Transforming growth factor β regulates β-catenin expression in lung fibroblast through NF-κB dependent pathway

Abstract

β-catenin contributes to the pathogenesis of lung fibrosis. However, the expression of β-catenin in fibroblasts under fibrotic conditions has not been studied. We investigated the expression of β-catenin in lung fibroblasts from bleomycin (BLM)‑challenged mice and human lung fibroblasts treated with transforming growth factor β (TGF-β) or lysophosphatidic acid (LPA) by western blot analysis. The result showed that the expression of β-catenin was significantly increased in lung fibrotic foci and lung fibroblasts from bleomycin‑challenged mice. TGF-β stimulated β-catenin expression and induced differentiation in human lung fibroblasts in vitro. Pretreatment of the NF-κB activation inhibitor attenuated the TGF-β‑induced expression of β-catenin and differentiation in human lung fibroblasts. Similarly, LPA induced β-catenin expression in human lung fibroblasts, and pre-treatment of the neutralized anti-TGF-β antibody attenuated the LPA‑induced expression of β-catenin and differentiation in human lung fibroblasts. The results suggested that β-catenin expression is upregulated in lung fibroblast during differentiation, and that TGF-β induced β-catenin expression in human lung fibroblasts through the activation of NF-κB.

Figure 1

Staining of β-catenin and collagen in lung tissue from mice with or without BLM challenge. Representative immunohistochemical staining for (a) β-catenin and (b) Masson’s trichrome staining for collagen in murine lung tissue isolated from mice with or without BLM challenge. Arrow shows the marked expression of (a) β-catenin (brown color) and (b) the deposition of collagen (blue color) in fibrotic foci. Scale bar, 200 μm.

Figure 2

Expression of β-catenin, α-SMA and FN in lung fibroblasts from mice with or without BLM challenge. (a) Representative western blot analysis (b–d) quantification of the expression of (b) β-catenin, (c) FN and (d) α-SMA) in lung fibroblasts from mice with or without BLM challenge. Data are expressed as means ± SEM of three independent experiments. ^*P<0.05 vs. cells from control mice. (e) Immunofloresent staining of β-catenin (red) and FN (green) in lung fibroblasts from mice with or without BLM challenge. Images were examined by immunofluorescence microscopy and recorded by using a 60× oil objective.

Figure 3

TGF-β induced the expression of β-catenin, FN and α-SMA in human lung fibroblasts. (a) Representative western blot and (b–d) quantification of the expression of (b) β-catenin, (c) FN and (d) α-SMA in human lung fibroblasts with or without TGF-β challenge. Data are expressed as means ± SEM of three independent experiments. ^*P<0.05 vs. cells without TGF-β challenge.

Figure 4

NF-κB inhibitor attenuates TGF-β-induced expression of β-catenin, FN and α-SMA in human lung fibroblasts. (a) Representative and (b–d) quantification of the expression of (b) β-catenin, (c) FN and (d) α-SMA in human lung fibroblasts with or without TGF-β challenge. Data are expressed as means ± SEM of three independent experiments. ^*P<0.05 vs. cells without TGF-β challenge. ^#P<0.05 vs. cells without NF-κB inhibitor treatment and with TGF-β challenge.

Figure 5

Lysophosphatidic acid (LPA) induced the differentiation and expression of β-catenin in human lung fibroblasts. (a) Representative western blot (b–d) quantification of the expression of (b) β-catenin, (c) FN and (d) α-SMA in LPA-challenged WI-38 cells. Data are expressed as means ± SEM of three independent experiments. ^*P<0.05 vs. cells with control antibody but without LPA treatment. ^#P<0.05 vs. LPA-challenged cells with pretreatment of control antibody.