Study On The Effect Of Promoting Blood Circulation, Removing Blood Stasis, Clearing Heat And Detoxification In Regulating NLRP3 Inflammasome-mediated Pyroptosis And Anti-AS

Purpose:To investigate the mechanism of promoting blood circulation and removing blood stasis,heat-clearing and detoxification method to regulate NLRP3 inflammasome mediated pyroptosis and anti-AS by observing the changes ofblood lipid level and pyroptosis related protein expression and inflammatory cytokines m RNA expression in Aope gene knockout mice,and the intervention effect of Danlou tablet with gardenia and forsythia on Aope gene knockout mice.Material and method:Forty SPF healthy Aope E^-/-mice were fed with water and food freely.After adaptive feeding for 7 days,the 40 mice were randomly divided into 5groups:control group,model group,Danlou tablet group,Danlou tablet with gardenia and forsythia group and simvastatin group,with 8 mice in each group.The control group was given normal diet daily,the model group,Danlou tablet group,Danlou tablet with gardenia and forsythia group and simvastatin group were given high-fat diet daily,free eating and drinking water.Eight weeks later,the model group,Danlou tablet group,Danlou tablet with gardenia and forsythiae group and simvastatin group continue to high fat feed,at the same time to fill the stomach,to medicine dose conversion coefficients according to the equivalent dose conversion,Danlou tablet group,Danlou tablet with gardenia and forsythia group was given traditional Chinese medicine mixed suspension twice a day and simvastatin group was given simvastatin solution once a day.the model group was given such as volume of normal saline lavage.After 4 weeks of treatment,the levels of TC,TG,LDL and HDL in serum of mice were detected.Morphological changes of aorta were observed by hematoxylin-eosin(HE)staining.Theproteinexpressionsof NLRP3,GSDMD,caspase-1,IL-1?,IL-18,ASC and AIM2 in aorta were detected by Western Blot,and the m RNA expressions of IL-18,IL-1?,MCP-1 and MIP-1?in aorta were detected by RT-PCR.Results:1.The results of blood lipid level showed:Compared with the control group,the levels of TC,TG and LDL in serum of model group were significantly increased(P<0.05),while the level of HDL was significantly decreased(P<0.05).Compared with model group,serum TC,TG and LDL levels in Danlou tablet group,Danlou tablet with gardenia and forsythia and simvastatin groups were significantly decreased(P<0.05),and HDL levels were significantly increased(P<0.05).2.The results of HE staining showed:Compared with the control group,a large number of inflammatory cells were observed in the aortic tissue of mice in the model group,with the increase of foam-like cells and the formation of atherosclerotic plaques.Compared with the model group,the infiltration of inflammatory cells,foam-like cells and atherosclerotic plaques in aortic tissues of mice in Danlou tablet group,Danlou tablet with gardenia and forsythiae group and simvastatin group were significantly reduced.3.Western Blot analysis of NLRP3,GSDMD,caspase-1,IL-1?,IL-18,ASC,AIM2 protein expression in aorta showed:compared with the control group,the histone expression levels of NLRP3,GSDMD,caspase-1,IL-1?,IL-18,ASC,AIM2 in the aortic tissue of mice in the model group were significantly increased(P<0.05);Compared with model group,the histone expression levels of NLRP3,GSDMD,caspase-1,IL-1?,IL-18,ASC and AIM2 in the aortic tissue of mice in Danlou tablet group,Danlou tablet with gardenia and forsythia group and simvastatin group were significantly decreased(P<0.05).4.The m RNA expressions of IL-18,IL-1?,MCP-1and MIP-1?in aorta detected by RT-qPCR showed:compared with the control group,the expression levels of IL-18,IL-1?,MCP1,and MIP-1?in the aortic tissue of mice in the model group were significantly increased(P<0.05).Compared with the model group,the expression levels of IL-18,IL-1?,MCP-1 and MIP-1?in the aortic tissue of mice in Danlou tablet group,Danlou tablet with gardenia and forsythia group and simvastatin group were significantly decreased(P<0.05).Conclusion:1.Danlou tablet with gardenia and forsythia can ameliorate the level of blood lipid in atherosclerosis mice,reduce cell infiltration and plaque formation,can effectively prevent and cure atherosclerosis.2.According to the results of the study,the expressions of inflammatory factors and apoptosis related factors in serum were decreased,indicating that the anti-AS mechanism of Danlou Tablet with Gardenia and Forsythia is related to the regulation of NLRP3 mediated apoptosis.