Shenfuyixin Granules enhance mitochondrial autophagy after myocardial infarction by regulating protein deacetylation via the SIRT3/FOXO1 signaling axis

Abstract

Background

Heart failure (HF) represents the terminal stage of various cardiovascular diseases, with current treatment options remaining limited. Shenfuyixin Granules (SFYX) have been integrated into clinical practice, demonstrating significant therapeutic efficacy. However, the underlying mechanisms of action are still not fully understood.

Purpose

This study aims to investigate whether SFYX promotes mitochondrial autophagy and enhances cardiac function in HF following myocardial infarction via the SIRT3/FOXO1 signaling axis.

Methods

The rat model of HF was established by ligation of the left anterior descending artery, while in vitro experiments were conducted using H9C2 cells. The blood-entry components of SFYX were identified using ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS). Network analysis, integrating proteomics and transcriptomics, was conducted to determine the active components of SFYX and elucidate the key regulatory mechanisms involved in its treatment of HF. After a 4-week intervention with SFYX, cardiac function was assessed via echocardiography. Myocardial infarct size was measured using triphenyl tetrazolium chloride (TTC) staining, while H&E and Masson staining were employed to evaluate myocardial tissue fibrosis and hypertrophy. Mitochondrial function was assessed using transmission electron microscopy and JC-1 dye. Cell apoptosis was detected via TUNEL assay. Additionally, molecular docking was performed to assess the binding affinity between key components of SFYX and autophagy-related proteins. Mechanistically, the expression levels of SIRT3, FOXO1, P62, and BNIP3 were determined using quantitative PCR and Western blotting.

Results

UPLC-MS/MS analysis revealed 21 blood-entry components in SFYX. Integrated analyses of network pharmacology, proteomics, and transcriptomics indicated that SFYX may ameliorate HF by stimulating mitochondrial autophagy through activation of the SIRT3/FOXO1 pathway. Compared with the model group, SFYX significantly attenuated myocardial hypertrophy, apoptosis, and fibrosis while enhancing autophagy, which may be partially attributed to the recovery of mitochondrial function. We propose that SFYX enhances mitochondrial function by reducing membrane potential and reactive oxygen species (ROS) production. Further results demonstrated that SFYX treatment upregulated SIRT3 and FOXO1 levels while inhibiting FOXO1 acetylation. Furthermore, the levels of mitophagy-associated proteins (ATG5, ATG7, BNIP3, and LC3B-II), which are downstream mediators of FOXO1, were enhanced by SFYX. Activation of SIRT3 or overexpression of FOXO1 enhanced the cardioprotective efficacy of SFYX, whereas inhibition of SIRT3 or silencing of FOXO1 partially reversed SFYX-induced favorable activities. Molecular docking analysis revealed that Glyceryl linolenate, a blood-entry component of SFYX, exhibited a strong binding affinity for SIRT3.

Conclusion

This study demonstrates that SFYX exerts cardioprotective effects against HF through the deacetylation-regulated activation of SIRT3/FOXO1 signaling-mediated mitophagy and apoptosis. These findings indicate that SFYX represents a promising therapeutic candidate for the treatment of HF.