Effect Of Yiqi Huayu Recipe On Lipid-lowering Effect And Mechanism Of Action In Hypercholesterolemic Rats

ObjectiveThis experiment in hypercholesterolemic rats as the research object,observe the effect of Yiqi Huayu Prescription on lip id-lowering,liver protection and protection of vascular and other aspects of the role,and at the same time by metabonomic method,metabolic markers of liver in rats,the experimental data obtained from the analysis,find out the related pathway of Yiqi Huayu Prescription decreased in rats the possible mechanism of blood lipid,and from the aspect of Metabonomics Study of Yiqi Huayu Prescription on hypercholesterolemia,laying the groundwork for further research.Method60 rats in 7 days after the real animal feeding,according to randomly divided into 6 groups:control group,hyperlipidemia model group,positive drug group,Yiqi Huayu Prescription,high,low dose group,10 rats in each group.After the experiment began,the rats were in normal water.Fed the basal diet,the rats in the blank group with distilled water,the other 5 groups with high fat emulsion by gavage.High fat emulsion formula:20%lard,5%cholesterol,5%egg yolk powder,1%propylthiouracil,6%Twain-80,63%distilled water gavage dose.According to 10ml/Kg,for 3 weeks.From the beginning of the fourth week,the rats according to law before feeding.At the same time,the control group used normal saline,model group were administered according to the dose of 10ml/Kg.Yiqihuayu group respectively according to the corresponding dose by intragastric administration of high,low dose,respectively,equivalent to the amount of crude drug 18G,9g,4.5g/Kg..Using Lipitor suspension dosage in accordance with 10mg/Kg,administered orally,daily intragastric dose of 10ml/Kg,1 times daily,administered continuously for 4 weeks.In the last administration before 12 hours,the rats underwent fasting but not water feeding.The principle of the morning fasting were weighed by 10%chloral hydrate solution according to the doses of 0.35ml/100g were anesthetized with intraperitoneal injection.After aortic blood,the blood taken 1 hours after the static at room temperature.At 3000 rpm for 15 minutes after centrifugal separation of serum,split after placed in the refrigerator at-20 DEG C to be measured.Take the blood after the rats were killed and the liver removed,and observed liver morphology,rats were photographed retained.Rinse immersion in saline to wash away the blood,after weighing filter paper blot moisture.Results with the combination of the weighing weight,is used to calculate the liver index of rats,the calculation method is:the liver index =liver weight/body weight(g)*100%.(g)of liver tissue of each rat liver left lobe(5 mm*5mm*3mm),fixed and immediately placed in 4%poly Formaldehyde Solution.Open the rat thoracic aortic arch,gently separated about 1cm in rat aortic tissue,and immediately put into more than 4%fixed together in Formaldehyde SolutionThe use of automatic biochemical analyzer,detection of four blood lipids of rats and other serum enzymes.The liver index and thoracic aorta was embedded in paraffin,HE staining respectively.Then to observe the pathological changes of the liver and thoracic aorta of rats under microscope.To detect the expression of rat thoracic aorta sPLA2 Ⅱ A and NF-K B.Using immunohistochemical method and Image Pro.Plus6.0 analysis of the average optical density(MOD).The detection of serum TNF-α,IL-6 level,using enzyme-linked immunosorbent assay(ELISA)method.The use of ultra high performance liquid chromatography time of flight mass spectrometry(UHPLC-QTOF-MS)detection,high fat stomach induced hypercholesterolemia in rat liver metabolic markers.After obtaining the data by using PC A,PLS-DA and OPLS-DA model to analyze the data,finally,according to the VIP value,there are differences in screening biomarkers through the MetPA analysis,identify accurate biomarkers and related metabolic pathways,and evaluate the efficacy and the modelResult1 serum test results and liver morphological changesCompared with the control group,the rats of TC model group,LDL-C level increased significantly(P<0.01).Compared with the model group,positive control group and Yiqi Huayu Prescription of low dose group rats TC,LDL-C levels were significantly decreased(P<0.01,P<0.05).The positive control group and Yiqi Huayu Prescription in each dose group of rats ALT,AST,CRE,UREA,CK,LDH levels were significantly decreased(P<0.01 or P<0.05).The pathological sections showed that liver fat cells of rats in model group were fat infiltration,and Yi Qi stasis prescription in each dose group and positive group of rat liver were basically normal.2 Expression of inflammatory factors in thoracic aorta of ratsThe model group of rat aortic smooth muscle layers.Some disorders of Yiqihuayu prescription in each dose group and positive group of rat thoracic aorta was normal.Compared with the model group,the expression level of positive group and Yiqi Huayu in each dose group of rat thoracic aorta sPLA2 Ⅱ A and NF-K B were significantly lower in model group than in the blank group.The increase in the expression of these two indexes.The serum TNF-a and IL-6 levels,also shows the results.3 differential metabolites in liver tissueBased on the difference between positive and negative ion metabolite MetPAana lysis showed that higher between the model group and the blank group high fat emulsion model in rat serum TC and LDL-C,may be related to metabolic pathways including folate one carbon metabolism and glycerophospho lip id metabolism,lipid-lowering effect of Yiqi Huayu Prescription;low dose group and folic acid maybe a carbon metabolism pathway,pentose and glucuronate conversion pathway of these two pathways.ConclusionYiqihuayu prescription can significantly decrease the serum cholesterol level of rats in the model group,the liver and kidlney in rats at the same time for high fat caused damage,also has a certain regulating effect of Yiqi Huayu Prescription.Can inhibit the infiltration of fat on the liver,improve the morphological changes of rat thoracic main artery,through inhibition of inflammatory factors role play,good prevention of vascular endothelial injury.Through the analysis of the liver metabolites of UHPLC-QTOF-MS detection and relevant data found that establish hyperlipidemia model may be associated with metabolic pathways including folate one carbon metabolism and glycerophospho lipid metabolism,and lipid-lowering effects of Yiqi Huayu Recipe low dose group and folic acid may play a carbon metabolism pathway,pentose and glucuronate conversion pathway of these two pathways.