ZhiXiaoSanZheng Decoction Ameliorates HGPA-induced HK-2 Cell Injury and Mitochondrial Metabolic Imbalance by Regulating the AMPK/PPARα/CPT1A Pathway.

INTRODUCTION ZhiXiaoSanZheng Decoction (ZXSZD) is a modern clinical Traditional Chinese Medicine (TCM) formula that exerts favorable clinical efficacy in the treatment of Diabetic Kidney Disease (DKD). However, its potential renal tubular protective mechanism remains unclear.

Methods

Ultra-high-performance liquid chromatography-tandem mass spectrometry was employed to analyze and characterize the chemical ingredients of ZXSZD. An in vitro cell model was established by stimulating HK-2 cells with High Glucose-Palmitic Acid (HGPA) to mimic the metabolic microenvironment of DKD. Oil Red O staining was performed to assess intracellular lipid accumulation. Western blot and immunofluorescence assays were used to determine the protein expression levels of key molecules in the AMPK/ PPARα/CPT1A signaling pathway. Additionally, mitochondrial membrane potential, intracellular ATP content, activities of mitochondrial respiratory chain complexes (I and IV), oxidative stress levels, and concentrations of inflammatory cytokines were quantified.

Results

A total of 141 chemical ingredients were preliminarily identified in ZXSZD. In HGPA-induced HK- 2 cells, ZXSZD significantly attenuated intracellular lipid accumulation, upregulated the protein expression of p-AMPK, PPARα, and CPT1A, ameliorated energy metabolism impairment and mitochondrial dysfunction, and suppressed oxidative stress as well as inflammatory responses.

Discussion

The results indicate that ginkgolide effectively binds and stabilizes the ApoE4 N-terminal domain, supporting its potential role in modulating a key pathological factor in Alzheimer's disease. Its superior pharmacokinetic profile and interaction dynamics compared with those of curcumin and resveratrol suggest broader therapeutic relevance. These in silico insights provide a mechanistic basis for further investigation into ginkgolide's neuroprotective effects.

Conclusion

ZXSZD ameliorates energy metabolism impairment and mitochondrial dysfunction in HGPAinduced HK-2 cells by activating the AMPK/PPARα/CPT1A signaling pathway, while also inhibiting oxidative stress and inflammation.