Effect And Mechanism Of Xinsuning Capsule On Arrhythmia Induced By Isoproterenol In Rats

Objective:Based on the research on the substance basis of Xinsuning capsule,the substance distribution and time-varying rule of Xinsuning capsule in plasma and myocardial tissue were analyzed.The model of arrhythmia induced by subcutaneous injection of isoprenaline hydrochloride into the left lower limb of rats was adopted,and the pharmacodynamics of all components and active components of Xinsuning capsule were evaluated.The pathway of "blood-entering components-disease target" was predicted,so as to explore the anti-arrhythmia mechanism of active components of Xinsuning capsule.Method and Results:1.The first part is the basic research on the pharmacodynamic substance of Xinsuning.Experiment 1: Analysis of migrating components in blood of Xinsuning capsule and discussion on the regularity of changes with time.Method: The rats were given Xinsuning suspension(12g/kg)by gavage,with the dosage of 10ml/kg.After 0h,0.083 h,0.167 h,0.25 h,0.333 h,0.5h,1h,2h,4h,6h,the samples were treated and analyzed by UPLC-MS/MS/MS.Results: An analytical method of multi-component content determination of Xinsuning capsule was established by LC-MS/MS,which realized the analysis of six blood components in rat plasma,namely sophocarpine,matrine,berberine,palmatine,citrin and nobiletin.Experiment 2:Analysis of material distribution of Xinning capsule in myocardial tissue and its time-varying regularity.Method: After fasting for 12 hours,30 SD rats were randomly divided into 5 groups(divided into groups according to the administration time),6 rats in each group were given Xinsuning suspension(6g/kg)by single intragastric administration,and then decapitated at0.25 h,0.5h,1h,2h,4h after intragastric administration,then blood was taken,and after perfusion and washing with normal saline,a small amount of congestion and tissue inclusions were removed.The other 6 experimental rats were taken as the blank control group,and the blank control group was given the same volume of normal saline,and the samples were taken in the same way.Results: There are five components in myocardial tissue,namely sophocarpine,matrine,berberine,citrin and nobiletin.2.The second part is based on the prediction of effective targets of "blood components-disease targets" of network pharmacology.Method: Methods: The target of drug components was detected from TCMSP database,and then related targets of arrhythmia were detected by OMIM,Pharm GKB,Genecard and TTD database systems,so that the common target of drug components and related targets of arrhythmia was detected,which was used as the predicted target of Xinsuning capsule in treating arrhythmia,and uploaded to STRING database to establish protein-protein interaction(PPI)network.Through Cytoscape 3.8.0,PPI network is screened according to connectivity,and its key targets are found.DAVID6.8 makes GO and KEGG pathway enrichment analysis on key targets.Results: The results of GO and KEGG pathway enrichment analysis show that Calcium signaling pathway can be used as the main pathway of arrhythmia.3.The third part is the study on the effect and mechanism of Xinsuning capsule on isoproterenol-induced arrhythmia in rats.Experiment 1: The effect of all components of Xinsuning on arrhythmia induced by isoproterenol in rats and the content of Na+-K+-ATPase in myocardial tissue.Method: Thirty SD rats were divided into five groups,namely blank group,model group,positive drug group(betaloc 2.1mg),low-dose group(0.6 g/kg)and high-dose group(H group,1.2 g/kg,twice the clinical equivalent dose),with 6 rats in each group.After seven days of adaptive feeding,the blank group and model group were given intragastric administration of the same volume.On the 7th day,30 minutes after intragastric administration,rats in each group were anesthetized by intraperitoneal injection of 10% chloral hydrate at 3.5ml/kg.Except for the blank group,the rest of the four groups were injected with isoproterenol solution at 4 mg/m L subcutaneously in their left lower limbs,and the injection was completed within 5 seconds.In the blank group,the same volume of normal saline was injected subcutaneously in their left lower limbs,and ECG was recorded.Results: Compared with the model group,the low and high doses of Xinsuning can obviously delay the occurrence and duration of arrhythmia,with statistical difference(P<0.05,P < 0.01).Compared with the model group,Xinsuning high-dose group can obviously reduce the duration of arrhythmia,with statistical difference(P < 0.001).Compared with the model group,the content of Na+-K+-ATPase in both low and high doses of Xinsuning can be increased,with statistical difference(P<0.001,P<0.01).Experiment 2: Study on the effect and mechanism of methanol extract from Xinsuning Capsule 6 on arrhythmia induced by isoproterenol in rats.Method: SD rats were divided 30 minto 5 groups: blank group,model group,positive drug group(Betaloc 2.1mg),low-dose group(0.4/kg)and high-dose group(H group,0.8g/kg,twice the clinical equivalent dose),with 6 rats in each group.After 7 days of adaptive feeding,the blank group and model group were fed with the same volume of physiological saline.Rats in each group were anesthetized by intraperitoneal injection of 10% chloral hydrate at3.5ml/kg.Except for the blank group,the other four groups were injected with isoproterenol solution at 4 mg/m L under the skin of the left lower limb,and the injection was completed within 5 s.The blank group was injected with the same volume of normal saline under the skin of the left lower limb,and the electrocardiogram was recorded.Results: The quantitative results of active components in Xinsuning capsule showed that the content of six blood components extracted from Xinsuning by methanol active components was higher than that of Xinsuning,and the content of matrine extracted from methanol active components was higher than that of sophocarpine,and the content of berberine was higher than that of palmatine,which changed from the original content ratio of Xinsuning.Compared with the model group,the low-dose and high-dose groups of Xinsuning significantly delayed and shortened the occurrence and duration of arrhythmia,with statistical differences(P<0.01,P<0.001).Compared with the model group,Xinsuning capsule(active component)can provide Na+-K+-ATPase in both low and high doses,with statistical difference(P<0.05,P<0.01).The results of pathway verification show that the active components of Xinsuning Ca N activate the Calcium signaling pathway to resist arrhythmia by increasing the contents of RYR,CAN and PKC in myocardial tissue.Conclusion:1.In plasma,matrine has the longest biological half-life and nobiletin has the shortest biological half-life;In myocardial tissue,the highest concentration of matrine(217.00±6.43ng/ml)and the lowest concentration of nobiletin(1.88±0.16 ng/ml)were found.2.The enrichment analysis of GO and KEGG pathways shows that the Calcium signaling pathway can be used as the main pathway of arrhythmia.3.All components and active components of Xinsuning can delay the occurrence of arrhythmia and shorten the duration of arrhythmia;It can increase the content of na+-k+-ATPase.The results showed that the active components of Xinsuning could activate the Calcium signaling pathway by increasing the concentration of RYR,Ca N and PKC in tissues to resist arrhythmia.