Abstract Body (Do not enter title and authors here): Background: Ecdysterone (Ecd), a bioactive compound derived from trianthema, exhibits potent anti-inflammatory properties. This study investigates the underlying mechanisms through which Ecd mitigates atherosclerosis (AS).
Methods: Human umbilical vein endothelial cells (HUVECs) were treated with 100 μg/mL (ox-LDL) for 24 h to induce endothelial injury. The cells were then treated with Ecd (40 μg/mL) for 24 h, and the protective effect of Ecd on Ox-LDL-treated HUVEC was evaluated by in vitro cell viability, ferroptosis, secretion of inflammatory factors, and levels of endothelial cell function markers (ET-1, eNOS, and NO). 1.5 μg/mL pcDNA-NCF2 was transfected into Ox-LDL-stimulated HUVECs for the recovery experiments. The PI3K/Akt/Nrf2 pathway inhibitor LY294002 (3 μM) and the pathway activator 1,3-Dicaffeoylquinic acid (10 mM) were used to confirm whether the pathway was involved in Ox-LDL-induced cell damage. ApoE^−/− mice were fed a high-fat diet to establish an AS model, Ecd (25 mg/kg) was fed every day, and 500 μL of Neutrophil cytoplasmic factor 2 (NCF2) lentivirus overexpression vector (1 × 10¹² PFU/mL) was injected into the tail vein every two weeks. Subsequently, levels of arterial ferroptosis and AS lesions in mice were analyzed.
Results: Ecd protected HUVECs from Ox-LDL-mediated inflammation and ferroptosis. NCF2 expression was upregulated in Ox-LDL-treated HUVECs and overexpression of NCF2 reversed this effect. In addition, Ecd activated the PI3K/Akt/Nrf2 pathway, whereas NCF2 promoted ferroptosis in Ox-LDL-treated HUVECs by inhibiting the PI3K/Akt/Nrf2 pathway. Further experiments showed that Ecd feeding reduced arterial ferroptosis in AS model mice, attenuated the levels of inflammatory markers, and alleviated AS.
Conclusion: The study demonstrates that Ecd ameliorates AS primarily by downregulating NCF2 and stimulating the PI3K/Akt/Nrf2 pathway, thereby inhibiting ferroptosis. These findings suggest Ecd as a promising therapeutic agent for AS by targeting ferroptosis-driven inflammation.