Anti-fibrotic activity of nobiletin and nintedanib: In vitro and in vivo evidence in pulmonary fibrosis models.

Abstract

BACKGROUND: Pulmonary fibrosis, a well-known chronic and progressive lung disease, primarily impacts the interstitial tissues of the lungs, with a lack of effective therapies. Nobiletin is a polymethoxyflavonoid that exhibits characteristics of BH3 mimetics. It is mainly extracted from citrus peels and is known for its diverse pharmacological activities. Given the unsatisfactory clinical trial results of nintedanib, a combined treatment approach may represent a viable strategy for combating pulmonary fibrosis. METHODS: An in vitro pulmonary fibrosis model was established by inducing MRC-5 cells with transforming growth factor-β1 (TGF-β1). The impact of nobiletin combined with nintedanib on the migration of MRC-5 cells was assessed by wound healing and Transwell assays. Immunofluorescence and Western blot analyses were employed to assess the effect of drug combination on fibrosis-related markers, while Co-Immunoprecipitation (Co-IP) experiments were performed to assess the effects on autophagy. The anti-fibrotic effects and potential mechanisms of the combination of nobiletin and nintedanib were further explored using a bleomycin-induced pulmonary fibrosis model in C57BL/6 J mice. RESULTS: In vitro, the combination of nobiletin and nintedanib significantly inhibited MRC-5 cells' migration and extracellular matrix deposition, while simultaneously promoting apoptosis and autophagy. In addition, this combination exerted an anti-pulmonary fibrosis effect by regulating epigenetic mechanisms and the PI3K-AKT-mTOR signaling pathway. In vivo studies further revealed that the combination of nobiletin and nintedanib significantly reduced hydroxyproline levels in mice, attenuated lung inflammation, and helped to limit or prevent collagen accumulation. CONCLUSIONS: Our study demonstrates that the combination of nobiletin and nintedanib exhibits promising anti-fibrotic effects both in vitro and in vivo.