Yiqi Huoxue Yangyin Decoction Ameliorates Diabetic Nephropathy by Targeting AMPK/mTOR-Mediated Autophagy: A Network Pharmacology-Guided in vivo and in vitro Study.

PURPOSE: This study aimed to investigate the protective effects and underlying mechanisms of Yiqi Huoxue Yangyin Decoction (YHY), a multi-herb formula composed of Astragalus membranaceus, Ligusticum chuanxiong, Pueraria lobata, Rehmannia glutinosa, and Trichosanthes kirilowii, against diabetic nephropathy (DN), focusing on its role in modulating autophagy and attenuating podocyte senescence via the AMPK/mTOR signaling pathway. MATERIALS AND METHODS: Using network pharmacology and UPLC-Q-TOF-MS, this study examined the active constituents and targets of YHY. The type 2 diabetes db/db mouse model (leptin receptor-deficient BKS.Cg-Lepr/+ mice, random blood glucose ≥ 16.7 mmol/L) was used to construct a DN model in vivo. Mice received daily oral gavage of YHY or vehicle for 4 weeks (n=10). YHY's protective benefits were assessed by physiological measurements, serum insulin and HbA1c by ELISA, histopathological evaluation of glomerular and tubular regions, glomerular basement membrane thickness by TEM, and protein expression in renal cortex by immunohistochemistry, immunofluorescence, and Western blot. In vitro, using a high-glucose-induced MPC-5 model, CCK-8, wound healing assays, qRT-PCR, and Western blot were performed (n=3-6) to investigate the mechanisms of YHY. RESULTS: YHY demonstrated significant protective effects against DN. The expression levels of autophagy markers (ULK1, P62, and LC3B) in renal tissue were restored following YHY intervention. Through the AMPK/mTOR pathway, YHY regulated the expression of autophagy-related genes (ULK1, Beclin-1, P62, LC3B, ATG5, ATG7) and senescence-related genes (P21, P53), and improved the survival and migratory capabilities of high-glucose-induced MPC-5 cells. Furthermore, YHY enhanced the expression of podocyte structural proteins, including Nephrin, NPHS2, and SYNPO. CONCLUSION: YHY treatment was associated with activation of autophagy and attenuation of podocyte senescence, potentially through modulation of the AMPK/mTOR signaling pathway, a finding that requires validation in future mechanistic studies.