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Objectives Anlotinib hydrochloride (AL3818) is a novel multitarget tyrosine kinase inhibitor which has the same targets as nintedanib, an effective drug has been approved for the treatment of idiopathic pulmonary fibrosis. Here, we examined whether anlotinib could also attenuate bleomycin-induced pulmonary fibrosis in mice and explored the antifibrosis mechanism. Methods We have evaluated the effect of anlotinib on bleomycin-induced pulmonary fibrosis in mice. Inflammatory cytokines in alveolar lavage fluid including IL-1β, IL-4, IL-6 and TNF-α were determined by ELISA. Biomarkers of oxidative stress were measured by corresponding kit. Histopathologic examination was analysed by H&E staining and immunohistochemistry. In vitro, we investigated whether anlotinib inhibited TGFβ/Smad3 and non-Smad pathways by luciferase assay or Western blotting. We also evaluated whether anlotinib inhibited TGF-β1-induced epithelial–mesenchymal transition (EMT) and promoted myofibroblast apoptosis in order to explore the possible molecular mechanism. Key findings The results indicated that anlotinib treatment remarkably attenuated inflammation, oxidative stress and pulmonary fibrosis in mouse lungs. Anlotinib could inhibit the TGF-β1 signalling pathway. Additionally, anlotinib not only profoundly inhibited TGF-β1-induced EMT in alveolar epithelial cells, but also simultaneously reduced the proliferation and promoted the apoptosis in fibroblasts. Conclusions In summary, the results suggest that anlotinib-mediated suppression of pulmonary fibrosis is related to the inhibition of TGF-β1 signalling pathway.

Background: Pirfenidone(PFD) and nintedanib(NDN), the only two drugs for IPF, which should be accurately selected in treatment, has no relevant basic research for reference. Objective: Compare the effect of PFD and NDN in bleomycin(BLM)-induced pulmonary fibrosis model of different periods, so as to provide theoretical basis for clinical application. Methods: The BLM model of different periods were performed in Fig1. Results: 1) Anti-inflammatory and antioxidant capacity: The inhibition rate of NDN on inflammatory cell numbers was higher than that of PFD, while PFD has better inhibitory effect on IL-1β and IL-4 and NDN has higher inhibitory rate on IL-6 and IFN-γ. The activity of SOD in PFD group was higher than that of NDN group, while the content of MDA in lung tissues and the concentration of MPO in serum were significantly lower in NDN group than of PFD group. 2) Anti-fibrosis capacity: In prevention model, hydroxyproline content in PFD group was 140.29±3.59μg/right lung and NDN group was 162.87±2.91μg/right lung. In treatment models, HYP content was much less in NDN group than in PFD group. Pathological change was consistent with the HYP results. Conclusion: Both PFD and NDN have anti-inflammatory and antioxidative effects and selectively suppressed different biomarkers. PFD is more effective in the prophylactic model of fibrosis, while NDN has a better effect in early and late treatment models.

Background: Idiopathic pulmonary fibrosis is incurable and has no effective treatment. The marketed drugs are only pirfenidone and nintedanib. By evaluating the inhibition effects of pirfenidone and nintedanib on the proliferation, migration and activation of human lung fibroblasts (HFL1), the anti-fibrotic mechanism of two drugs was tested and compared to provide reference for accurate clinical medication. Methods: HFL1 cells were divided into control group, TGF-β1/ PDGF-stimulate-model group, and drug-add group. The effect of pirfenidone and nintedanib on the proliferation and migration of HFL1 were evaluated by MTT assay and scratch test, respectively. Anti-pulmonary fibrosis effects of pirfenidone and nintedanib were evaluated by detecting the inhibitory activity of each drug on the expression of fibroblasts activation markers. The inhibitory effects of pirfenidone and nintedanib on TGF-β/Smad3 signaling pathway were evaluated by Western Blotting. Results: The results of MTT assays showed that both pirfenidone and nintedanib could inhibit TGF-β-induced proliferation of fibroblasts. In addition, both pirfenidone and nintedanib inhibited the migration of HFL1; Both pirfenidone and nintedanib can inhibit the expression of fibroblast activation markers, and they can inhibit the activation of TGF-β/Smad3 protein signaling pathway by inhibiting the phosphorylation of Smad3. The inhibition rate of Smad3 phosphorylation by nintedanib was 51%, and the inhibition rate of pirfenidone on Smad3 phosphorylation was 13%. Conclusion: Both pirfenidone and nintedanib could inhibit the proliferation, migration and activation of myofibroblasts, And all of them can exert anti-pulmonary fibrosis through TGF-β signaling pathway.