Bufalin Alleviates Renal Tubulointerstitial Fibrosis in Diabetic Kidney Disease by Inhibiting STAT3-Mediated Ferroptosis

Background: The pathological hallmark of diabetic kidney disease (DKD) is tubulointerstitial fibrosis (TIF), which arises from epithelial-mesenchymal transition (EMT) and extracellular matrix (ECM) synthesis in renal tubular epithelial cells (RTECs). Bufalin exhibits potential for delaying the progression of kidney disease; however, its precise role and underlying mechanisms remain to be elucidated. Ferroptosis is likely involved in the regulatory progression of bufalin. Methods: db/db mice and high glucose (HG) -induced RTECs were used as DKD models. Ferrostatin-1 (Fer-1) was further used to inhibit ferroptosis to assess its effect on TIF. Regarding the mechanism, molecular docking was employed to simulate the binding interaction between bufalin and signal transducer and activator of transcription 3 (STAT3). Additionally, the influence of bufalin on the stability of STAT3 mRNA was assessed in the presence of streptomyces D intervention. Results: Our findings revealed that the treatment of bufalin altered the expression of TIF-related indicators. During the progression of TIF, ferroptosis was activated, characterized by iron overload, elevated lipid peroxidation product generation, and mitochondrial alterations induced by HG. Conversely, Fer-1 reversed these changes and further mitigated TIF. Notably, bufalin exhibited an inhibitory effect on ferroptosis, suggesting a mechanism analogous to Fer-1. Network pharmacology analysis identified STAT3 as a target of bufalin, and knockdown of STAT3 altered the expression of ferroptosis and TIF-related markers. Molecular docking and STAT3 mRNA stability analysis further elucidated the mechanism by which bufalin regulates ferroptosis and TIF. Conclusion: Bufalin inhibited ferroptosis by decreasing the stability of STAT3 mRNA and further alleviates TIF in DKD. Funding: Government Support – Non-U.S.