Na₂SiO₃ Attenuates High-fat Diet–induced Arterial Intima Injury In Rats

Atherosclerosis is an inflammatory disease that is the main reason for cardiovascular events leading to death in the world.Accumulated evidences show that atherosclerosis is driven by inflammation,with involvement of inflammatory factors and immunity.It also involves several cell types including smooth muscle cells,lipids,monocytes,macrophages,dendritic cells,and extracellular matrix,and eventually leads to plaque rupture and thrombosis.In previous studies,silicon play a role in the prevention and treatment of diseases of the cardiovascular system and can maintain the vessel wall elasticity.We also found that Na₂SiO₃ can improve hardening of the arteries of rabbits and rats in our lab.Nox4?NADPH oxidative enzyme,Nox?is the most important source of vascular active Oxygen?Reactive Oxygen Species,ROS?,which leads to a series of inflammatory factors change and will affect the structure and function of vascular endothelium.ROS activates the NF-?B directly,or make the NF-?B activation from the other indirect way.Research has shown that endogenous ROS can activate the NF-?B,resulting in monocyte chemotactic protein 1?MCP-1?expression.Objective: To research the effect of Na₂SiO₃ on the rats' vascular intimal injury,we established animal model by feeding high-fat-diet and injection of vitamin D3.Serum TG,TC,LDL,HDL were detected,then we further measured expression of nuclear factor ?B-p65?NF-?B-p65?,Monocyte chemoattractant protein 1?MCP-1?and NADPH oxidative enzyme 4?Nox4?in rat artery tissue by using immunohistochemical stains and molecular biological techniques in order to explore the potential mechanisms of Na₂SiO₃in the prevention and treatment of intimal hyperplasia by feeding with high-fat-diet in rat.Method: Male Sprague-Dawley rats were randomly divided into five groups?n=7?.Control group?Con?,High-fat-diet group?HFD?,HFD+Na₂SiO₃ Low dose group?HFD+ Na₂SiO₃ 5mg/kg?,HFD+ Na₂SiO₃ high dose group?HFD+ Na₂SiO₃ 10mg/kg?,HFD+Statins group?HFD+Statins 5mg/kg?.All of the rats were fed 7 days to adapt the environment,then VD3?600000IU/kg?were injected into the abdominal cabity beside control group,and meanwhile,feeding with high-fat-diet,then 300000 IU/kg VD3?injection,every other day for 3 times?were injected in the 4th week,Control group and HFD group were fed with purified water,different dose of drug-treated groups were given Na₂SiO₃,statins group were given statins?5mg/kg?by gavage,all treatments lasted for 10 weeks.Fasting for 24 hours,weigh and anesthesia the rats with 3% chloral hydrate,take blood,separate the aorta and take 1 * 1 cm size of liver tissue.All of the tissues were cut to 2 pieces,one fixed in 4% paraformaldehyde,another tissue samples were immediately frozen in liquid nitrogen and stored at-80?C.We use biochemical method to detect serum lipid levels.The aorta and liver histological changes in rat were observed by hematoxylin eosin?HE?dyeing.Results:1 Effects of Na₂SiO₃ on blood TC,TG,LDL and HDL in high-fat-diet rats In this experiment,we test TC,TG,LDL and HDL in serum.TG of the HFD group was increased compared with the control group;and Na₂SiO₃treated group have no significant differences.statins treated group were decreased.TC was higher compared with the control group and the HFD group;statins and Na₂SiO₃ treated group were decreased.Compared with the control group,LDL was higher in the HFD group,and Na₂SiO₃ groups have reduced LDL level.Compared with the control group,HDL was higher in the HFD group;high dose group of Na₂SiO₃ and statins group have reduced serum HDL level.2 Effects of Na₂SiO₃ on histopathologic changes in rats' liver Compared with normal group,more vacuolated lipid droplets can be seen in the liver cells in model group with HE staining;compared with model group,fatty degeneration of liver tissue was ameliorated in Na₂SiO₃treatment group.Vacuoles degeneration reduced obviously in statin group.3 Effects of Na₂SiO₃ on histopathologic changes in rats' aorta HE staining showed that compared with the normal group,significant intimal thickening and disorder arrangement of the medial smooth muscle were observed in the model group;compared with the model group,vascular intima was smooth with mild intimal thickening in low dose of Na₂SiO₃ group;the intima was smooth without thickening and medial smooth muscle arranged in neat rows in high dose group.Vascular intima is relatively smooth in statin group.4 Effects of Na₂SiO₃ on the aorta Nox4 protein expression Immunohistochemistry show that Nox4 was low expressed in the normal aorta;the expression of Nox4 was elevated in the HFD model group compared with the control group;compared with the model group,Nox4 was decreased in Na₂SiO₃ and statin treated groups Western blot show that the protein levels of Nox4 were increased significantly in the model group compared with the control group;while in the statin and Na₂SiO₃ treated groups,the protein levels of Nox4 were markedly decreased compared with the model group.5 Effects of Na₂SiO₃ on the aorta of NF-?B-p65 protein expression and activation Immunohistochemical results show that the NF-?B-p65 was low expressed and dispersed in the cytoplasm in the control group;the NF-?B-p65 protein expression was increased significantly in the model group,and the particle aggregation,from the cytoplasm to the nucleus;statin,low and high dose group of Na₂SiO₃ have reduced NF-?B-p65 expression compared with the model group.Western blot results show that NF-?B-p65 protein expression is less in the control group,the NF-?B-p65 expression and translocation into the nucleus was obviously increased in the model group,and Na₂SiO₃ and statin can decrease the protein expression and inhibit translocation into the nucleus of NF-?B-p65.6 Effects of Na₂SiO₃ on the aorta MCP-1 protein and m RNA expression The results indicate that the expression of MCP-1 was significantly increased in the model group compared with those in control group;statin and the low and high dose of Na₂SiO₃-treatment can markedly decreased MCP-1levels.RT-PCR results show that MCP-1 m RNA expression was increased obviously in the model group;statin and low and high dose group of Na₂SiO₃treatment can down-regulate MCP-1 m RNA expression level.Conclusions: The results of these studies suggested that Na₂SiO₃ could inhibit high-fat diet–induced arterial intima injury in rats,and this mechanism may be related with inhibiting the Nox4/NF-?B signaling pathway and reduce inflammatory factor expression.