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Figure 3 | Arthritis Research & Therapy

Figure 3

From: Transcriptional regulation of matrix metalloproteinase-1 and collagen 1A2 explains the anti-fibrotic effect exerted by proteasome inhibition in human dermal fibroblasts

Figure 3

Bortezomib does not abrogate TGF-β-induced phosphorylation, nuclear translocation, or binding of Smad2 to the COL1A2 promoter. Fibroblasts were treated with TGF-β (5 ng/mL) or bortezomib (1 μM) or both (TGF-β was added 1 hour after bortezomib) or were left untreated for the indicated amount of time. (a) Total protein extracts were analyzed by Western blotting. Band intensities are provided below. (b) Fibroblasts were labeled with rabbit anti-p300 or Smad2/3 antibodies. Immunofluorescence photographs (× 40) from one representative experiment out of three independent experiments are presented. (c) Nuclear proteins from fibroblasts were extracted and used to perform DNA pull-down assays. DNA-bound proteins were eluted and analyzed by Western blotting using anti-P-Smad2 antibodies. Total nuclear protein content was assessed using anti-TFIIEα antibodies on unbound fractions. Band intensities were quantified and normalized to those obtained with the anti-TFIIEα antibody. The increase in P-Smad2 levels in treated relative to untreated cells is provided. Bor, bortezomib; B/T, bortezomib/transforming growth factor-beta; COL1A2, collagen 1A2; TGF-β, transforming growth factor-beta; UT, untreated.

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Arthritis Research & Therapy

ISSN: 1478-6362

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