Integrating molecular dynamics and experimental validation reveals ShenQi granule stabilizes podocyte cytoskeleton by modulating Bax/Bcl2-caspase-3 rheostat in IMN.

ETHNOPHARMACOLOGICAL RELEVANCE: Idiopathic membranous nephropathy (IMN) is one of the most common pathological subtypes of nephrotic syndrome. ShenQi Granule (SQ) a traditional Chinese medicine (TCM) prescription, has been utilized in clinical settings to treat IMN, but the molecular mechanisms underlying its podocyte-protective effects remain poorly defined.

AIM OF THE STUDY: This study aimed to move beyond descriptive efficacy validation to elucidate whether and how SQ orchestrates podocyte protection through integrated regulation of the PI3K/AKT/mTOR-apoptosis axis.

METHODS

A Cationic bovine serum albumin(C-BSA) induced rat model of IMN was used to assess renal function, histopathology, and podocyte ultrastructure. Computationally, network pharmacology identified SQ-IMN common targets, followed by molecular docking and molecular dynamics simulations to evaluate binding stability between SQ's active components and core targets (Bax, Bcl2). Mechanistically, we examined in vivo and in vitro the effects of SQ on PI3K/AKT/mTOR phosphorylation, apoptosis-related proteins (Bax, Bcl2, cleaved-caspase-3), and cytoskeletal integrity (F-actin, Nephrin).

RESULTS

SQ treatment significantly ameliorated proteinuria, renal dysfunction, and podocyte ultrastructural damage in IMN rats, accompanied by reduced glomerular IgG deposition and restored nephrin expression. Network pharmacology coupled with molecular docking revealed stable binding of SQ's active components to Bax and Bcl2, molecular dynamics revealed highly stable binding between quercetin and Bcl2 over 100 ns, with a binding free energy of -10.9 kcal/mol and consistently low RMSD values, moving beyond conventional docking to demonstrate dynamic binding stability. In vivo, SQ suppressed PI3K/AKT/mTOR phosphorylation, shifted the Bax/Bcl2 balance toward anti-apoptosis, and attenuated cleaved-caspase-3 expression. Puromycin aminonucleoside(PAN)-challenged podocyte experiments confirmed SQ's dual mechanism: simultaneous suppression of AKT/mTOR phosphorylation and apoptosis, coupled with F-actin stabilization-functionally coupling the predicted Bcl2 interaction to podocyte structural preservation.

CONCLUSION

SQ protects against IMN podocyte injury not via generalized anti-apoptosis, but through targeted PI3K/AKT/mTOR orchestration, where cytoskeletal stability intersects with mitochondrial death control. This work establishes a hybrid computational-experimental framework for decoding multi-target herbal mechanisms.