Formononetin in Jiawei Qihuangyin inhibits podocyte epithelial–mesenchymal transition and ameliorates diabetic nephropathy via SIRT1/NF-κB axis

BACKGROUND: Traditional Chinese medicine (TCM) has shown great promise in treating diabetic nephropathy (DN). However, the key targets and mechanisms underlying the therapeutic effects of the active ingredients of modified prescription Jiawei Qihuangyin (JWQHY) remain unclear. METHODS: Network pharmacology analysis was employed to identify potential targets of JWQHY in DN. Protein-protein interaction (PPI) and TCM component-target networks were constructed, and KEGG pathway enrichment analysis was performed to determine key therapeutic targets and signaling pathways. Molecular docking suggested an interaction between the major active compound formononetin (FMN) and the central target silent information regulator 1 (SIRT1), which was experimentally validated using cellular thermal shift assay. SIRT1 expression in podocytes was assessed by qRT-PCR and western blotting (WB). Cell viability (CCK-8), apoptosis (flow cytometry), and proinflammatory cytokine secretion (ELISA) were measured to evaluate podocyte injury. The acetylation level of NF-κB p65 and epithelial-mesenchymal transition (EMT)-related proteins were analyzed by WB. In vivo, a DN rat model was established to assess the therapeutic efficacy of JWQHY through biochemical urine analysis, histopathological examination (HE staining), and WB detection of SIRT1, acetylated NF-κB p65, and EMT markers. RESULTS: Network pharmacology identified 52 potential overlapping targets of JWQHY in DN, primarily associated with the NF-κB pathway. Among these, SIRT1 was predicted and experimentally confirmed as the main target of FMN. In a high-glucose-induced podocyte injury model, FMN upregulated SIRT1 expression, promoted NF-κB p65 deacetylation, and inhibited podocyte EMT. Consistently, FMN treatment improved renal function, reduced podocyte injury, and modulated SIRT1/NF-κB signaling in DN rats. CONCLUSION: JWQHY exerts therapeutic effects in diabetic nephropathy by modulating the SIRT1/NF-κB signaling axis through its active compound formononetin, thereby inhibiting podocyte EMT. These findings provide mechanistic insight into the pharmacological basis of FMN and support its clinical potential in DN treatment.