Figure 3

Effects of SB-431542 and bone morphogenetic protein 6 on Dupuytren's and control fibroblasts. (A) Quantitative PCR was used to determine the average expression of TGF-β1, TGF-β2 and TGF-β3 mRNA from control (mixture 1 through 4) and Dupuytren's (mixture 1 through 4) fibroblasts relative to GAPDH mRNA expression in the presence or absence of 100 ng/mL bone morphogenetic protein 6 (BMP6) for 18 hours. All values are expressed relative to the average of the control (1 through 4) TGF-β1 mRNA values. (B) Immunofluorescence of α-SMA in control and Dupuytren's fibroblasts treated for 72 hours with 20 μmol SB-431542 (+) in combination with or without 100 ng/mL rec. BMP6. Dimethyl sulfoxide (DMSO)-treated cells (-) were used as an internal control. Alexa Fluor 488 (green), α-SMA; 4',6-diamidino-2-phenylindole (blue), DNA. Representative staining is shown for Dupuytren's patient 4 and control patient 1. (C) Quantification of α-SMA-expressing control (mixture 1 through 4) and Dupuytren (mixture 1 through 4) cells after treatment with SB-431542 (20 μmol) in the presence or absence of rec. BMP6 (100 ng/mL) for 72 hours as depicted in Figure 3B. (D) Fibroblast-populated collagen lattice (FPCL) of control (1 through 4) and Dupuytren's (1 through 4) fibroblasts treated with DMSO (-) or 20 μmol SB-431542 (+) in the presence or absence of 100 ng/mL rec. BMP6. Quantification of the average contractions of control (1 through 4) and Dupuytren's (1 through 4) fibroblasts was calculated after 72 hours of treatment. Representative images are shown in Additional file 2, Supplementary Figure 2.