Study On The Antioxidant Effect Of Low Molecular Weight Fucoidan On Atherosclerosis In ApoE-/-Mice

Objective:Atherosclerosis（AS）is the pathological basis of cardiovascular and cerebrovascular disease,the most important cause of coronary heart disease,aortic aneurysm,cerebral infarction,and peripheral vascular disease.At present,there are still some side effects of drugs used to treat AS clinically,so it is urgent to find a new drug,which is non-toxic,safer for the prevention and treatment of AS.As a natural marine medicine,low molecular weight fucoidan（LMWF）has a variety of structures and activities.This study aims to investigate the therapeutic effect and antioxidant mechanism of LMWF on atherosclerosis in Apo E-/-mice,so as to provide theoretical basis for treatment.Methods:（1）LMWF anti-oxidation experiment:the scavenging rates of LMWF to O₂^-˙and˙OH were determined by colorimetric method with catechol and the method of ferrous sulphate-salicylic acid.（2）Cell experiment:peritoneal macrophages were randomly divided into control group,model group,low concentration group,high concentration group,and probucol group,and cultured in 10%fetal bovine serum in DMEM medium.In addition to the control group,the other four groups were treated with oxidized low-density lipoprotein（ox-LDL）.After cultured for 24,48 and 72 h,the levels of reactive oxygen species（ROS）,superoxide dismutase（SOD）and glutathione peroxidase（GSH-PX）in five groups of cells were detected,cell inhibition was detected using CCK8 method,and cell apoptosis was detected by flow cytometry.（3）Animal experiment:50 SPF male Apo E-/-mice were randomly divided into control group,model group,fucoidan group,probucol group,and combination group.In addition to mice in the control group fed standard mouse food,mice in the other four groups were given atherogenic high-fat diet to establish AS model.All treatments were performed by gavage.The mice were randomly selected from each group,the heart,liver and aortic tissues of the mice were taken out.Hematoxylin-eosin（HE）staining and 3:2 saturated oil red O（Oil red O）staining were used to observe the morphological changes of aortic tissues in mice.The total antioxidant capacity（T-AOC）,malondialdehyde（MDA）,SOD,GSH-PX,ROS and ox-LDL levels of mouse heart and liver tissues were detected.Results:（1）Antioxidant results of LMWF:results showed that LMWF had significant antioxidant activity,and the IC₅₀ of the scavenging rates of O₂^-˙and˙OH were 0.047 mg/mL and 0.098 mg/mL,respectively.（2）Cell experiment results:compared with the control group,ROS level and apoptosis rate in the model group were significantly increased（P<0.05）,the level of SOD,GSH-PX and cell inhibition rate decreased significantly（P<0.01）.Compared with the model group,ROS levels in the low concentration group,the high concentration group and the probucol group were significantly reduced（P<0.05）.The levels of SOD,GSH-PX,cell inhibition rate and cell apoptosis rate increased significantly（P<0.05）.Except for the cell inhibition rate,the group with high concentration of other indicators had the best effect among the three groups（P<0.05）.（3）Animal experimental results:the plaque area of the model group was significantly larger than that of the control group（P<0.001）.The plaque area of the fucoidan group,the probucol group and the combination group were significantly smaller than that of the model group（P<0.001）,but larger than that of the control group（P<0.05）.Compared with the control group,the antioxidant capacity,SOD level and GSH-PX level of the model group were significantly reduced（P<0.01）.The MDA level was significantly increased（P<0.001）.Compared with the model group,the antioxidant capacity,SOD level and GSH-PX level of the fucoidan group,the probucol group and the combination group were significantly increased（P<0.05）,and the MDA level was significantly reduced（P<0.001）,almost reaching the normal level,there was no statistically significant difference between the three groups（P>0.05）.Conclusion:LMWF has a strong antioxidant capacity,and shows significant inhibition and improvement on AS.LMWF can enhance the inhibition rate of peritoneal macrophages and promote their apoptosis,inhibit the production of MDA,ROS and ox-LDL,increase the activity of SOD and GSH-PX,and improve T-AOC of the body.The strong antioxidant activity of LMWF may be one of the important mechanisms to improve the occurrence and development of AS.