FFA4 inhibits bleomycin-induced pulmonary fibrosis in mice by suppressing IL-33

Background: Interstitial lung disease is one of the most severe pulmonary complications of connective tissue diseases and is associated with high mortality and poor prognosis due to the lack of effective therapies. Free Fatty Acid Receptor 4, a receptor activated by long-chain unsaturated fatty acids, participates in the regulation of inflammatory responses; however, its direct role and underlying mechanisms in ILD-related pulmonary fibrosis have not yet been reported.

Methods: In this study, we investigated the effect of FFA4 on pulmonary fibrosis using both in vivo and in vitro models. A bleomycin-induced pulmonary fibrosis model was established in wild-type and FFAR4 knockout mice. In vitro, a Transwell co-culture system consisting of RAW264.7 macrophages and NIH3T3 fibroblasts was constructed. Transfection, transcriptomic analysis, dual-luciferase reporter assays, RT-qPCR, and Western blotting were performed to explore the molecular mechanisms mediated by FFA4. In addition, pharmacological interventions with the FFA4 agonist CpdA and the NF-κB Inhibitor BAY11-7082 were used to evaluate the role of the NF-κB-IL-33 signaling axis in inflammation-driven fibrotic responses.

Results: FFA4 expression was reduced in the bleomycin-induced fibrosis model. In both the co-culture system and the mouse model, FFA4 deficiency significantly increased IL-33 expression and aggravated pulmonary fibrosis. In contrast, activation of FFA4 with CpdA reduced IL-33 expression and attenuated pulmonary fibrosis. Moreover, BAY11-7082 also suppressed IL-33 expression. Genetic deletion and pharmacological activation experiments further confirmed that the inhibitory effect of CpdA on IL-33 expression was dependent on FFA4.

Conclusions: Collectively, these findings demonstrate that FFA4 restrains inflammatory signal amplification and suppresses pulmonary fibrosis progression by regulating the NF-κB-IL-33 signaling axis, suggesting that FFA4 may serve as a potential therapeutic target for pulmonary fibrosis.