Figure 2

Effect of loss of TRIP-1 on α-SMA expression is independent of TGFβ receptor signaling and Smad3. A: HLF-F transfected with control siRNA or TRIP-1 siRNA were treated with either vehicle or with 10 μM solution of TGFβ receptor inhibitor (SB431542) for 24 h, after which whole cell lysates were prepared and samples run on SDS-PAGE and analyzed by western-blot for α-SMA protein; Western blot shows a reduction in TGFβ- induced α-SMA in controls treated with SB431542, while the α-SMA was not significantly different in the TRIP-1 siRNA treated fibroblasts when treated with SB431542. TGFβ1 treatment was used as a positive control for the inhibitor; α-Tubulin was used as a loading control; B: normalized values of α-SMA expression in control siRNA (left) or TRIP-1 siRNA (right) transfected cells, with means ± SE shown (**P < 0.05, n = 3) showing a significant difference in TGFβ-induced α-SMA in the controls when treated with SB431542 but no difference in the TRIP-1 siRNA fibroblast treated with SB431542; C: HLF-F cells were transfected with control siRNA, TRIP-1 siRNA or TRIP-1 and Smad3 siRNA and 48 hours later lysates were prepared and analyzed by western-blot for α-SMA expression showing increased α-SMA in TRIP-1 siRNA is independent of Smad3 activity; D: normalized values of α-SMA expression in control siRNA (left column), TRIP-1 siRNA (middle column) or TRIP-1 and Smad3 siRNA (right column) transfected cells, with means ± SE shown (**P < 0.05, n = 3). Experiments were performed three times and one representative experiment is shown.