Qiangyang decoction attenuates ischemia/reperfusion-induced cardiomyocyte ferroptosis via ULK1/KEAP1/Nrf2 axis

Myocardial ischemia-reperfusion (I/R) induced cardiomyocyte death is a major cause of cardiac dysfunction and fibrosis. Ferroptosis of cardiomyocytes has been demonstrated to contribute to I/R induced injury. Qiangyang decoction (QYD) is commonly used in clinical treatment of hypertension and cardiac fibrosis. However, the intervention effect of QYD on I/R injury, especially its therapeutic efficacy on ferroptosis in cardiomyocytes, remains unclear. This study aimed to investigate the effect of QYD against I/R induced cardiomyocyte ferroptosis and to reveal the underlying mechanism. In this study, UPLC-MS/MS was conducted to identify the component of QYD. An in vivo I/R mouse model and an in vitro cardiomyocytes OGD model were utilized to evaluate the cardiac protective effect of QYD. Transcriptome analysis and western blotting assays were used to explore the mechanism. A high dose of QYD exerts a cardioprotective effect through inhibiting I/R-induced cardiomyocyte ferroptosis. A high dose of QYD ameliorated I/R and OGD injury; these beneficial effects were not further enhanced by the ferroptosis inhibitor Fer-1. Anti-ferroptosis factors FPN1, Homx1, and GPX4 were transcriptionally upregulated by QYD both in mouse heart and adult mouse cardiomyocytes (AMCMs). Consistently, the kelch like ECH associated protein 1 (KEAP1) level and nuclear factor erythroid 2-related factor 2 (Nrf2) degradation were remarkably suppressed. RNA-sequencing results revealed that unc-51 like autophagy activating kinase 1 (ULK1), a serine kinase mediating the autophagy-dependent degradation of KEAP1, was significantly up-regulated in QYD-treated mouse hearts. The ULK1 inhibitor SBI-0206965 abolished the inhibition of QYD on KEAP1 and notably weakened the cardiac protective effect. These findings suggest that QYD exerted cardiac protection by activating ULK1 and subsequently inhibited KEAP1, thereby enhancing Nrf2 level and initiating the transcription of ferroptosis-resistance genes. These findings highlight that QYD may serve as a cardioprotective strategy for I/R.