Sanqi qushi formula ameliorates podocyte injury in membranous nephropathy by inhibiting endoplasmic reticulum stress-induced ferroptosis

ETHNOPHARMACOLOGICAL RELEVANCE: Membranous nephropathy (MN) is a primary glomerular disease and a major cause of nephrotic syndrome in adults. According to traditional Chinese medicine (TCM) theory, "dampness" is considered a key pathogenic factor. Sanqi Qushi Formula (SQQS), developed by adding dampness-eliminating herbs to Sanqi Oral Liquid, has been shown to improve remission rates in idiopathic MN, although its pharmacological mechanisms are not yet fully clarified. AIM OF THE STUDY: This study investigated whether SQQS attenuates podocyte injury in MN by inhibiting endoplasmic reticulum stress (ERS)-induced ferroptosis. METHODS: The main components of SQQS were identified and quantified by widely targeted metabolomics. A passive Heymann nephritis (PHN) rat model was established via intravenous injection of anti-Fx1A serum, followed by oral SQQS intervention. Renal function and histopathology were evaluated, and underlying mechanisms were investigated by integrating transcriptomic and proteomic analyses. Structural changes in the endoplasmic reticulum and mitochondria were assessed by transmission electron microscopy. The regulatory effect of SQQS on ERS-induced ferroptosis was further examined using immunohistochemistry, flow cytometry, oxidative stress assays, and related molecular analyses. In vitro, immortalized mouse podocytes MPC-5 were injured with zymosan-activated serum and treated with SQQS-containing serum, the ferroptosis inhibitor Fer-1, the ERS inhibitor 4-PBA, or the ERS activator tunicamycin. The protective effects of SQQS were validated by Western blotting and immunofluorescence. RESULTS: SQQS treatment significantly improved renal function, reduced proteinuria, and alleviated glomerular injury in PHN rats. Histopathological and ultrastructural analyses revealed that SQQS reduced glomerular IgG deposition, basement membrane thickening, fibrosis, and podocyte injury. SQQS dose-dependently restored the expression of key podocyte structural proteins (Nephrin, Synaptopodin, Podocin, α-actinin-4, WT-1) and decreased Desmin levels. Integrated transcriptomic and proteomic analyses identified ERS and ferroptosis as central pathways regulated by SQQS. SQQS suppressed ERS markers (GRP78, p-PERK, p-eIF2α, ATF4, CHOP) and pro-ferroptotic proteins (ACSL4, CHAC1), upregulated antioxidant proteins (GPX4, xCT), reduced iron deposition, and attenuated oxidative stress. In vitro, SQQS-containing serum protected MPC-5 podocytes from ZAS-induced ferroptosis by inhibiting ERS, restoring podocyte marker expression, and enhancing antioxidant defenses. CONCLUSION: SQQS ameliorates MN by inhibiting the ERS-ferroptosis axis, providing experimental and theoretical support for the application of TCM in the treatment of MN.