The novel inhibitor PRI-724 for Wnt/ß-catenin/CBP signaling ameliorates bleomycin-induced pulmonary fibrosis in mice.

Purpose/Aim of the Study: Wnt/ß-catenin signaling was reported to be activated in pulmonary fibrosis, and was focused on as a target for antifibrotic therapy. However, the mechanism how the inhibition of Wnt/ß-catenin signaling ameliorate pulmonary fibrosis has not been fully elucidated. The purpose of this study is to explore the target cells of Wnt/ß-catenin inhibition in pulmonary fibrosis and to examine the antifibrotic effect of the novel inhibitor PRI-724 specifically disrupting the interaction of ß-catenin and CBP. Materials and Methods: The effect of C-82, an active metabolite of PRI-724, on the expression of TGF-ß1 and α-smooth muscle actin (SMA) was examined on fibroblasts and macrophages. We also examined the effects of PRI-724 in mouse model of bleomycin-induced pulmonary fibrosis. Results: The activation and increased accumulation of ß-catenin in the canonical pathway were detected in lung fibroblasts as well as macrophages stimulated by Wnt3a using Western blotting. Treatment with C-82 reduced CBP protein and increased p300 protein binding to ß-catenin in the nucleus of lung fibroblasts. In addition, C-82 inhibited the expression of SMA in lung fibroblasts treated with TGF-ß, indicating the inhibition of myofibroblast differentiation. In the fibrotic lungs induced by bleomycin, ß-catenin was stained strongly in macrophages, but the staining of ß-catenin in alveolar epithelial cells and fibroblasts was weak. The administration of PRI-724 ameliorated pulmonary fibrosis induced by bleomycin in mice when administered with a late, but not an early, treatment schedule. Analysis of bronchoalveolar fluid (BALF) showed a decreased number of alveolar macrophages. In addition, the level of TGF-ß1 in BALF was decreased in mice treated with PRI-724. C-82 also inhibited the production of TGF-ß1 by alveolar macrophages. Conclusions: These results suggest that the ß-catenin/CBP inhibitor PRI-724 is a potent antifibrotic agent that acts by modulating the activity of macrophages in the lungs.

Role of platelet-derived growth factor/platelet-derived growth factor receptor axis in the trafficking of circulating fibrocytes in pulmonary fibrosis.

Circulating fibrocytes have been reported to migrate into the injured lungs, and contribute to fibrogenesis via CXCL12-CXCR4 axis. In contrast, we report that imatinib mesylate prevented bleomycin (BLM)-induced pulmonary fibrosis in mice by inhibiting platelet-derived growth factor receptor (PDGFR), even when it was administered only in the early phase. The goal of this study was to test the hypothesis that platelet-derived growth factor (PDGF) might directly contribute to the migration of fibrocytes to the injured lungs. PDGFR expression in fibrocytes was examined by flow cytometry and RT-PCR. The migration of fibrocytes was evaluated by using a chemotaxis assay for human fibrocytes isolated from peripheral blood. The numbers of fibrocytes triple-stained for CD45, collagen-1, and CXCR4 were also examined in lung digests of BLM-treated mice. PDGFR mRNA levels in fibrocytes isolated from patients with idiopathic pulmonary fibrosis were investigated by real-time PCR. Fibrocytes expressed both PDGFR-α and -ß, and migrated in response to PDGFs. PDGFR inhibitors (imatinib, PDGFR-blocking antibodies) suppressed fibrocyte migration in vitro, and reduced the number of fibrocytes in the lungs of BLM-treated mice. PDGF-BB was a stronger chemoattractant than the other PDGFs in vitro, and anti-PDGFR-ß-blocking antibody decreased the numbers of fibrocytes in the lungs compared with anti-PDGFR-α antibody in vivo. Marked expression of PDGFR-ß was observed in fibrocytes from patients with idiopathic pulmonary fibrosis compared with healthy subjects. These results suggest that PDGF directly functions as a strong chemoattractant for fibrocytes. In particular, the PDGF-BB-PDGFR-ß biological axis might play a critical role in fibrocyte migration into the fibrotic lungs.