Journal: Clinical and translational medicine

Article Title: FKBP51 increases the tumour-promoter potential of TGF-beta.

doi: 10.1186/2001-1326-3-1

Figure Lengend Snippet: Figure 2 FKBP51 interacts with the general transcriptional co-activator p300 and the TGF-β transcription factor Smad2/3. Left; FKBP51 co immunoprecipitates with p300. Right; p300 co immunoprecipitates with FKBP51. Total cell lysates were prepared by SAN melanoma cells transfected with FKBP51/Flag. Cell lysates were immunoprecipitated with anti-Kat3B/p300 (IP p300) or anti-Flag (IP FKBP51). Immunoprecipitated and total lysates were then subjected to Western blot with anti-FKBP51, anti-p300 or anti-Smad 2/3. Smad 2/3 co immunoprecipitate with either p300 (left) and FKBP51 (right).

Article Snippet: Primary antibodies against FKBP51 (F-13; goat polyclonal; Santa Cruz Biotechnology, CA, USA); Smad 2/3 (H465, rabbit polyclonal; Santa Cruz Biotechnology); SPARC (H-90, rabbit polyclonal; Santa Cruz Biotechnology); N-Cadherin (5D5, mouse monoclonal; Abcam, Cambridge, UK); G3PDH (D16H11, rabbit monoclonal; Cell Signaling, Danvers, USA); were used diluted.

Techniques: Transfection, Immunoprecipitation, Western Blot

Journal: Clinical and translational medicine

Article Title: FKBP51 increases the tumour-promoter potential of TGF-beta.

doi: 10.1186/2001-1326-3-1

Figure Lengend Snippet: Figure 5 Mechanism proposed for FKBP51 enhancement of TGF-β pro-oncogenic signal. Left, FKBP51 facilitates Smad recruitment to coactivators. Right, FKBP51 takes part to the transcriptional complex formed by P300 and Smad 2,3 Increase in FKBP51, as it occurs in melanoma, generates an auto regulatory loop of TGF-β signaling, which in turn promotes tumour progression.

Article Snippet: Primary antibodies against FKBP51 (F-13; goat polyclonal; Santa Cruz Biotechnology, CA, USA); Smad 2/3 (H465, rabbit polyclonal; Santa Cruz Biotechnology); SPARC (H-90, rabbit polyclonal; Santa Cruz Biotechnology); N-Cadherin (5D5, mouse monoclonal; Abcam, Cambridge, UK); G3PDH (D16H11, rabbit monoclonal; Cell Signaling, Danvers, USA); were used diluted.

Techniques:

Journal: PLoS ONE

Article Title: Long-Term Gene Therapy with Thrombospondin 2 Inhibits TGF-β Activation, Inflammation and Angiogenesis in Chronic Allograft Nephropathy

doi: 10.1371/journal.pone.0083846

Figure Lengend Snippet: In gene therapy, treated rats with renal transplants TGF-β were evaluated by immunohistochemistry. Active TGF-β was either detected directly by using an antibody recognizing active TGF-β (A) or indirectly by evaluation of phosphorylation of the TGF-β signaling molecule smad2/3 (B) or expression of the TGF-β downstream target PAI-1 within the glomeruli (C) or the cortex (D). Representative microphotographs of PAI-1 staining in renal grafts treated with control (E) and TSP-2 overexpressing plasmid (F) are shown. Total TGF-β1 (G) and TGF-β2 (H) was similar in both groups. Control (n = 8) vs. TSP-2 treated (n = 8); *p<0,05.

Article Snippet: The TGF-β system was studied using antibodies to TGF-β1 (rabbit anti-human TGF-β1, Santa Cruz Biotechnology Inc. ); TGF-β2 (rabbit anti-human TGF-β2, Santa Cruz ); active TGF-β1 (chicken anti-human active TGF-β1, (R&D systems, Germany , , Phospho-Smad2/3 (rabbit anti-human Smad2 peptide phosphorylated at Ser-433/435, Santa Cruz ) or PAI-1, a rabbit polyclonal to human plasminogen activator inhibitor-1 (Santa Cruz).

Techniques: Immunohistochemistry, Phospho-proteomics, Expressing, Staining, Control, Plasmid Preparation

Journal: PLoS ONE

Article Title: Long-Term Gene Therapy with Thrombospondin 2 Inhibits TGF-β Activation, Inflammation and Angiogenesis in Chronic Allograft Nephropathy

doi: 10.1371/journal.pone.0083846

Figure Lengend Snippet: Representative microphotographs from immunohistological staining of kidney grafts for active TGF-β (A, control plasmid; B, TSP-2 therapy, brown cytosolic staining), P-smad 2/3 (C, control plasmid; D, TSP-2 therapy, brown nuclear staining), PAI-1 (E, control plasmid; F, TSP-2 therapy, brown staining), fibronectin (G, control plasmid; H, TSP-2 therapy, brown staining) and alpha-smooth muscle actin (I, control plasmid; J, TSP-2 therapy, brown staining) are shown.

Article Snippet: The TGF-β system was studied using antibodies to TGF-β1 (rabbit anti-human TGF-β1, Santa Cruz Biotechnology Inc. ); TGF-β2 (rabbit anti-human TGF-β2, Santa Cruz ); active TGF-β1 (chicken anti-human active TGF-β1, (R&D systems, Germany , , Phospho-Smad2/3 (rabbit anti-human Smad2 peptide phosphorylated at Ser-433/435, Santa Cruz ) or PAI-1, a rabbit polyclonal to human plasminogen activator inhibitor-1 (Santa Cruz).

Techniques: Staining, Control, Plasmid Preparation

Journal: Redox biology

Article Title: Mitoquinone ameliorates pressure overload-induced cardiac fibrosis and left ventricular dysfunction in mice.

doi: 10.1016/j.redox.2019.101100

Figure Lengend Snippet: Fig. 5. MitoQ blunts activation of myocardial TGF-β1 and profibrogenic signaling in response to pressure overload. A: mRNA expression levels of Agt, Pdgf, TGF-β1 and Edn1; B: mRNA expression levels of Nox4, Ctgf, Acta2, and collagen (Col1a1, Col1a2 and Col3a1); C: Western blotting of TGF-β1, NOX4, phosphorylated SMAD2 (p-SMAD2), and total SMAD2; and D: Quantification of normalized TGF-β1 (left), NOX4 (middle) and p-SMAD2/total SMAD2 (right) protein expression. *: P < 0.05 vs. Sham. **: P < 0.01 vs. Sham. #: P < 0.05 vs. AAC. ##: P < 0.01 vs. AAC. n = 4–6 each group. Agt: angiotensinogen; Pdgf: platelet-derived growth factor; Edn1: endothelin; Nox4: NADPH oxidase 4; Ctgf: connective tissue growth factor; Acta: actin assembly-inducing protein. AAC: ascending aortic constriction.

Article Snippet: Antibodies against SMAD2 (mAb #8685), p-SMAD2 (mAb # 8828), TGF-β1 (pAb #3711) were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Activation Assay, Expressing, Western Blot, Derivative Assay

Journal: Redox biology

Article Title: Mitoquinone ameliorates pressure overload-induced cardiac fibrosis and left ventricular dysfunction in mice.

doi: 10.1016/j.redox.2019.101100

Figure Lengend Snippet: Fig. 6. MitoQ attenuates NOX4-induced oxidative stress and SMAD2 signaling pathway in TGF-β-treated cardiac fibroblasts (CF). A: Representative confocal images of MitoSox stained CF (left) and quantification of ROS measurement (right); B: mRNA expression of TGF-β1, Nox4, Ctgf, Acta2 and Col1a2 in CF; C: Representative western blotting of NOX4, phosphorylated SMAD2 (p-SMAD2), and total SMAD2; D: Quantification of normalized NOX4 and p-SMAD2/total SMAD2 protein ex- pression; E: mRNA expression of Nrf2 and Nqo1 in CF; and F: Nrf2 protein expression in AAC myocardium *: P < 0.05 vs. Control. #: P < 0.05 vs. TGF-β (A-E) or AAC (F). n = 3 each group for CF (A-E) and n = 6 each group for mouse (F). NOX4: NADPH oxidase 4; Ctgf: connective tissue growth factor; Acta: actin assembly- inducing protein; Col1: collagen type I alpha 2 chain; Nrf2: nuclear factor erythroid 2-related factor 2; Nqo1: NAD(P)H quinone dehydrogenase 1. AAC: ascending aortic constriction.

Article Snippet: Antibodies against SMAD2 (mAb #8685), p-SMAD2 (mAb # 8828), TGF-β1 (pAb #3711) were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Staining, Expressing, Western Blot, Control