Research Mechanism Of Salidroside Inhibiting The Formation Of Foam Cells In Atherosclerosis Based On NLRP3 Inflammatory

Objective:Atherosclerosis?AS?is one of the important pathological bases of coronary heart disease,cerebral infarction and peripheral vascular disease,which seriously endangering human life and health.The formation of foam cells is a marker for the formation of atherosclerotic plaque,and NLRP3 participates in the process of the formation of atherosclerotic foam cells,and NLRP3is activated to produce a series of inflammatory reactions.In this paper,in vivo and in vitro experiments,the effect of salidroside on the inhibition of the formation of foamy cells,inhibiting the inflammatory reaction,reducing the formation of plaque,and playing the role of anti AS based on NLRP3 in vitro and in vitro..Methods:1.In vivo experiment:6 weeks old ApoE^-/-male mice were adaptively fed for 1 week and randomly divided into model group,positive group?atorvastatin,high dose of salidroside?and high dose of salidroside.Salimdroside-H,salidronside-L,a high-fat diet?21%fat,0.15%cholesterol?was given throughout the entire course?12 weeks?to prepare atherosclerotic models;with the same inheritance Eight C57BL/6/J mice were used as normal control group and were given normal feed for 12 weeks.From the first week of model establishment,the normal control group and model group were given intragastric administration of an equal volume of normal saline.The positive group was given atorvastatin?10mg/kg?orally,and the high and low dose group of salidroside was given 50 mg/kg and 25mg/kg gavage;12 weeks later,blood was taken from the eyeball,serum was separated,serum lipids?TC,TG,HDL-C,LDL-C?were detected;internal aorta oil red O staining was used to observe the overall plaque;frozen Oil red O staining was used to observe the lipid area in atherosclerotic plaques;ELISA was used to detect IL-1?and IL-6,TNF-?,MCP-1 secretion;Immunohistochemical method to detect NLRP3,LC3b,ABCA1expression in plaques.2.In vitro experiments:1)Induced transformation of THP-1 cells;2)Extraction and purification of mouse peritoneal macrophages;3)Oil red O staining;4)Determination of IL-1?,TNF-?,IL-6,MCP-1 by ELISA;5 Western blot was used to detect the expression of NLRP3 and LC3b proteins.Results:1.In vivo experiments:1)Four results of biochemistry:Compared with the normal group,the TC,TG,and LDL-C levels in the model group were significantly higher?P<0.001?.Compared with the normal group,the HDL-C in the model group was decreased,but there was no statistic.Significance;Compared with the model group,atorvastatin can significantly reduce TC,with statistical significance?P<0.01?,high dose of salidroside,low dose of salidroside can reduce TC,statistically significant?P<0.05?;compared with the model group,atorvastatin and salidroside reduced TG at a high dose?P<0.05?;compared with the model group,salidroside had a high dose There was a statistically significant increase in HDL-C?P<0.05?.Compared with the model group,atorvastatin,high dose of salidroside,and low dose of salidroside all reduced LDL-C,but there was no statistic Meaning.2)Oil Red O staining on the inner surface of the aorta:atorvastatin and salidroside reduced the plaque area to some extent.3)Frozen oil red O staining:The amount of macrophages in plaques was reduced by salidroside to a certain degree?P<0.05?.Macrophages were the main source of foam cells,so they inhibited the formation of foam cells.Reduced lipid content in plaques.4)Immunohistochemistry:Compared with the model group,the expression of ABCA1 was significantly increased in the high-dose group of salidroside?P<0.05?.Compared with the normal group,the NLRP3 of the model group The expression was obvious.Compared with the model group,the expression of NLRP3 in the salidroside high-dose group was significantly decreased?P<0.05?.Compared with the normal group,the model group had no obvious expression of LC3b.Compared with the model group,there was no obvious expression in the atorvastatin group.LC3b expression was significantly increased in the low-dose salidroside group,and the brown particles were significantly?P<0.05?.Therefore,it was known that salidroside highly expressed ABCA1 and LC3b.Can reduce NLRP3,expression.5)ELISA:compared with the normal group,the IL-1?,IL-6,MCP-1 and TNF-?in the model group were significantly higher?P<0.05?,compared with the model group,Atu The high doses of fluvastatin and salidroside significantly decreased the levels of IL-1?,IL-6,TNF-?,and MCP-1,which was statistically significant?P<0.05?.Low dose of salidroside significantly reduced IL-6 The level of MCP-1 was statistically significant?P<0.05?.2.In vitro experiments:1)THP-1 macrophage can be induced when THP-1 cells are stimulated with 200 nm PMA for 48 h.2)2ml broth was injected into C57BL/6/J mice.72 hours later,mouse peritoneal macrophages were extracted and purified by adhesion.3)The effect of salidroside on THP-1 cells and mouse peritoneal macrophages:The safe drug concentration of salidroside by MTT assay is the dose of salidronside high dose?Salidroside-H?200?M.The salidroside low-dose group?Salidroside-L?dose was100?M.4)Oil Red O staining:50?g/ml ox-LDL stimulated foam cells after THP-1 macrophage48 h,and salidroside high dose group and salidroside low dose group significantly reduced lipid droplets?visible to the naked eye?.After 4 hr/m L LPS stimulation,salidroside was added for 2 h.Finally,1 mg/m L cholesterol crystal was added to stimulate mouse peritoneal macrophages to become foam cells,salidroside high dose group,and salidroside low dose group.It also significantly reduces lipid droplets?visible to the naked eye?.5)ELISA assay:THP-1cells:compared with the normal group,the IL-1?,IL-6,MCP-1 and TNF-?in the model group were significantly higher?P<0.05?;In comparison,Salidroside-H significantly reduced the levels of IL-1?,TNF-?,and MCP-1,which was statistically significant?P<0.05?.Salidroside-H had a low dose?Salidroside-L?.5)The level of IL-1?and TNF-?was significantly decreased?P<0.05?.Mouse peritoneal macrophages:Compared with the normal group,the model group was significantly different from the normal group?P<0.001?;compared with the model group,salidroside high-dose group?Salidroside-H?,red Salidroside low-dose group?Salidroside-L?significantly reduced the secretion of IL-1?and TNF-??P<0.01?;6)Western blot analysis of NLRP3 and LC3b protein expression:Compared with the model group,Salidroside inhibited the expression of NLRP3 protein?P<0.05?Conclusions:In vitro and in vivo studies have shown that salidroside inhibits the expression of NLRP3,inhibits IL-1?and other inflammatory factors,increases autophagy,promotes cholesterol excretion,inhibits the formation of foam cells,and plays an anti-AS role.