Polydatin in combination with hawthorn flavonoids alleviates atherosclerosis by inhibiting mitochondrial division through the YTHDF2-DRP1 axis.

INTRODUCTION: The combination of polydatin and hawthorn flavonoids (PH), a traditional Chinese medicine formulation for activating blood circulation and detoxification, has shown the potential to counteract atherosclerosis, but the mechanisms underlying its effects remain unclear. PURPOSE: To investigate the protective effects of pH on atherosclerosis and identify the key targets underlying mitochondrial homeostasis. METHODS: An ApoE-/- mouse model fed a high-fat diet (HFD) was established to evaluate the effect of pH on aortic plaques, and an oxidized low-density lipoprotein (ox-LDL)-induced human umbilical vein endothelial cell (HUVEC) injury model was established. Dynamin-related protein 1 (DRP1) knockdown, YTHDF2 knockdown, and overexpression models were used to identify the key targets. RESULTS: PH exerted potent dose-dependent anti-atherosclerotic effects in high-fat diet-challenged ApoE-/-mice, reducing atherosclerotic lesion burden by 44.8%, 49.29%, and 72.99% at low, medium, and high doses, respectively. In vivo, PH ameliorated systemic dyslipidemia by lowering circulating total cholesterol, triglycerides, low-density lipoprotein cholesterol, and very-low-density lipoprotein levels, while robustly suppressing proinflammatory cytokine expression. PH also rescued aortic mitochondrial damage, mitigated mitochondrial fragmentation, and restored mitochondrial structural integrity, as evidenced by increased mitochondrial length, improved aspect ratio, and elevated mtDNA abundance. In ox-LDL-injured human umbilical vein endothelial cells (HUVECs), PH rescued endothelial mitochondrial dysfunction and restored normal mitochondrial architecture by reversing the ox-LDL-induced declines in mitochondrial matrix diameter and aspect ratio. Mechanistically, PH inhibited DRP1 expression and Ser616 phosphorylation, blocked mitochondrial translocation of phosphorylated DRP1, and thereby preserved mitochondrial membrane potential and mtDNA content. DRP1 knockdown abrogated PH's endothelial protective functions, verifying DRP1 as a core downstream effector. PH significantly upregulated YTHDF2 expression in ox-LDL-stimulated HUVECs. Functional assays confirmed a negative YTHDF2-DRP1 regulatory axis: YTHDF2 depletion increased DRP1 and phosphorylated DRP1 levels by 31.92% and 30.79%, whereas YTHDF2 overexpression reduced their levels by 43.88% and 38.31%. Notably, YTHDF2 loss abolished PH-mediated DRP1 suppression, indicating PH alleviates DRP1-dependent mitochondrial dysfunction via a YTHDF2-dependent mechanism. CONCLUSION: PH exerts a protective effect on HUVECs and prevents AS by regulating the YTHDF2/DRP1 axis to restore mitochondrial homeostasis, thereby providing a novel therapeutic approach for AS.