Study on the effect and mechanism of fraxetin on atherosclerosis based on ferroptosis regula-tion

Objective:To further explore the mechanism of fraxetin on atherosclerosis (AS)based on the regula-tory effect of fracetin on ferroptosis.Methods:An atherosclerosis model of ApoE knockout (ApoE^-/-) mice was established using a high-fat diet, and the ApoE^-/-mice were randomly divided into normal diet group(NC Group), high-fat diet group(HFD group), ferroptosis inhibitor group(Fer-1 group), fraxetin group(Fra group)and sulfora-phane group (Sul group).Primary human umbilical vein endothelial cells (HUVECs) were exposed to hydrogen peroxide (H₂O₂) to establish an in vitro AS model and were randomly divided into NC, H₂O₂, Fer-1, Fra and Sul groups.Oil red O staining was used to assess AS lesion size, and serum low density lipoprotein cholesterol(LDLC), serum high density lipoprotein cholesterol(HDL-C), and total cholesterol(TC)assay kits were used to assess lipid metabolism.Cell counting kit-8(CCK-8)assay was used to detect the toxic effect of fraxetin and cell prolif-eration ability, and the iron, glutathione(GSH)and malondialdehyde(MDA)assay kits were used to measure fer-roptosis levels in mouse aortic tissues and HU-VECs.Real-time fluorescence quantitative poly-merase chain reaction(RT-qPCR)and western blot-ting were used to examine the expression levels of glutathione peroxidase 4 (GPX4) and heme oxy-genase-1(HMOX1)to evaluate the anti-ferroptosis effect of fraxetin.Results:Fraxetin attenuated serum lipid de-position and aortic atherosclerosis in AS mice and protected HUVECs from H₂O₂-induced cell death.Consistent with the results of the Fer-1 group, fraxetin could inhibit the expression levels of pro-ferroptosis biomarkers iron and MDA and up-regulate the expression levels of anti-ferroptosis biomarkers GSH and GPX4 both in vivo and in vitro.In addition, fraxetin attenuated H₂O₂-induced ferroptosis in HUVECs, and this protective effect was re-versed by the HMOX1-specific activator sulforaphane.Conclusion:Fraxetin may inhibit ferroptosis by downregulating HMOX1, thereby reducing AS.