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Fisetin ameliorates fibrotic kidney disease in mice via inhibiting ACSL4-mediated tubular ferroptosis

Acta Pharmacologica Sinica volume 45, pages 150–165 (2024)Cite this article

Abstract

Kidney fibrosis is the hallmark of chronic kidney disease (CKD) progression, whereas no effective anti-fibrotic therapies exist. Recent evidence has shown that tubular ferroptosis contributes to the pathogenesis of CKD with persistent proinflammatory and profibrotic responses. We previously reported that natural flavonol fisetin alleviated septic acute kidney injury and protected against hyperuricemic nephropathy in mice. In this study, we investigated the therapeutic effects of fisetin against fibrotic kidney disease and the underlying mechanisms. We established adenine diet-induced and unilateral ureteral obstruction (UUO)-induced CKD models in adult male mice. The two types of mice were administered fisetin (50 or 100 mg·kg−1·d−1, i.g.) for 3 weeks or 7 days, respectively. At the end of the experiments, the mice were euthanized, and blood and kidneys were gathered for analyzes. We showed that fisetin administration significantly ameliorated tubular injury, inflammation, and tubulointerstitial fibrosis in the two types of CKD mice. In mouse renal tubular epithelial (TCMK-1) cells, treatment with fisetin (20 μM) significantly suppressed adenine- or TGF-β1-induced inflammatory responses and fibrogenesis, and improved cell viability. By quantitative real-time PCR analysis of ferroptosis-related genes, we demonstrated that fisetin treatment inhibited ferroptosis in the kidneys of CKD mice as well as in injured TCMK-1 cells, as evidenced by decreased ACSL4, COX2, and HMGB1, and increased GPX4. Fisetin treatment effectively restored ultrastructural abnormalities of mitochondrial morphology and restored the elevated iron, the reduced GSH and GSH/GSSG as well as the increased lipid peroxide MDA in the kidneys of CKD mice. Notably, abnormally high expression of the ferroptosis key marker ACSL4 was verified in the renal tubules of CKD patients (IgAN, MN, FSGS, LN, and DN) as well as adenine- or UUO-induced CKD mice, and in injured TCMK-1 cells. In adenine- and TGF-β1-treated TCMK-1 cells, ACSL4 knockdown inhibited tubular ferroptosis, while ACSL4 overexpression blocked the anti-ferroptotic effect of fisetin and reversed the cytoprotective, anti-inflammatory, and anti-fibrotic effects of fisetin. In summary, we reveal a novel aspect of the nephroprotective effect of fisetin, i.e. inhibiting ACSL4-mediated tubular ferroptosis against fibrotic kidney diseases.

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Fig. 1: Fisetin improved renal dysfunction and kidney injury in adenine diet-induced CKD mice.
Fig. 2: Fisetin improved kidney injury in UUO-induced mice.
Fig. 3: Fisetin improved kidney injury in UUO-induced mice.
Fig. 4: The expression of ferroptosis-related genes was significantly changed in the kidneys of both CKD mice and patients.
Fig. 5: Fisetin inhibited kidney ferroptosis in adenine diet-treated mice.
Fig. 6: Fisetin inhibited kidney ferroptosis in UUO-treated mice.
Fig. 7: Fisetin suppressed inflammation and fibrogenesis in adenine- or TGF-β1-stimulated tubular TCMK-1 cells.
Fig. 8: Fisetin alleviated ferroptosis in adenine- or TGF-β1-stimulated TCMK-1 cells.
Fig. 9: Inhibition of ACSL4-mediated ferroptosis reduced inflammation and fibrogenesis in adenine- or TGF-β1-stimulated TCMK-1 cells.
Fig. 10: Fisetin attenuated ferroptosis, inflammation, and fibrogenesis in adenine- or TGF-β1-stimulated TCMK-1 cells by inhibiting ACSL4 transcription activity.

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Acknowledgements

This current work was supported by the Science/Technology Project of Sichuan Province (2021YFQ0027 and 2022YFS0589) and the Health Commission of Sichuan Province (20PJ048). The graphical abstract was created with a free version of Biorender.com under agreement number EL24NF7RFD.

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  1. These authors contributed equally: Bo Wang, Li-na Yang

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  1. Kidney Research Institute, Division of Nephrology, West China Hospital of Sichuan University, Chengdu, 610041, China

    Bo Wang, Li-na Yang, Le-tian Yang, Fan Guo, Ping Fu & Liang Ma

  2. Research Core Facility of West China Hospital, Sichuan University, Chengdu, 610041, China

    Yan Liang

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Contributions

LM and BW designed experiments. BW, LNY, and YL performed experiments. BW, LNY, LTY, FG, and LM analyzed the data. BW, LNY, and LM wrote the draft of the manuscript and edited it. All authors have read and approved the submitted manuscript.

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Correspondence to Liang Ma.

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Wang, B., Yang, Ln., Yang, Lt. et al. Fisetin ameliorates fibrotic kidney disease in mice via inhibiting ACSL4-mediated tubular ferroptosis. Acta Pharmacol Sin 45, 150–165 (2024). https://doi.org/10.1038/s41401-023-01156-w

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