Study On The Material Basis Of The New Medicated System Of Maxingshigan Decoction And Its Exploration Of Network Pharmacology

Objective:To explore the migration and transformation of the main active components of maxingshigan decoction in vivo by analyzing the main components of different ratios of water decoction,medicated intestinal absorption solution and medicated liver cell incubation solution in the treatment of asthma,and to study the applicability of the in vitro pharmacodynamics experiment of the new medicated liver cell incubation solution and to predict the potential action mechanism of maxingshigan decoction in treating asthma by network pharmacology.The purpose of this study is to provide experimental basis and theoretical reference for the further study of the material basis and mechanism of Maxingshigan decoction in treating asthma.Methods:1.Study on the screening of better proportion and the main difference components of water decoction in treating asthma:according to Un(n^s-1)of mixture uniform design table,different compatibility proportion of maxingshigan decoction was designed,and Rat asthma model was established by equal volume atomization of 2%acetylcholine chloride and 0.4%histamine phosphate.The rats were fed with different proportions of maxingshigan decoction（N1-N13 group）,dexamethasone sodium phosphate solution（positive group）and normal saline（normal group and model group）.The latent period of inducing asthma was recorded and the better ratio was selected.Using a triple quadrupole LC/MS instrument,the principal component analysis（PCA）and orthogonal partial least square discriminant analysis（OPLS-DA）of the components in the decoction of maxingshigan decoction were carried out by SIMCA13.0 to determine the main difference components in the treatment of asthma.2.Study on the main difference components of medicated intestinal absorption solution in treating asthma:the water decoction（N1-N13）was prepared by clinical decoction,and the medicated intestinal absorption solution（N1a-N13a）was prepared by the method of intestinal sac turning in vitro.the main difference components of asthma were determined by the same method as above,and the absorption of the main active components of the compound was investigated.3.Study on the material basis and applicability of the new medicated liver cell incubation solution:taking N13a as the object,three factors and three levels of experiments were designed according to the orthogonal design table L₉（3⁴）.The contents of the main index components of the samples were detected by high performance liquid phase（HPLC）,the methodology was investigated,and the preparation process was determined.Different proportions of medicated hepatocyte incubation solution（N1b-N13b）were prepared,and the main antiasthmatic components were determined by the same method as above,and the transformation status of the main active components of the compound was investigated.The growth curve of（RTE）of rat airway epithelial cells was measured by CCK-8 method.The effects of different concentrations of histamine phosphate（His）on the activity of RTE cells were detected.The effects of different concentrations of budesonide,medicated intestinal absorption solution and medicated liver cell incubation solution on the activity of RTE cells damaged by His were detected,to investigate the applicability of external pharmacodynamics of medicated liver cell incubation solution.4.Comparative analysis of medicated liver cell incubation solution and medicated serum of Maxingshigan decoction:the medicated liver cell incubation solution and medicated serum were compared from two aspects of the adaptability of pharmacodynamics in vitro and the relative content of main active components.5.Study on the potential action mechanism based on network pharmacology:the active components and corresponding targets of maxingshigan decoction were screened by（TCMSP）database of traditional Chinese medicine system pharmacology analysis platform,and the asthma related targets were searched by Gene Cards database and NCBI gene database,and the corresponding targets were intersected with maxingshigan decoction.The drug-active ingredient-action target-disease network and protein-protein interaction（PPI）network were constructed by Cytoscape3.7.2 software,and the GO function and KEGG pathway were analyzed by DAVID database.Results:1.The whole animal experiment showed that all the groups of Maxingshigan decoction tended to prolong the latent period of inducing asthma.Among them,the latent period of inducing asthma was significantly prolonged in N1 group,N2 group,N5group,N7 group,N8 group,N9 group,N10 group,N11 group,N12 group and N13 group（P<0.05 or P<0.01）.But the effect of N7 was the best（P<0.01）.The main differential components of asthma treated with water decoction are methyl ephedrine,ephedrine,pseudoephedrine,amygdalin,glycyrrhizin,glycyrrhizin,isoglycyrr hizin and ammonium glycyrrhizinate.2.The main difference components of medicated intestinal absorption solution of Maxingshigan decoction in the treatment of asthma are methyl ephedrine,ephedrine,pseudoephedrine,amygdalin,glycyrrhizin,glycyrrhizin,isoglycyrrhizin and ammonium glycyrrhizinate.Consistent with the water decoction.3.When the BRL-3A cell density was 1×10^-5cells/m L,the cell suspension:medicated intestinal absorption solution was 1:12,and the incubation time was 10min,the contents of the four index components were the highest,so this condition was selected as the preparation condition of medicated liver cell incubation solution.The main differential components of medicated liver cell incubation solution for treating asthma were ephedrine,ephedrine,pseudoephedrine,amygdalin,Liquiritin,glycyrrhizin and ammonium glycyrrhizinate.Compared with the medicated intestinal absorption solution containing drugs,there is a lack of isoglycyrrhizin,which may be involved in the metabolism of hepatocytes.RTE cells grow normally and can meet the needs of follow-up experiments.The His damage model of RTE cells was established with His concentration of 1.6×10^-2 mol·L^-1,budesonide concentration of 8×10^-4mol·L^-1 was used as the positive concentration of anti-His damage,and the intestinal absorption solution and liver cell incubation solution of maxingshigan decoction were both 10%as the concentration of anti-His damage.4.The inhibition rate of 10%medicated liver cell incubation solution of Maxingshigan decoction was the lowest（2%）,and that of 30%serum was the lowest（39%）.It can be seen that the effect of medicated hepatocyte incubation solution against His damage of RTE cells is better;the relative content of the main active components of Maxingshigan decoction is much higher than that of medicated serum.5.According to the network pharmacological search and screening,a total of 134active components of Maxingshigan decoction,237 corresponding targets,930 asthma related targets,81 intersection targets,187 items of GO and 89 KEGG pathways were obtained.Among them,the values of IL-6 and EGFR of the targets were higher,and the intersection targets were mainly concentrated in the signal pathways of PI3K-Akt,NF-κB and TNF.Conclusion:1.In this study,it was determined that there were differences and main differences in 13 different combinations of the three Medicated systems,and the three Medicated systems could reflect the migration and transformation of the main anti-asthma components of Maxingshigan decoction in vivo.2.In this study,the preparation technology of the new medicated liver cell incubation solution was determined.the medicated system has good applicability in vitro pharmacodynamics study,and can be used in the in vitro pharmacodynamics study of traditional Chinese medicine.3.Maxingshigan decoction has a definite antiasthmatic effect,and the network pharmacological analysis shows that it has the characteristics of multi-targets and multi-links,and can provide reference for the study of the action mechanism of the compound.