Figure 4.

Effects of small molecule TAM (Tyro3, Axl, and Mertk) receptor antagonists and BIBF1120 on normal and IPF myofibroblast differentiation and activation. Fibroblast cultures derived from normal and IPF lung samples were treated with R428 (2 or 10 μM), LDC (5 μM), BIBF1120 (300 nM), S6 (4 μg/ml), or vehicle in the absence or presence of CpG (10 nM) for 1, 4, 24, or 72 hours. (A and B) Phosphorylated Axl was determined using a PathScan phospho-Axl (panTyr) sandwich ELISA kit, whereas (C and D) α-SMA was quantified via in situ in-cell ELISA and (E and F) collagen 1 via direct ELISA. Relative protein expression was calculated from ratios of phosphorylated Axl, α-SMA, or collagen 1 to β-tubulin, and results are expressed as fold changes compared with vehicle controls. Data are mean ± SEM, n = 3 per group. *P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001; ****P ≤ 0.0001. α-SMA = α-smooth muscle actin; Axl = anexelekto; BIBF = BIBF1120 (nintedanib); IPF = idiopathic pulmonary fibrosis; LDC = LDC1267; Mertk = MER proto-oncogene, tyrosine kinase; pAxl = phosphorylated Axl; Tyro3 = TYRO3 protein tyrosine kinase 3.