NOX4-Derived ROS Promotes Collagen I Deposition in Bronchial Smooth Muscle Cells by Activating Noncanonical p38MAPK/Akt-Mediated TGF-β Signaling

Figure 6

ROS-mediated activation of p38MAPK/Akt signaling was responsible for TGF-β₁-induced α-SMA and Col I expression. (a–e) Cells were pretreated with different signaling pathway inhibitors PD98059 (10 mM), SB203580 (10 mM), and LY294002 (10 mM) before exposure to TGF-β₁ (2 ng/mL). (a–c) The impact of inhibitors on the protein expression of NOX4 (a), α-SMA (b), and collagen I (c) after treatment with 2 ng/mL TGF-β₁ for 36 h, as determined by Western blot. (d, e) The impact of inhibitors on the mRNA levels of NOX4 (d) and α-SMA (e) after incubation with 2 ng/mL TGF-β₁ for 24 h; the mRNA levels were determined by real-time PCR. (f, g) Cells were transfected with scramble control, NOX4, and Smad3 siRNA for 24 h and treated with 2 ng/mL TGF-β₁ for 12 h; the protein levels of p-p38MAPK (f) and p-Akt473 (g) were determined by Western blot. (h, i) Cells were pretreated with NAC (10 mM) before exposure to TGF-β₁ (2 ng/mL) for 12 h; the protein levels of p-p38MAPK (h) and p-Akt473 (i) were detected by Western blot. Data was represented as from three independent experiments and presented as . CON: control. For (a–e) and (h, i): , , and compared to the control; ^# and ^## compared to TGF-β₁. For (f, g): , , and ^# compared to the control siRNA with/without TGF-β₁.

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