3; Kruskal-Wallis test
    Recombinant Human Rh Tgf β1, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Effects of MAPKs inhibitors on TGF-β1 induced increase of proliferation in ASMCs. Confluent and growth-arrested ASMCs were pretreated for 1 hour with SB 203580, PD 98059, or SP 600125, prior to 24-hour treatment with 1 ng/ml of TGF-β1 (T). DNA synthesis was measured by [ 3 H]-thymidine incorporation assay. Inhibition of phosphorylated p38 and ERK1/2 reduced TGF-β1 induced DNA synthesis. ## p

    Journal: Respiratory Research

    Article Title: TGF-?1 increases proliferation of airway smooth muscle cells by phosphorylation of map kinases

    doi: 10.1186/1465-9921-7-2

    Figure Lengend Snippet: Effects of MAPKs inhibitors on TGF-β1 induced increase of proliferation in ASMCs. Confluent and growth-arrested ASMCs were pretreated for 1 hour with SB 203580, PD 98059, or SP 600125, prior to 24-hour treatment with 1 ng/ml of TGF-β1 (T). DNA synthesis was measured by [ 3 H]-thymidine incorporation assay. Inhibition of phosphorylated p38 and ERK1/2 reduced TGF-β1 induced DNA synthesis. ## p

    Article Snippet: Roles of FGF-2, PDGF, EGF and IGF-I in TGF-β1 induced proliferation To examine if the TGF-β1 induced proliferation of ASMCs is a secondary effect mediated by other growth factors that had been reported to be induced by TGF-β1 [ , , - ], ASMCs were treated with TGF-β1 in the absence or presence of neutralizing antibodies against FGF-2, PDGF, EGF, and IGF-I (all from R & D Systems).

    Techniques: DNA Synthesis, Thymidine Incorporation Assay, Inhibition

    Role of EGF, FGF-2, PDGF and IGF-I in TGF-β1 induced proliferation of ASMCs. Confluent and growth-arrested ASMCs were treated with TGF-β1 (1 ng/ml) in the absence or presence of neutralizing antibodies to EGF, FGF-2, PDGF and IGF-I for 48 hours prior to [ 3 H]-thymidine incorporation assay. # p

    Journal: Respiratory Research

    Article Title: TGF-?1 increases proliferation of airway smooth muscle cells by phosphorylation of map kinases

    doi: 10.1186/1465-9921-7-2

    Figure Lengend Snippet: Role of EGF, FGF-2, PDGF and IGF-I in TGF-β1 induced proliferation of ASMCs. Confluent and growth-arrested ASMCs were treated with TGF-β1 (1 ng/ml) in the absence or presence of neutralizing antibodies to EGF, FGF-2, PDGF and IGF-I for 48 hours prior to [ 3 H]-thymidine incorporation assay. # p

    Article Snippet: Roles of FGF-2, PDGF, EGF and IGF-I in TGF-β1 induced proliferation To examine if the TGF-β1 induced proliferation of ASMCs is a secondary effect mediated by other growth factors that had been reported to be induced by TGF-β1 [ , , - ], ASMCs were treated with TGF-β1 in the absence or presence of neutralizing antibodies against FGF-2, PDGF, EGF, and IGF-I (all from R & D Systems).

    Techniques: Thymidine Incorporation Assay

    TGF-β1 concentration-dependently increased proliferation of ASMCs. Confluent and growth-arrested ASMCs were incubated with various concentrations of TGF-β1 for 24 or 48 hours prior to [ 3 H]-thymidine incorporation assay (A) or cell counting (B). Significant differences were detected at all concentrations of TGF-β1 treatment compared to the untreated control, p

    Journal: Respiratory Research

    Article Title: TGF-?1 increases proliferation of airway smooth muscle cells by phosphorylation of map kinases

    doi: 10.1186/1465-9921-7-2

    Figure Lengend Snippet: TGF-β1 concentration-dependently increased proliferation of ASMCs. Confluent and growth-arrested ASMCs were incubated with various concentrations of TGF-β1 for 24 or 48 hours prior to [ 3 H]-thymidine incorporation assay (A) or cell counting (B). Significant differences were detected at all concentrations of TGF-β1 treatment compared to the untreated control, p

    Article Snippet: Roles of FGF-2, PDGF, EGF and IGF-I in TGF-β1 induced proliferation To examine if the TGF-β1 induced proliferation of ASMCs is a secondary effect mediated by other growth factors that had been reported to be induced by TGF-β1 [ , , - ], ASMCs were treated with TGF-β1 in the absence or presence of neutralizing antibodies against FGF-2, PDGF, EGF, and IGF-I (all from R & D Systems).

    Techniques: Concentration Assay, Incubation, Thymidine Incorporation Assay, Cell Counting

    TGF-β1 enhanced serum-induced proliferation of ASMCs. Confluent and growth-arrested ASMCs were treated with 10% FBS in the absence or presence of TGF-β1 (1 ng/ml) for 48 hours and the changes of [ 3 H]-thymidine incorporation (n = 9) and cell number (n = 6) were determined. All values are % of untreated control cultured in 0.2% BSA/DMEM. p values indicated were compared to control (10% FBS only).

    Journal: Respiratory Research

    Article Title: TGF-?1 increases proliferation of airway smooth muscle cells by phosphorylation of map kinases

    doi: 10.1186/1465-9921-7-2

    Figure Lengend Snippet: TGF-β1 enhanced serum-induced proliferation of ASMCs. Confluent and growth-arrested ASMCs were treated with 10% FBS in the absence or presence of TGF-β1 (1 ng/ml) for 48 hours and the changes of [ 3 H]-thymidine incorporation (n = 9) and cell number (n = 6) were determined. All values are % of untreated control cultured in 0.2% BSA/DMEM. p values indicated were compared to control (10% FBS only).

    Article Snippet: Roles of FGF-2, PDGF, EGF and IGF-I in TGF-β1 induced proliferation To examine if the TGF-β1 induced proliferation of ASMCs is a secondary effect mediated by other growth factors that had been reported to be induced by TGF-β1 [ , , - ], ASMCs were treated with TGF-β1 in the absence or presence of neutralizing antibodies against FGF-2, PDGF, EGF, and IGF-I (all from R & D Systems).

    Techniques: Cell Culture

    TGF-β1 increased expression of phosphorylated MAPKs in ASMCs. Confluent and growth-arrested ASMCs were incubated with 1 ng/ml of TGF-β1 for 1, 5, 30 minutes, 24 or 48 hours prior to protein extraction and Western analysis for phosphorylated or total p38 (Panel A), ERK1/2 (Panel B), and JNK (Panel C). * p

    Journal: Respiratory Research

    Article Title: TGF-?1 increases proliferation of airway smooth muscle cells by phosphorylation of map kinases

    doi: 10.1186/1465-9921-7-2

    Figure Lengend Snippet: TGF-β1 increased expression of phosphorylated MAPKs in ASMCs. Confluent and growth-arrested ASMCs were incubated with 1 ng/ml of TGF-β1 for 1, 5, 30 minutes, 24 or 48 hours prior to protein extraction and Western analysis for phosphorylated or total p38 (Panel A), ERK1/2 (Panel B), and JNK (Panel C). * p

    Article Snippet: Roles of FGF-2, PDGF, EGF and IGF-I in TGF-β1 induced proliferation To examine if the TGF-β1 induced proliferation of ASMCs is a secondary effect mediated by other growth factors that had been reported to be induced by TGF-β1 [ , , - ], ASMCs were treated with TGF-β1 in the absence or presence of neutralizing antibodies against FGF-2, PDGF, EGF, and IGF-I (all from R & D Systems).

    Techniques: Expressing, Incubation, Protein Extraction, Western Blot

    Effects of MAPKs inhibitors on TGF-β1 induced activation of MAPKs. Confluent and growth-arrested ASMCs were pretreated for 1 hour with SB 203580, PD 98059, or SP 600125, prior to 24-hour treatment with 1 ng/ml of TGF-β1 (T), followed by protein extraction and Western analysis for phosphorylated or total p38 (Panel A), ERK1/2 (Panel B), and JNK (Panel C). The blots are representatives of 3 independent experiments. C = control. ** p

    Journal: Respiratory Research

    Article Title: TGF-?1 increases proliferation of airway smooth muscle cells by phosphorylation of map kinases

    doi: 10.1186/1465-9921-7-2

    Figure Lengend Snippet: Effects of MAPKs inhibitors on TGF-β1 induced activation of MAPKs. Confluent and growth-arrested ASMCs were pretreated for 1 hour with SB 203580, PD 98059, or SP 600125, prior to 24-hour treatment with 1 ng/ml of TGF-β1 (T), followed by protein extraction and Western analysis for phosphorylated or total p38 (Panel A), ERK1/2 (Panel B), and JNK (Panel C). The blots are representatives of 3 independent experiments. C = control. ** p

    Article Snippet: Roles of FGF-2, PDGF, EGF and IGF-I in TGF-β1 induced proliferation To examine if the TGF-β1 induced proliferation of ASMCs is a secondary effect mediated by other growth factors that had been reported to be induced by TGF-β1 [ , , - ], ASMCs were treated with TGF-β1 in the absence or presence of neutralizing antibodies against FGF-2, PDGF, EGF, and IGF-I (all from R & D Systems).

    Techniques: Activation Assay, Protein Extraction, Western Blot

    ASMC responses to TGF-β1 and serum in different culture conditions. ASMCs were cultured with DMEM/10% FBS to confluence and then changed to DMEM/0.2% BSA, DMEM/0.5% FBS, or DMEM/10% FBS for 72 hours, followed by treatment with 5 ng/ml of TGF-β1 or 10% FBS for 24 hours prior to [ 3 H]-thymidine incorporation assay. * p

    Journal: Respiratory Research

    Article Title: TGF-?1 increases proliferation of airway smooth muscle cells by phosphorylation of map kinases

    doi: 10.1186/1465-9921-7-2

    Figure Lengend Snippet: ASMC responses to TGF-β1 and serum in different culture conditions. ASMCs were cultured with DMEM/10% FBS to confluence and then changed to DMEM/0.2% BSA, DMEM/0.5% FBS, or DMEM/10% FBS for 72 hours, followed by treatment with 5 ng/ml of TGF-β1 or 10% FBS for 24 hours prior to [ 3 H]-thymidine incorporation assay. * p

    Article Snippet: Roles of FGF-2, PDGF, EGF and IGF-I in TGF-β1 induced proliferation To examine if the TGF-β1 induced proliferation of ASMCs is a secondary effect mediated by other growth factors that had been reported to be induced by TGF-β1 [ , , - ], ASMCs were treated with TGF-β1 in the absence or presence of neutralizing antibodies against FGF-2, PDGF, EGF, and IGF-I (all from R & D Systems).

    Techniques: Cell Culture, Thymidine Incorporation Assay

    Overexpression of KLF15 regulates endothelial cell associated factors in Eahy926 cells. (A) mRNA expression levels of eNOS, MCP-1, ICAM-1 and TGF-β1 were determined via reverse transcription-quantitative polymerase chain reaction. (B) The protein expression levels of eNOS, MCP-1, ICAM-1 and TGF-β1 were investigated via western blotting. β-actin was used as an internal control. *P

    Journal: Molecular Medicine Reports

    Article Title: Protective effect of KLF15 on vascular endothelial dysfunction induced by TNF-α

    doi: 10.3892/mmr.2018.9195

    Figure Lengend Snippet: Overexpression of KLF15 regulates endothelial cell associated factors in Eahy926 cells. (A) mRNA expression levels of eNOS, MCP-1, ICAM-1 and TGF-β1 were determined via reverse transcription-quantitative polymerase chain reaction. (B) The protein expression levels of eNOS, MCP-1, ICAM-1 and TGF-β1 were investigated via western blotting. β-actin was used as an internal control. *P

    Article Snippet: Subsequently, membranes were blocked with 5% non-fat milk at 37°C for 1 h. The membrane was then incubated with anti-KLF15 (cat. no. ab2647; 1:1,000; Abcam, Cambridge, UK), anti-endothelial nitric oxide synthase (eNOS; cat. no. AF950; 1:700; R & D Systems, Inc., Minneapolis, MN, USA), anti-monocyte chemoattractant protein-1 (MCP-1; cat. no. MAB679, 1:1,000; R & D Systems, Inc.), anti-intercellular adhesion molecule-1 (ICAM-1; cat. no. BBA3; 1:800; R & D Systems, Inc.), anti-transforming growth factor-β1 (TGF-β1; cat. no. MAB1835; 1:1,200; R & D Systems, Inc.), anti-phosphorylated (p)-p65 (cat. no. MAB7226; 1:1,000; R & D Systems, Inc.), anti-Nrf2 (cat. no. MAB3925; 1:1,000; R & D Systems, Inc.) and anti-β-actin (cat. no. MAB8969; 1:2,000; R & D Systems, Inc.) at 4°C overnight.

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

    EGCG reduced the expression of liver fibrogenic genes and the activity of the TGF-β/Smad signaling pathway in a dose-dependent manner in human hepatic stellate LX-2 cells. (A) mRNA expression of COL1A1, MMP-2, TIMP-1, TGF-β1, TGFBR2, and SMAD4 in LX-2 cells. b P

    Journal: Acta Pharmacologica Sinica

    Article Title: The anti-fibrotic effects of epigallocatechin-3-gallate in bile duct-ligated cholestatic rats and human hepatic stellate LX-2 cells are mediated by the PI3K/Akt/Smad pathway

    doi: 10.1038/aps.2014.155

    Figure Lengend Snippet: EGCG reduced the expression of liver fibrogenic genes and the activity of the TGF-β/Smad signaling pathway in a dose-dependent manner in human hepatic stellate LX-2 cells. (A) mRNA expression of COL1A1, MMP-2, TIMP-1, TGF-β1, TGFBR2, and SMAD4 in LX-2 cells. b P

    Article Snippet: Chemicals and reagents TGF-β1 (240-B) was purchased from R & D Systems (Minneapolis, MN).

    Techniques: Expressing, Activity Assay

    Effects of EGCG on the PI3K/Akt/Smad signaling pathway in LX-2 cells and BDL rat livers. (A) LX-2 cells were treated with EGCG, LY294002 or both for 24 h, the mRNA expression of COL1A1 and TGF-β1 were detected by real-time PCR, and (B) the expression of liver fibrogenic protein and members of the PI3K/Akt/Smad signaling pathway were detected by Western blotting. (C) The protein expression of members of the PI3K/Akt and TGF-β/Smad signaling pathways in BDL rat livers, and the quantification of the whole group ( n =10). b P

    Journal: Acta Pharmacologica Sinica

    Article Title: The anti-fibrotic effects of epigallocatechin-3-gallate in bile duct-ligated cholestatic rats and human hepatic stellate LX-2 cells are mediated by the PI3K/Akt/Smad pathway

    doi: 10.1038/aps.2014.155

    Figure Lengend Snippet: Effects of EGCG on the PI3K/Akt/Smad signaling pathway in LX-2 cells and BDL rat livers. (A) LX-2 cells were treated with EGCG, LY294002 or both for 24 h, the mRNA expression of COL1A1 and TGF-β1 were detected by real-time PCR, and (B) the expression of liver fibrogenic protein and members of the PI3K/Akt/Smad signaling pathway were detected by Western blotting. (C) The protein expression of members of the PI3K/Akt and TGF-β/Smad signaling pathways in BDL rat livers, and the quantification of the whole group ( n =10). b P

    Article Snippet: Chemicals and reagents TGF-β1 (240-B) was purchased from R & D Systems (Minneapolis, MN).

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

    EGCG significantly reduced the expression of genes involved in liver fibrosis and inflammation in BDL rat livers. (A) mRNA expression of COL1A1, MMP-2, MMP-9, TNF-α, IL-1β, and TGF-β1 in the sham, BDL, and EGCG-treated BDL livers. The mRNA expression levels were normalized against GAPDH. (B) Representative images of liver tissue sections labeled with an anti-desmin antibody or with an anti-α-SMA antibody. The positive areas were quantified using Image J Software. (C) Western blotting analysis of α-SMA, collagen 1a1 (COL1a1) and TGF-β1 expression in BDL rat livers, and the quantification of the whole group. b P

    Journal: Acta Pharmacologica Sinica

    Article Title: The anti-fibrotic effects of epigallocatechin-3-gallate in bile duct-ligated cholestatic rats and human hepatic stellate LX-2 cells are mediated by the PI3K/Akt/Smad pathway

    doi: 10.1038/aps.2014.155

    Figure Lengend Snippet: EGCG significantly reduced the expression of genes involved in liver fibrosis and inflammation in BDL rat livers. (A) mRNA expression of COL1A1, MMP-2, MMP-9, TNF-α, IL-1β, and TGF-β1 in the sham, BDL, and EGCG-treated BDL livers. The mRNA expression levels were normalized against GAPDH. (B) Representative images of liver tissue sections labeled with an anti-desmin antibody or with an anti-α-SMA antibody. The positive areas were quantified using Image J Software. (C) Western blotting analysis of α-SMA, collagen 1a1 (COL1a1) and TGF-β1 expression in BDL rat livers, and the quantification of the whole group. b P

    Article Snippet: Chemicals and reagents TGF-β1 (240-B) was purchased from R & D Systems (Minneapolis, MN).

    Techniques: Expressing, Labeling, Software, Western Blot

    TGF- β1 induced dose-dependent changes in endothelial cell receptors. a Representative western blots of endothelial cells exposed to exogenous TGF-β1 (0.1–10.0 ng/ml) for 24 h. Densitometry showed significantly reduced VEGFR2 expression ( b ), but no change in VEGFR1 ( c ) in cells treated with higher doses of TGF-β1. The TGF-β1 co-receptor endoglin was significantly upregulated in a dose dependent fashion ( e ). Notch1 was significantly downregulated in endothelial cells treated with 5.0 and 10.0 ng/ml TGF-β1 ( d ), and its ligand Dll4 showed a trend towards reduced expression with higher doses ( f ). TGF-β1 signaling through ALK5/SMAD2-dependent pathways, as revealed by phosphorylation of SMAD2, was maximal at doses of 5.0 ng/ml and higher ( g ). * p ≤ 0.05; ** p ≤ 0.01; N = 3; Kruskal-Wallis test

    Journal: BMC Cell Biology

    Article Title: Release of endothelial cell associated VEGFR2 during TGF-β modulated angiogenesis in vitro

    doi: 10.1186/s12860-017-0127-y

    Figure Lengend Snippet: TGF- β1 induced dose-dependent changes in endothelial cell receptors. a Representative western blots of endothelial cells exposed to exogenous TGF-β1 (0.1–10.0 ng/ml) for 24 h. Densitometry showed significantly reduced VEGFR2 expression ( b ), but no change in VEGFR1 ( c ) in cells treated with higher doses of TGF-β1. The TGF-β1 co-receptor endoglin was significantly upregulated in a dose dependent fashion ( e ). Notch1 was significantly downregulated in endothelial cells treated with 5.0 and 10.0 ng/ml TGF-β1 ( d ), and its ligand Dll4 showed a trend towards reduced expression with higher doses ( f ). TGF-β1 signaling through ALK5/SMAD2-dependent pathways, as revealed by phosphorylation of SMAD2, was maximal at doses of 5.0 ng/ml and higher ( g ). * p ≤ 0.05; ** p ≤ 0.01; N = 3; Kruskal-Wallis test

    Article Snippet: TGF-β1 dose-response Serum starved BAEC were treated with 0 (Control), 0.1, 1.0, 5.0, or 10.0 ng/ml of recombinant human (rh) TGF-β1 (R & D Systems) in serum free DMEM for up to 24 h, ± 5 μM SB-431542 (an inhibitor of TGβR-I isoforms ALK-5, −4 and −7 [ ]).

    Techniques: Western Blot, Expressing

    TGF-β1 induced reduction in receptor expression occurs via an ALK5 related pathway. a ) Representative western blots and densitometry ( b ) from BAEC exposed to control conditions (C; DMSO vehicle), 5 μM SB-431542 (an ALK5 inhibitor) in DMSO (SB), 5 ng/ml exogenous TGF-β1 (T) or 5.0 ng/ml TGF-β1 plus 5 μM SB-431542 (T + SB) for 24 h. TGF-β1 leads reduced VEGFR2 expression and activation, and significantly upregulated endoglin in an ALK5/SMAD2-dependent fashion (as revealed by pSMAD2). There were no significant changes in SMAD1/5-dependent signaling, or in VEGFR1, Notch1 or Dll4 expression. * p ≤ 0.05; N = 3; Kruskal-Wallis test

    Journal: BMC Cell Biology

    Article Title: Release of endothelial cell associated VEGFR2 during TGF-β modulated angiogenesis in vitro

    doi: 10.1186/s12860-017-0127-y

    Figure Lengend Snippet: TGF-β1 induced reduction in receptor expression occurs via an ALK5 related pathway. a ) Representative western blots and densitometry ( b ) from BAEC exposed to control conditions (C; DMSO vehicle), 5 μM SB-431542 (an ALK5 inhibitor) in DMSO (SB), 5 ng/ml exogenous TGF-β1 (T) or 5.0 ng/ml TGF-β1 plus 5 μM SB-431542 (T + SB) for 24 h. TGF-β1 leads reduced VEGFR2 expression and activation, and significantly upregulated endoglin in an ALK5/SMAD2-dependent fashion (as revealed by pSMAD2). There were no significant changes in SMAD1/5-dependent signaling, or in VEGFR1, Notch1 or Dll4 expression. * p ≤ 0.05; N = 3; Kruskal-Wallis test

    Article Snippet: TGF-β1 dose-response Serum starved BAEC were treated with 0 (Control), 0.1, 1.0, 5.0, or 10.0 ng/ml of recombinant human (rh) TGF-β1 (R & D Systems) in serum free DMEM for up to 24 h, ± 5 μM SB-431542 (an inhibitor of TGβR-I isoforms ALK-5, −4 and −7 [ ]).

    Techniques: Expressing, Western Blot, Activation Assay

    TGF-β1 induced reduction in endothelial cord formation occurs via an ALK5 related pathway. a Representative phase contrast images showing endothelial cell cord formation 8 h after plating on Matrigel™ under control conditions (C; DMSO vehicle), 5 μM SB-431542 (an ALK5 inhibitor) in DMSO (SB), 5 ng/ml exogenous TGF-β1 (T) or 5.0 ng/ml TGF-β1 plus 5 μM SB-431542 (T + SB). b WimTube automated quantification of cord formation showed SB inhibitor significantly blocked the ability of 5.0 ng/ml TGF-β1 to significantly inhibit total cord length, cord branching, formation of loops, and generation of tip cells. SB inhibitor, either alone or in combination with 5.0 ng/ml TGF-β1 was not significantly different from DMSO control. ** p ≤ 0.01; *** p ≤ 0.001; N = 4; Kruskal-Wallis test

    Journal: BMC Cell Biology

    Article Title: Release of endothelial cell associated VEGFR2 during TGF-β modulated angiogenesis in vitro

    doi: 10.1186/s12860-017-0127-y

    Figure Lengend Snippet: TGF-β1 induced reduction in endothelial cord formation occurs via an ALK5 related pathway. a Representative phase contrast images showing endothelial cell cord formation 8 h after plating on Matrigel™ under control conditions (C; DMSO vehicle), 5 μM SB-431542 (an ALK5 inhibitor) in DMSO (SB), 5 ng/ml exogenous TGF-β1 (T) or 5.0 ng/ml TGF-β1 plus 5 μM SB-431542 (T + SB). b WimTube automated quantification of cord formation showed SB inhibitor significantly blocked the ability of 5.0 ng/ml TGF-β1 to significantly inhibit total cord length, cord branching, formation of loops, and generation of tip cells. SB inhibitor, either alone or in combination with 5.0 ng/ml TGF-β1 was not significantly different from DMSO control. ** p ≤ 0.01; *** p ≤ 0.001; N = 4; Kruskal-Wallis test

    Article Snippet: TGF-β1 dose-response Serum starved BAEC were treated with 0 (Control), 0.1, 1.0, 5.0, or 10.0 ng/ml of recombinant human (rh) TGF-β1 (R & D Systems) in serum free DMEM for up to 24 h, ± 5 μM SB-431542 (an inhibitor of TGβR-I isoforms ALK-5, −4 and −7 [ ]).

    Techniques:

    TGF-β1 induced dose-dependent reduction in endothelial cord formation. a Representative phase contrast images showing endothelial cell cord formation 8 h after plating on Matrigel™ in the presence of exogenous TGF-β1 (0–10 ng/ml). Note higher magnification control panel (0 ng/ml TGF-β1), showing tip cells ( arrows ) and loop of endothelial cords ( asterisk ). WimTube automated quantification of cord formation showed that TGF-β1 significantly inhibits total cord length ( b ), cord branching ( c ), formation of loops ( d ), and generation of tip cells ( e ). TGF-β1 doses of 1.0 ng/ml or higher significantly inhibited cord formation compared to control (0 ng/ml) or 0.1 ng/ml. ** p ≤ 0.01; **p ≤ 0.001; N = 4; Kruskal-Wallis test. Scale bars = 300 μm

    Journal: BMC Cell Biology

    Article Title: Release of endothelial cell associated VEGFR2 during TGF-β modulated angiogenesis in vitro

    doi: 10.1186/s12860-017-0127-y

    Figure Lengend Snippet: TGF-β1 induced dose-dependent reduction in endothelial cord formation. a Representative phase contrast images showing endothelial cell cord formation 8 h after plating on Matrigel™ in the presence of exogenous TGF-β1 (0–10 ng/ml). Note higher magnification control panel (0 ng/ml TGF-β1), showing tip cells ( arrows ) and loop of endothelial cords ( asterisk ). WimTube automated quantification of cord formation showed that TGF-β1 significantly inhibits total cord length ( b ), cord branching ( c ), formation of loops ( d ), and generation of tip cells ( e ). TGF-β1 doses of 1.0 ng/ml or higher significantly inhibited cord formation compared to control (0 ng/ml) or 0.1 ng/ml. ** p ≤ 0.01; **p ≤ 0.001; N = 4; Kruskal-Wallis test. Scale bars = 300 μm

    Article Snippet: TGF-β1 dose-response Serum starved BAEC were treated with 0 (Control), 0.1, 1.0, 5.0, or 10.0 ng/ml of recombinant human (rh) TGF-β1 (R & D Systems) in serum free DMEM for up to 24 h, ± 5 μM SB-431542 (an inhibitor of TGβR-I isoforms ALK-5, −4 and −7 [ ]).

    Techniques:

    TGF-β1 reduces VEGFR2 expression via multiple mechanisms. a Western blot showing TGF-β1 induced loss of cell associated VEGFR2 protein and concomitant increase in full-length VEGFR2 detected in conditioned medium. b Quantitative PCR analysis showed that there was no change in the relative ratio of full length VEGFR2 mRNA to alternative spliced sVEGFR2 mRNA upon TGF-β1 treatment. c TGF-β1 treatment induced increased expression of ADAM family sheddase enzyme ADAM 10. TGF-β1 treatment had no effect on the expression of ADAM 17 or the membrane-associated metalloproteinase MMP-14. d Ultracentrifugation of endothelial conditioned medium demonstrated that soluble VEGFR2 is associated with the extracellular vesicle/exosome marker HSP90 in TGF-β1 treated cells

    Journal: BMC Cell Biology

    Article Title: Release of endothelial cell associated VEGFR2 during TGF-β modulated angiogenesis in vitro

    doi: 10.1186/s12860-017-0127-y

    Figure Lengend Snippet: TGF-β1 reduces VEGFR2 expression via multiple mechanisms. a Western blot showing TGF-β1 induced loss of cell associated VEGFR2 protein and concomitant increase in full-length VEGFR2 detected in conditioned medium. b Quantitative PCR analysis showed that there was no change in the relative ratio of full length VEGFR2 mRNA to alternative spliced sVEGFR2 mRNA upon TGF-β1 treatment. c TGF-β1 treatment induced increased expression of ADAM family sheddase enzyme ADAM 10. TGF-β1 treatment had no effect on the expression of ADAM 17 or the membrane-associated metalloproteinase MMP-14. d Ultracentrifugation of endothelial conditioned medium demonstrated that soluble VEGFR2 is associated with the extracellular vesicle/exosome marker HSP90 in TGF-β1 treated cells

    Article Snippet: TGF-β1 dose-response Serum starved BAEC were treated with 0 (Control), 0.1, 1.0, 5.0, or 10.0 ng/ml of recombinant human (rh) TGF-β1 (R & D Systems) in serum free DMEM for up to 24 h, ± 5 μM SB-431542 (an inhibitor of TGβR-I isoforms ALK-5, −4 and −7 [ ]).

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