Background:To explore and predict the mechanism of classic ancient prescription of Chinese medicine Zhen-Wu decoction for chronic heart failure treatment based on network pharmacology.Methods:Traditional Chinese medicine systems pharmacology database was used to search the effective components and targets of herbs in classic ancient prescription of Chinese medicine Zhen-Wu decoction.Relevant target genes of chronic heart failure were obtained from GeneCards and Online Mendelian Inheritance in Man databases.Then we obtained the intersection target genes of classic ancient prescription of Chinese medicine Zhen-Wu decoction in treating chronic heart failure,constructing the classic ancient prescription of Chinese medicine Zhen-Wu decoction-active ingredient-chronic heart failure-targets network using Cytoscape,and performing network analysis using a network ananlyzer plug-in to acquire hub compounds and key targets.Proton pump inhibitor network was constructed through STRING database.Gene ontology function and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis were carried out in Database for Annotation,Visualization and Integrated Discovery.Results:There are 61 active ingredients and 134 action targets in classic ancient prescription of Chinese medicine Zhen-Wu decoction,among which 49 target genes and 11 important compounds related to the chronic heart failure treatment.After network analysis,we obtained 12 key targets,77 gene ontology entries and 22 signal pathways,including tumor necrosis factor signaling pathway,NF-kappaB signal pathway,PI3K-Akt signal pathway,et al.Concludion:classic ancient prescription of Chinese medicine Zhen-Wu decoction was used for chronic heart failure treatment for the multi-component,multi-target and multi-channel interaction.The components such as kaempferol and β-sitosterol were combined with target proteins such as CHRM1 and AChE,involving biological processes such as DNA transcription regulation,cholinergic synaptic transmission and apoptosis regulation,as well as signal pathways such as tumor necrosis factor signaling pathway,NF-kappaB signal pathway,PI3K-Akt signal pathway,et al.

Keyword:Zhen-Wu decoction,Chronic heart failure,Network pharmacology,Mechanism of action,Signal pathway

Background

Chronic heart failure (CHF) is a group of clinical syndromes with pulmonary circulation and/or systemic circulation congestion and hypoperfusion of organs and tissues.It is mainly manifested as dyspnea,limited activity tolerance and fluid retention,due to various cardiac structural or functional abnormalities leading to impaired ventricular filling and/or ejection capacity and cardiac output unable to meet the metabolic needs of the body tissue [1,2,3].Studies have shown that the pathological mechanism of CHF is mainly related to cardiac remodeling and neurohormonal mechanisms,specifically involving changes in cardiac systolic and diastolic functions,myocardial cell apoptosis or necrosis,sympathetic nervous system activation,renin-angiotensin system activation,insufficient neurohormonal secretion in peripheral blood vessels,inflammatory reactions,et al.[4,5].

The incidence rate,prevalence rate,rehospitalization rate and mortality rate of CHF are increasing year by year.The medical economic and social burden caused by CHF is worrying [6,7].Taking the electronic health records of 4 million people as research samples,the UK conducted a survey on the incidence and prevalence of heart failure from 2002 to 2014.The results showed that the absolute number of heart failure increased by 23% [8].An epidemiological study on heart failure in theEuropean Journal of Heart Failurein 2016 pointed out that the incidence rate and prevalence rate of heart failure among people over 60 years old rose sharply,reaching 11.8% [9].According to China cardiovascular disease report 2018[10],at present,the prevalence rate and mortality rate of cardiovascular diseases in our country are the first,and its mortality rate has exceeded that of tumors.It is estimated that the number of cardiovascular patients is 290 million,of which heart failure accounts for 4.5 million.

Traditional Chinese medicine (TCM) has certain advantages in the treatment of CHF.The history of heart failure research in TCM can be traced back to Huangdi Neijing (Inner Canon of Yellow Emperor,unknown author) from Warring States to Qin and Han Dynasties more than 2,000 years ago.According to the statistical analysis of “literature research on classic prescription for CHF” [11],classic ancient prescription of Chinese medicine Zhen-Wu decoction(CAPCMZWD) is the most frequently used classic prescription for CHF.CAPCMZWD,which consists of Fuling (Poria),Baishao (Paeoniae radix alba),Baizhu(Atractylodis macrocephalae rhizoma),Shengjiang(Zingiber officinale roscoe) and Fuzi (Aconiti lateralis radix praeparata) (Figure 1),has curative effect of“Warming Yang and Diuretic Treatment”,and is mainly used for “Yang-Deficiency”,and can be applied to CHF patients caused by “Yang Deficiency”and “Water Flooding”.In recent years,a number of studies have confirmed that CAPCMZWD can effectively treat CHF as well as reduce the adverse reactions of modern medicine treatments [12-14].However,the chemical composition of CAPCMZWD is complex,and its mechanism in CHF treatment has not been clearly explained.

Figure 1 The composition chart of Zhen-Wu decoction.Fuling,Poria; Baishao,Paeoniae radix alba; Baizhu,Atractylodis macrocephalae rhizoma;Shengjiang,Zingiber officinale roscoe; Fuzi,Aconiti lateralis radix praeparata.

With the gradual deepening of the world’s acceptance and recognition of TCM,the modernization of TCM is an inevitable trend in the research of TCM.How to expound the material basis and action mechanism of Chinese medicine in treating diseases from the perspective of modern pharmacology is the bottleneck and challenge faced by the modernization of Chinese medicine.In recent years,the concept of network pharmacology has gradually arisen and has been gradually applied to the research in the field of TCM.Network Pharmacology is based on systems biology,network biology,pharmacology and other disciplines.It uses public databases and documents to retrieve target information of known drug components and target information of diseases.It uses high-throughput proteomics and bioinformatics technologies to screen information.It uses visualization software to build a“gene-drug-target-disease” network diagram.Through the analysis and discussion of the network diagram,a research method to predict the pharmacodynamic active components,action targets,cell signal pathways and potential pharmacological mechanisms may be involved in regulation and control of diseases [15-17].

In this study,the CAPCMZWD is taken as the breakthrough point,and the mechanism of the treatment of CHF by CAPCMZWD is discussed and predicted based on the network pharmacology method,to provide ideas and basis for the research of the mechanism of the treatment of CHF by TCM.

Materials and methods

Major databases and software

The drug data in this study were extracted from the TCM systems pharmacology (TCMSP) database(http://tcmspw.com/tcmsp.php).The disease gene data were retrieved from GeneCards(https://www.genecards.org/),Online Mendelian Inheritance in Man (OMIM) (http://www.omim.org/),and UniProt (https://www.uniprot.org/).In addition,the key nodes excavated by STRING.11.0 database(https://string-db.org/) and Cytoscape3.7.2 software(https://cytoscape.org/) were visualized,and functional enrichment analysis of the Database for Annotation,Visualization and Integrated Discovery (DAVID) 6.8(https://david.ncifcrf.gov/),gene ontology (GO)(http://geneontology.org/),and Kyoto Encyclopedia of Genes and Genomes database (KEGG) database(https://www.kegg.jp/).Finally,Bioconductor(https://www.bioconductor.org/),Perl (https://www.perl.org/),R x64 3.6.0 (https://www.r-project.org/),and Java-jre (https://www.oracle.com/java/technologies/javase-jre8-downloads.html) were used as statistical tools for data analysis.

Collection and selecting of effective components in CAPCMZWD

The names of 5 TCMs “Fuling”,“Baishao”,“Baizhu”,“Shengjiang” and “Fuzi” in CAPCMZWD were respectively taken as key words,retrieved through TCMSP database.Relevant information of all herbs’chemical components was obtained under the entry of“ingredients”.According to pharmacokinetic absorption,distribution,metabolism,and excretion parameters,the effective chemical components of TCM were screened and stored in txt format with two conditions of simultaneously satisfying oral bioavailability > 30% and drug-like property > 0.18 as screening conditions.

Collection and selecting of action targets of CAPCMZWD

Searching through TCMSP database,taking “Fuling”,“Baishao”,“Baizhu”,“Shengjiang” and “Fuzi” as key words respectively,we obtained relevant action targets of all Chinese medicines.Then extract action targets matching with effective components of CAPCMZWD,namely “CAPCMZWD-effective components-action targets”,saving the data in txt format.The matching step was completed by Perl software running.The action targets corresponding to the effective components of CAPCMZWD were entered into UniProt database for retrieval,repetitive and irregular action targets were eliminated,and the standardized Gene Symbol of the action targets of the effective components of CAPCMZWD was obtained through retrieval and screening.

Selecting of target genes of CAPCMZWD for the treatment of CHF

The key word “chronic heart failure” was input into GeneCards database and OMIM of human Mendelian genetic comprehensive database to search,and repetitive genes and false positive genes were eliminated to obtain target genes related to CHF.The Bioconductor program package was run by software R x64 3.6.0 to map the target genes of CHF and the target genes of effective components of CAPCMZWD to obtain the intersection genes of drugs and diseases,that was,the target genes of CAPCMZWD for treating CHF.The data results were shown in venny diagram.

Network construction and analysis of CAPCMZWD in CHF treatment

Analysis on network construction of Cytoscape.The visualization tool Cytoscape3.7.2 software was used to draw the“CAPCMZWD-effective components-CHF-target” network.The Cytoscape3.7.2 software runs under Java-jre operating environment.The “network ananlyzer” plug-in in of Cytoscape3.7.2 software was used to analyze the network topology attributes to obtain the network topology parameters.The degree of freedom was used as the parameter reflecting the importance of nodes in the network,thus screening out important effective components and key targets.

Proton pump inhibitor network construction analysis.Target genes and proteins often interact with each other.Therefore,the target gene (gene symbol)for treating CHF with CAPCMZWD was imported into the protein interaction database STRING.11.0,the species was set as "Homo Sapiens",proton pump inhibitor (PPI) network between genes or proteins was obtained,and the network data was saved as tsv format file.The R language was used to analyze and count the number of edges connected by each node in PPI network.The more edges,the stronger the interaction between the target protein and other proteins.

Enrichment analysis of GO and KEGG.The target gene (gene symbol) of CAPCMZWD for CHF treatment was entered into DAVID 6.8 database,and the species was set as "Homo Sapiens" to obtain enrichment analysis data of GO function and KEGG signal pathway,wherein the enrichment analysis of GO function covered three aspects of biological process (BP),cellular component and molecular function.The software R x64 3.6.0 was used to run bioconductor's "clusterProfiler" program package.The GO function and KEGG signal pathway were described and annotated.At the same time,the GO function enrichment analysis bubble chart and KEGG signal path enrichment analysis histogram were drawn.Through the KEGG,the signal pathway map was queried.

Results

Effective components of CAPCMZWD

A total of 504 active ingredients of CAPCMZWD were retrieved and collected from TCMSP database,including 34 fromPoria,85 fromPaeoniae radix alba,55 fromAtractylodis macrocephalae rhizoma,265 fromZingiber officinale roscoeand 65 fromAconitilateralisradixpraeparata.Based on oral bioavailability > 30% and drug-like property > 0.18,61 of the active ingredients met the criteria,including 15 fromPoria,13 fromPaeoniae radix alba,7 fromAtractylodis macrocephalae rhizoma,5 fromZingiber officinale roscoeand 21 fromAconiti lateralis radix praeparata.Thus,there were 61 effective chemical components in CAPCMZWD (Table 1).

Table 1 The effective chemical components in Zhen-Wu decoction

Targets of CAPCMZWD

Four thousand two hundred and forty-nine targets of CAPCMZWD were retrieved from TCMSP database,including 121 fromPoria,990 fromPaeoniae radix alba,774 fromAtractylodis macrocephalae rhizoma,2,225 fromZingiber officinale roscoeand 139 fromAconiti lateralis radix praeparata.There were 134 targets corresponding to active ingredients in CAPCMZWD,including 15 fromPoria,61 fromPaeoniaeradixalba,11 fromAtractylodis macrocephalae rhizoma,32 fromZingiber officinale roscoeand 15 fromAconiti lateralis radix praeparata.

Target genes of CAPCMZWD in CHF treatment

Ten thousand three hundred and eleven CHF-related target genes were retrieved from GeneCards and OMIM databases,of which 9,861 were from GeneCards and 470 were from OMIM.After eliminating duplicate genes and false positive genes,a total of 10,223 CHF-related target genes were obtained,among which 1085 genes with relevance scoreda ≥ 20.There were 49 target genes for treating CHF with CAPCMZWD (Table 2).The intersection results of CAPCMZWD and CHF targets were shown in Figure 2.

Table 2 Target genes of Zhen-Wu decoction in treating chronic heart failure

Figure 2 Venny diagram of drug targets of Zhen-Wu decoction and chronic heart failure disease targets

Results of network construction and analysis

Analysis on network construction by Cytoscape.Cytoscape3.7.2 software was used to construct a“CAPCMZWD-active ingredient-CHF-target” network diagram of 49 common targets gene in drugs and diseases.The network included a total of 76 nodes and 372 edges (Figure 3).Through the analysis of the network topology attribute by the "network ananlyzer"plug-in in the Cytoscape3.7.2 software,it could be seen that there were 11 effective components with degree ≥ 4.They were kaempferol,beta-sitosterol,stigmasterol,hederagenin,3β-acetoxyatractylone,karanji,deltoin,8β-ethoxy atractylenolide Ⅲ,(+)-catechin,deoxyandrographolide,sitosterol respectively.These 11 effective components were used as important compounds of CAPCMZWD in the treatment of CHF.There were 12 targets with degree ≥4,namelyNCOA2,PGR,PTGS1,GABRA1,NR3C2,CHRM2,CHRM1,CHRM3,NCOA1,ESR1,AChEandADRA1A.These 12 targets were regarded as the key targets of CAPCMZWD for CHF treatment.Among the 49 targets,16 targets were linked to more than 3 compounds,which showsed that CAPCMZWD played a therapeutic mechanism through the joint action of multiple components and multiple targets in the treatment of CHF.

PPI network construction analysis.The 49 target genes of CAPCMZWD for CHF treatment were entered into STRING.11.0 database,and PPI network diagram is constructed.The network contained 47 target protein nodes and 430 edges (Figure 4).The R language analysis was used to count the number of edges connected by each node in PPI network.The top 10 target proteins in the number of edges were IL6,MAPK8,CASP3,ESR1,RELA,AR,AHR,CYP1A1,CYP3A4,PGR,which showed that the 10 target proteins had strong interaction with other proteins(Table 3).

Figure 3 “ZWD-active ingredient-CHF-target” network diagram.Blue nodes (1) represent ZWD formula; red nodes (1) represent CHF; yellow nodes (25) represent ZWD active ingredients; green nodes (49) represent ZWD targets for CHF treatment,and each edge represents the interaction between ZWD active ingredients and targets.ZWD,Zhen-Wu decoction (a classic ancient prescription of Chinese medicine); CHF,chronic heart failure.

Figure 4 PPI network diagram of ZWD in treating CHF.ZWD,Zhen-Wu decoction (a classic ancient prescription of Chinese medicine); CHF,chronic heart failure; PPI,proton pump inhibitor.

Table 3 Top 10 target proteins with the number of edges in proton pump inhibitor network

Enrichment analysis of GO and KEGG.Using DAVID 6.8 database,49 target genes of CAPCMZWD for CHF treatment were analyzed for GO function enrichment,and 77 GO entries were obtained,of which 47 were related to BP,mainly involving DNA-templated transcription,cholinergic synaptic transmission,cell adhesion,cellular response to hydrogen peroxide,metabolic process,regulation of apoptotic process,regulation of cell proliferation,regulation of vascular smooth muscle contraction,response to cytokine,cell growth and other aspects.There were 12 items related to cellular component,mainly involving nucleus,extracellular space,cell junction,cytosol,postsynaptic membrane,integral component of plasma membrane,perinuclear region of cytoplasm,synapse,nuclear membrane,nuclear chromatin,et al.There were 18 items related to molecular function,mainly involving chromatin binding,sequence-specific DNA binding,glutathione transferase activity,heme binding,iron ion binding,steroid binding,steroid hormone receptor activity,nuclear hormone receptor binding,scaffold protein binding,oxidoreductase activity,et al.The enrichment results of 20 related items of GO function enrichment analysis were plotted with R software (Figure 5).

Enrichment analysis of KEGG signal pathway:using DAVID 6.8 database,49 target genes of CAPCMZWD for CHF were enriched and analyzed by KEGG signal pathway,88 signal pathways were obtained,of which 22 were related to CHF,including tumor necrosis factor (TNF) signaling pathway,NF-kappaB (NF-κB) signal pathway,PI3K-Akt signal pathway,et al.The effective components of CAPCMZWD might act on these signaling pathways to achieve the purpose of treating CHF.Screen withP< 0.01 to obtain the first 20 pathways,and use R software to draw the enrichment analysis histogram of the first 20 signal channels (Figure 6).Among the 22 signal pathways related to CHF,the signal pathway diagrams of TNF signaling pathway,neuroactive ligand-receiver interaction,PI3K-Akt signaling pathway,which were the top 3 in the number sequence of enriched target genes,were retrieved through KEGG database.

Figure 5 GO function enrichment analysis bubble chart.Functional enrichment analysis of target gene GO for CHF treated by ZWD.Bubble size in bubble chart represents the number of genes occupied by this functional item,and color from cold to warm represents P value from large to small.GO,gene ontology; ZWD,Zhen-Wu decoction(a classic ancient prescription of Chinese medicine); CHF,chronic heart failure.

Figure 6 Column diagram for enrichment analysis of KEGG signal pathway.Enrichment analysis of KEGG signal pathway of target gene for CHF treated by ZWD.in the column chart,the X axis represents the number of genes occupied by signal pathway,the color from cold to warm represents the p value from large to small,and the Y axis represents the name of signal pathway.ZWD,Zhen-Wu decoction (a classic ancient prescription of Chinese medicine); CHF,chronic heart failure; KEGG,Kyoto Encyclopedia of Genes and Genomes database.

Discussion

In recent years,the study of mechanism of TCM compound by network pharmacology has gradually become a research hotspot.The mechanism of treating heart failure with TCM is related to regulating neuroendocrine system,inhibiting cardiac remodeling,inhibiting myocardial cell apoptosis,improving cardiac systolic and diastolic function,influencing humoral inflammatory factors,optimizing myocardial energy metabolism,et al.Based on the network pharmacology method,this study explores the mechanism of CAPCMZWD on CHF.

Network pharmacological analysis shows that the important active components related to the treatment of CAPCMZWD in CHF belong to flavonols,sterols,alcohols,pentacyclic triterpenes,diterpene lactones,phenols,et al.Flavonols can improve and regulate cardiovascular function.Kaempferol can significantly reduce the level of brain natriuretic peptide,and inhibit myocardial cell apoptosis by blocking phosphorylation of Akt/GSK-3β and p38 mitogen activated protein kinase/extracellular signal-regulated kinase signal pathways,thus playing a role in protecting the heart[18].Beta-sitosterol,stigmasterol and sitosterol are sterols,which can inhibit the expression of proinflammatory factors and regulate the apoptosis pathway.For example,β-sitosterol can down-regulate extracellular signal-regulated kinase 1/2 and up-regulate Bcl-2/Bax to inhibit myocardial cell apoptosis,and can also down-regulate the levels of interleukin (IL)-1β,IL-6 and TNF-α to improve inflammatory response [19].Both hederagenin and deltoin can inhibit the expression levels of NF-κB,TNF-α,IL-1β,IL-6 and other inflammatory factors [20,21].(+)-Catechin can inhibit immune activation,regulate the imbalance level of IL-17/IL-10,reverse the abnormal polarization of Th17h and Treg in peripheral blood and spleen,thus improving the cardiac function of congestive heart failure and playing a cardiac protective role [22].The roles of karanjin,3β-acetoxyatractylone,8β-ethoxy atractylenolide Ⅲand deoxyandrographolide in CHF need to be further explored.

There are 12 key targets for CHF treatment with CAPCMZWD,including NCOA2,PGR,PTGS1,GABRA1,NR3C2,CHRM2,CHRM1,CHRM3,NCOA1,ESR1,AChE,ADRA1A.Some studies have pointed out that prostaglandin signal activates JNK by up-regulating PTGER3,thus reducing the expression of GAD1 and GABRA1 in peripheral blood and promoting sympathetic excitation of CHF [23].NR3C2 genotype is pathophysiological related to ventricular remodeling in heart failure [24].The 3 subtypes of CHRM1,CHRM2 and CHRM3 belong to muscarinic acetylcholine receptors of G protein coupled receptor family.The parasympathetic nervous system controlled by acetylcholine plays a vital role in regulating cardiovascular function.After combining with acetylcholine,muscarinic receptor selectively acts on catecholamines and initiates multiple signal transduction pathways in cells,thus affecting the systolic and diastolic functions of the heart [25].The expression of AChE can affect the signal conduction in neuroendocrine system,thus regulating the levels of inflammatory factors such as TNF-α,IL-1β,NF-κB in heart tissue [26].ADRA1A belongs to α1 adrenoceptor,whose expression can reduce ventricular fibrosis and myocardial cell apoptosis [27].The regulatory roles of NCOA1,NCOA2,PGR,PTGS1,ESR1 in the occurrence and development of CHF still need to be further explored in order to provide new breakthroughs for future researchers.

There are 22 signaling pathways involved in the treatment of CHF with CAPCMZWD,including TNF signaling pathway,toll-like receptor signaling pathway,PI3K-Akt signaling pathway,cyclic adenosine monophosphate signaling pathway,calcium signaling pathway,neuroactive ligand-receptor interaction,aldosterone-regulated sodium reabsorption,et al.Studies have shown that the severity of heart failure is related to pro-inflammatory cytokines TNF-α,IL-6,et al.The lower the heart function,the higher the level of inflammatory factors [28].Therefore,inhibition of inflammatory reaction is one of the mechanisms that play a role in the treatment of CHF.TNF signaling pathway is closely related to NF-κB signaling pathway,PI3K-Akt signaling pathway and apoptosis,and mainly regulates apoptosis,cell survival,inflammation and immunity [28].The downstream cells of TNF-α in the TNF signaling pathway send signals through 2 homologous receptors TNFR1 and TNFR2.On the one hand,TNFR1 and TNFR2 act on JNK,P38,ERK,et al.to regulate the expression of various inflammatory factors such as IL1b,IL6,IL15,et al.and drive the pro-inflammatory program.On the other hand,they act on CASP3,CASP8,CASP10,et al.,affecting apoptosis and necrosis of cells and initiating tissue regeneration [29].Toll-like receptor signaling pathway can activate PI3K-Akt signaling pathway,NF-κB signaling pathway,mitogen activated protein kinase signaling pathway,et al.through a variety of downstream target transmembrane glycoprotein TLRs,and activate a variety of transcription factors such as NF-κB,AP-1,thus mediating inflammatory response in heart failure [30].

In the treatment of cardiovascular diseases,inhibition of myocardial cell death has extensive therapeutic potential [31].By inhibiting myocardial cell apoptosis,regulating cell growth and prolonging cell cycle,the process of myocardial remodeling is delayed,thus achieving the purpose of treating heart failure.FoxO signaling pathway plays an important role in regulating cell growth,cell cycle,apoptosis and resistance to oxidative stress [32].PI3K-Akt signaling pathway plays an important role in the pathophysiology of heart failure.It has been proved to be one of the most important proliferation promoting and anti-apoptosis signaling pathways controlling the survival and function of myocardial cells,and plays a key regulatory role in the process of myocardial cell survival and programmed death [33].Phosphatidylinositol triphosphate PI3K is an important cell signal transduction molecule,which generates phosphatidylinositol triphosphate PIP3 after activation of various extracellular signal stimuli.PIP3,as the second messenger in the cell,can bind and activate its downstream target serine/threonine protein kinase AKT,which is an important anti-apoptosis regulator.It can activate or inhibit downstream target proteins such as BAD,Caspase-9,NF-κB,FOXO,mTOR,et al.through phosphorylation,thus mediate a variety of growth factors to regulate cell growth and survival,and further regulate the process of myocardial cell proliferation,differentiation,survival,et al.[34-36].Activated AKT can also phosphorylate MDM2,up-regulate MDM2 activity,inactivate or degrade p53 function,and block p53-mediated apoptosis-promoting reaction [37].Some studies have shown that inhibiting PI3K-Akt signaling pathway can reduce myocardial hypertrophy and cardiac structural remodeling caused by pressure overload [38].Therefore,regulating PI3K-Akt signaling pathway can improve myocardial cell apoptosis,delay myocardial remodeling and improve myocardial function.

Lin et al.[39] observed in heart failure rats that inhibiting miR-665 could stabilize heart function of heart failure rats by up-regulating cAMP signaling pathway of GLP1R.CAMP is an important second messenger in cells.cAMP can act on its downstream target prekallikrein activator PKA to activate PKA and directly act on CREB and BAD to mediate cell death and survival.cAMP can also participate in calcium signaling pathway and regulate Ca2+concentration to regulate cardiac systolic and diastolic functions.The neuroactive ligand-receptor interaction pathway can enhance or inhibit neuronal activity by acting on targets such as NMUR,GHR,GNRHR and taking neuroactive steroid hormones as modulators of neurotransmitter receptors to maintain mitochondrial function during ischemia and perfusion to mediate cardiac protection [40].

Conclusion

To sum up,CAPCMZWD has predicted multi-component,multi-target and multi-pathway effects for CHF treatment.It could regulate neuroendocrine system,inhibite cardiac remodeling,inhibite myocardial cell apoptosis,improve cardiac systolic and diastolic function,and regulate inflammatory factor level.Chemical components such as kaempferol,β-sitosterol,stigmasterol and(+)-catechin in CAPCMZWD may play a role in CHF treatment by combining with key target proteins such as CHRM1,AChE and ADRA1A.these targets involved DNA transcription regulation,cholinergic synaptic transmission,apoptosis regulation,TNF signaling pathway,NF-κB signal pathway,neuroactive ligand-receptor interaction,PI3K-Akt signal pathway,calcium signal pathway,cAMP signal pathway,aldosterone-regulated sodium reabsorption and other signal pathways.This study is only a virtual prediction of the mechanism of CAPCMZWD in the treatment of CHF,and further experiments are needed to verify it.