rabbit polyclonal antibody against total fn  (Abcam)

Journal: Journal of cellular biochemistry

Article Title: The Novel mTOR Complex 1/2 Inhibitor P529 Inhibits Human Lung Myofibroblast Differentiation

doi: 10.1002/jcb.25878

Figure Lengend Snippet: A) HLF were treated with 1 ng/ml TGF-β for 24 h and the indicated concentrations of P529. Lysates were subjected to western blotting for α-SMA, collagen-1, fibronectin, and GAPDH. N=3 biologic repeats. B) Densitometry of Col1/GAPDH and α-SMA/GAPDH. Measurements are normalized to the TGF-β treatment condition. Statistical significance was tested using student’s t-test. *, # denote statistically significant differences (p<0.05) between expression of α-SMA (*) or Col1 (#) in the TGF-β and TGF-β/P529 treatment conditions. C) Real time PCR of the indicated genes. Statistical significance was tested using student’s t-test. *, # denote statistically significant differences (p<0.05) between expression of α-SMA (*) or Col1 (#) in the TGF-β and TGF-β/P529 treatment conditions. N, denotes not statistically significant (FN and PAI-1).

Article Snippet: Rabbit polyclonal antibody against total fibronectin (FN) was from Abcam (ab2413, Cambridge, MA).

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

Journal: Journal of cellular biochemistry

Article Title: The Novel mTOR Complex 1/2 Inhibitor P529 Inhibits Human Lung Myofibroblast Differentiation

doi: 10.1002/jcb.25878

Figure Lengend Snippet: HLF were treated with 1 ng/ml TGF-β for 24 h and P529 or vehicle control, as indicated. A) Cells were then subjected to staining with phalloidin (red) and DAPI (Blue) and visualized with a 60x objective. Scale bar = 50 μm. B) Cells were stained with anti-fibronectin antibody (green) and DAPI (Blue) and visualized with a 10x objective (top row) or a 60x objective (bottom row). Scale bar = 250 μm. C, D) HLF were transfected with siRNA targeting mTOR or scrambled control, followed by treatment with 1 ng/ml TGF-β for 24 h, followed by western blotting and densitometry of the indicated proteins. Statistical significance was tested using student’s t-test. * denote statistically significant differences (p<0.05) between conditions. E) HLF were plated and grown overnight on tissue culture plastic. Serum-free media with DMSO or 10 μM P529 was then added to induce growth arrest. Linear wounds were made at the time of serum removal and imaged. Cells were fixed after 24 h and imaged with a 10x objective. Images shown have been processed with ImageJ to enhance edges and contrast for clarity. F) Closure after 24 h was measured using Image J. Scale Bar = 200 μM. Statistical significance was tested using student’s t-test. * denote statistically significant differences (p<0.05) between conditions.

Article Snippet: Rabbit polyclonal antibody against total fibronectin (FN) was from Abcam (ab2413, Cambridge, MA).

Techniques: Control, Staining, Transfection, Western Blot